US20030180733A1 - Specific and universal probes and amplification primers to rapidly detect and identify common bacterial pathogens and antibiotic resistance genes from clinical specimens for routine diagnosis in microbiology laboratories - Google Patents

Abstract

The present invention relates to DNA-based methods for universal bacterial detection, for specific detection of the common bacterial pathogensEscherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus mirabilis, Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Staphylococcus saprophyticus, Streptococcus pyogenes, Haemophilus influenzaeandMoraxella catarrhalisas well as for specific detection of commonly encountered and clinically relevant bacterial antibiotic resistance genes directly from clinical specimens or, alternatively, from a bacterial colony. The above bacterial species can account for as much as 80% of bacterial pathogens isolated in routine microbiology laboratories.

The core of this invention consists primarily of the DNA sequences from all species-specific genomic DNA fragments selected by hybridization from genomic libraries or, alternatively, selected from data banks as well as any oligonucleotide sequences derived from these sequences which can be used as probes or amplification primers for PCR or any other nucleic acid amplification methods. This invention also includes DNA sequences from the selected clinically relevant antibiotic resistance genes.

With these methods, bacteria can be detected (universal primers and/or probes) and identified (species-specific primers and/or probes) directly from the clinical specimens or from an isolated bacterial colony. Bacteria are further evaluated for their putative susceptibility to antibiotics by resistance gene detection (antibiotic resistance gene specific primers and/or probes). Diagnostic kits for the detection of the presence, for the bacterial identification of the above-mentioned bacterial species and for the detection of antibiotic resistance genes are also claimed. These kits for the rapid (one hour or less) and accurate diagnosis of bacterial infections and antibiotic resistance will gradually replace conventional methods currently used in clinical microbiology laboratories for routine diagnosis. They should provide tools to clinicians to help prescribe promptly optimal treatments when necessary. Consequently, these tests should contribute to saving human lives, rationalizing treatment, reducing the development of antibiotic resistance and avoid unnecessary hospitalizations.

Description

BACKGROUND OF THE INVENTION
• Classical Identification of Bacteria[0001]
• Bacteria are classically identified by their ability to utilize different substrates as a source of carbon and nitrogen through the use of biochemical tests such as the API20E™ system. Susceptibility testing of Gram negative bacilli has progressed to microdilution tests. Although the API and the microdilution systems are cost-effective, at least two days are required to obtain preliminary results due to the necessity of two successive overnight incubations to isolate and identify the bacteria from the specimen. Some faster detection methods with sophisticated and expensive apparatus have been developed. For example, the fastest identification system, the autoSCAN-Walk-Away system™ identifies both Gram negative and Gram positive from isolated bacterial colonies in 2 hours and susceptibility patterns to antibiotics in only 7 hours. However, this system has an unacceptable margin of error, especially with bacterial species other than[0002]Enterobacteriaceae(York et al., 1992. J. Clin. Microbiol. 30:2903-2910). Nevertheless, even this fastest method requires primary isolation of the bacteria as a pure culture, a process which takes at least 18 hours if there is a pure culture or 2 to 3 days if there is a mixed culture.
• Urine Specimens[0003]
• A large proportion (40-50%) of specimens received in routine diagnostic microbiology laboratories for bacterial identification are urine specimens (Pezzlo, 1988, Clin. Microbiol. Rev. 1:268-280). Urinary tract infections (UTI) are extremely common and affect up to 20% of women and account for extensive morbidity and increased mortality among hospitalized patients (Johnson and Stamm, 1989; Ann. Intern. Med. 111:906-917). UTI are usually of bacterial etiology and require antimicrobial therapy. The Gram negative bacillus[0004]Escherichia coliis by far the most prevalent urinary pathogen and accounts for 50 to 60% of UTI (Pezzlo, 1988, op. cit.). The prevalence for bacterial pathogens isolated from urine specimens observed recently at the “Centre Hospitalier de 1'Université Laval (CHUL)” is given in Tables 1 and 2.
• Conventional pathogen identification in urine specimens. The search for pathogens in urine specimens is so preponderant in the routine microbiology laboratory that a myriad of tests have been developed. The gold standard is still the classical semi-quantitative plate culture method in which a calibrated loop of urine is streaked on plates and incubated for 18-24 hours. Colonies are then counted to determine the total number of colony forming units (CFU) per liter of urine. A bacterial UTI is normally associated with a bacterial count of ≧10[0005]⁷CFU/L in urine. However, infections with less than 10⁷CFU/L in urine are possible, particularly in patients with a high incidence of diseases or those catheterized (Stark and Maki, 1984, N. Engl. J. Med. 311:560-564). Importantly, close to 80% of urine specimens tested are considered negative (<10⁷CFU/L; Table 3).
• Accurate and rapid urine screening methods for bacterial pathogens would allow a faster identification of negative results and a more efficient clinical investigation of the patient. Several rapid identification methods (Uriscreen™, UTIscreen™, Flash Track™ DNA probes and others) were recently compared to slower standard biochemical methods which are based on culture of the bacterial pathogens. Although much faster, these rapid tests showed low sensitivities and specificities as well as a high number of false negative and false positive results (Koening et al., 1992. J. Clin. Microbiol. 30:342-345; Pezzlo et al., 1992. J. Clin. Microbiol. 30:640-684).[0006]
• Urine specimens found positive by culture are further characterized using standard biochemical tests to identify the bacterial pathogen and are also tested for susceptibility to antibiotics.[0007]
• Any Clinical Specimens[0008]
• As with urine specimen which was used here as an example, our probes and amplification primers are also applicable to any other clinical specimens. The DNA-based tests proposed in this invention are superior to standard methods currently used for routine diagnosis in terms of rapidity and accuracy. While a high percentage of urine specimens are negative, in many other clinical specimens more than 95% of cultures are negative (Table 4). These data further support the use of universal probes to screen out the negative clinical specimens. Clinical specimens from organisms other than humans (e.g. other primates, mammals, farm animals or live stocks) may also be used.[0009]
• Towards the Development of Rapid DNA-Based Diagnostic Tests[0010]
• A rapid diagnostic test should have a significant impact on the management of infections. For the identification of pathogens and antibiotic resistance genes in clinical samples, DNA probe and DNA amplification technologies offer several advantages over conventional methods. There is no need for subculturing, hence the organism can be detected directly in clinical samples thereby reducing the costs and time associated with isolation of pathogens. DNA-based technologies have proven to be extremely useful for specific applications in the clinical microbiology laboratory. For example, kits for the detection of fastidious organisms based on the use of hybridization probes or DNA amplification for the direct detection of pathogens in clinical specimens are commercially available (Persing et al, 1993. Diagnostic Molecular Microbiology: Principles and Applications, American Society for Microbiology, Washington, D.C.).[0011]
• The present invention is an advantageous alternative to the conventional culture identification methods used in hospital clinical microbiology laboratories and in private clinics for routine diagnosis. Besides being much faster, DNA-based diagnostic tests are more accurate than standard biochemical tests presently used for diagnosis because the bacterial genotype (e.g. DNA level) is more stable than the bacterial phenotype (e.g. biochemical properties). The originality of this invention is that genomic DNA fragments (size of at least 100 base pairs) specific for 12 species of commonly encountered bacterial pathogens were selected from genomic libraries or from data banks. Amplification primers or oligonucleotide probes (both less than 100 nucleotides in length) which are both derived from the sequence of species-specific DNA fragments identified by hybridization from genomic libraries or from selected data bank sequences are used as a basis to develop diagnostic tests. Oligonucleotide primers and probes for the detection of commonly encountered and clinically important bacterial resistance genes are also included. For example, Annexes I and II present a list of suitable oligonucleotide probes and PCR primers which were all derived from the species-specific DNA fragments selected from genomic libraries or from data bank sequences. It is clear to the individual skilled in the art that oligonucleotide sequences appropriate for the specific detection of the above bacterial species other than those listed in Annexes 1 and 2 may be derived from the species-specific fragments or from the selected data bank sequences. For example, the oligonucleotides may be shorter or longer than the ones we have chosen and may be selected anywhere else in the identified species-specific sequences or selected data bank sequences. Alternatively, the oligonucleotides may be designed for use in amplification methods other than PCR. Consequently, the core of this invention is the identification of species-specific genomic DNA fragments from bacterial genomic DNA libraries and the selection of genomic DNA fragments from data bank sequences which are used as a source of species-specific and ubiquitous oligonucleotides. Although the selection of oligonucleotides suitable for diagnostic purposes from the sequence of the species-specific fragments or from the selected data bank sequences requires much effort it is quite possible for the individual skilled in the art to derive from our fragments or selected data bank sequences suitable oligonucleotides which are different from the ones we have selected and tested as examples (Annexes I and II).[0012]
• Others have developed DNA-based tests for the detection and identification of some of the bacterial pathogens for which we have identified species-specific sequences (PCT patent application Serial No. WO 93/03186). However, their strategy was based on the amplification of the highly conserved 16S rRNA gene followed by hybridization with internal species-specific oligonucleotides. The strategy from this invention is much simpler and more rapid because it allows the direct amplification of species-specific targets using oligonucleotides derived from the species-specific bacterial genomic DNA fragments.[0013]
• Since a high percentage of clinical specimens are negative, oligonucleotide primers and probes were selected from the highly conserved 16S or 23S rRNA genes to detect all bacterial pathogens possibly encountered in clinical specimens in order to determine whether a clinical specimen is infected or not. This strategy allows rapid screening out of the numerous negative clinical specimens submitted for bacteriological testing.[0014]
• We are also developing other DNA-based tests, to be performed simultaneously with bacterial identification, to determine rapidly the putative bacterial susceptibility to antibiotics by targeting commonly encountered and clinically relevant bacterial resistance genes. Although the sequences from the selected antibiotic resistance genes are available and have been used to develop DNA-based tests for their detection (Ehrlich and Greenberg, 1994. PCR-based Diagnostics in Infectious Diseases, Blackwell Scientific Publications, Boston, Mass.; Persing et al, 1993. Diagnostic Molecular Microbiology: Principles and Applications, American Society for Microbiology, Washington, D.C.), our approch is innovative as it represents major improvements over current “gold standard” diagnostic methods based on culture of the bacteria because it allows the rapid identification of the presence of a specific bacterial pathogen and evaluation of its susceptibility to antibiotics directly from the clinical specimens within one hour.[0015]
• We believe that the rapid and simple diagnostic tests not based on cultivation of the bacteria that we are developing will gradually replace the slow conventional bacterial identification methods presently used in hospital clinical microbiology laboratories and in private clinics. In our opinion, these rapid DNA-based diagnostic tests for severe and common bacterial pathogens and antibiotic resistance will (i) save lives by optimizing treatment, (ii) diminish antibiotic resistance by reducing the use of broad spectrum antibiotics and (iii) decrease overall health costs by preventing or shortening hospitalizations.[0016]
SUMMARY OF THE INVENTION
• In accordance with the present invention, there is provided sequence from genomic DNA fragments (size of at least 100 base pairs and all described in the sequence listing) selected either by hybridization from genomic libraries or from data banks and which are specific for the detection of commonly encountered bacterial pathogens (i.e.[0017]Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus mirabilis, Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Staphylococcus saprophyticus, Streptococcus pyogenes, Haemophilus influenzaeandMoraxella catarrhalis) in clinical specimens. These bacterial species are associated with approximately 90% of urinary tract infections and with a high percentage of other severe infections including septicemia, meningitis, pneumonia, intraabdominal infections, skin infections and many other severe respiratory tract infections. Overall, the above bacterial species may account for up to 80% of bacterial pathogens isolated in routine microbiology laboratories.
• Synthetic oligonucleotides for hybridization (probes) or DNA amplification (primers) were derived from the above species-specific DNA fragments (ranging in sizes from 0.25 to 5.0 kilobase pairs (kbp)) or from selected data bank sequences (GenBank and EMBL). Bacterial species for which some of the oligonucleotide probes and amplification primers were derived from selected data bank sequences are[0018]Escherichia coli, Enterococcus faecalis, Streptococcus pyogenesandPseudomonas aeruginosa.The person skilled in the art understands that the important innovation in this invention is the identification of the species-specific DNA fragments selected either from bacterial genomic libraries by hybridization or from data bank sequences. The selection of oligonucleotides from these fragments suitable for diagnostic purposes is also innovative. Specific and ubiquitous oligonucleotides different from the ones tested in the practice are considered as embodiments of the present invention.
• The development of hybridization (with either fragment or oligonucleotide probes) or of DNA amplification protocols for the detection of pathogens from clinical specimens renders possible a very rapid bacterial identification. This will greatly reduce the time currently required for the identification of pathogens in the clinical laboratory since these technologies can be applied for bacterial detection and identification directly from clinical specimens with minimum pretreatment of any biological specimens to release bacterial DNA. In addition to being 100% specific, probes and amplification primers allow identification of the bacterial species directly from clinical specimens or, alternatively, from an isolated colony. DNA amplification assays have the added advantages of being faster and more sensitive than hybridization assays, since they allow rapid and exponential in vitro replication of the target segment of DNA from the bacterial genome. Universal probes and amplification primers selected from the 16S or 23S rRNA genes highly conserved among bacteria, which permit the detection of any bacterial pathogens, will serve as a procedure to screen out the numerous negative clinical specimens received in diagnostic laboratories. The use of oligonucleotide probes or primers complementary to characterized bacterial genes encoding resistance to antibiotics to identify commonly encountered and clinically important resistance genes is also under the scope of this invention.[0019]
DETAILED DESCRIPTION OF THE INVENTION
• Development of Species-Specific DNA Probes[0020]
• DNA fragment probes were developed for the following bacterial species:[0021]Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus mirabilis, Streptococcus pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus saprophyticus, Haemophilus influenzaeandMoraxella catarrhalis.(ForEnterococcus faecalisand Streptococcus pyogenes, oligonucleotide sequences were exclusively derived from selected data bank sequences). These species-specific fragments were selected from bacterial genomic libraries by hybridization to DNA from a variety of Gram positive and Gram negative bacterial species (Table 5).
• The chromosomal DNA from each bacterial species for which probes were seeked was isolated using standard methods. DNA was digested with a frequently cutting restriction enzyme such as Sau3AI and then ligated into the bacterial plasmid vector pGEM3Zf (Promega) linearized by appropriate restriction endonuclease digestion. Recombinant plasmids were then used to transform competent[0022]E. colistrain DH5α thereby yielding a genomic library. The plasmid content of the transformed bacterial cells was analyzed using standard methods. DNA fragments of target bacteria ranging in size from 0.25 to 5.0 kilobase pairs (kbp) were cut out from the vector by digestion of the recombinant plasmid with various restriction endonucleases. The insert was separated from the vector by agarose gel electrophoresis and purified in low melting point agarose gels. Each of the purified fragments of bacterial genomic DNA was then used as a probe for specificity tests.
• For each given species, the gel-purified restriction fragments of unknown coding potential were labeled with the radioactive nucleotide α-[0023]³²P(dATP) which was incorporated into the DNA fragment by the random priming labeling reaction. Non-radioactive modified nucleotides could also be incorporated into the DNA by this method to serve as a label.
• Each DNA fragment probe (i.e. a segment of bacterial genomic DNA of at least 100 bp in length cut out from clones randomly selected from the genomic library) was then tested for its specificity by hybridization to DNAs from a variety of bacterial species (Table 5). The double-stranded labeled DNA probe was heat-denatured to yield labeled single-stranded DNA which could then hybridize to any single-stranded target DNA fixed onto a solid support or in solution. The target DNAs consisted of total cellular DNA from an array of bacterial species found in clinical samples (Table 5). Each target DNA was released from the bacterial cells and denatured by conventional methods and then irreversibly fixed onto a solid support (e.g. nylon or nitrocellulose membranes) or free in solution. The fixed single-stranded target DNAs were then hybridized with the single-stranded probe. Pre-hybridization, hybridization and post-hybridization conditions were as follows: (i) Pre-hybridization; in 1 M NaCl+10% dextran sulfate+1% SDS (sodium dodecyl sulfate)+1 μg/ml salmon sperm DNA at 650° C. for 15 min. (ii) Hybridization; in fresh pre-hybridization solution containing the labeled probe at 650° C. overnight. (iii) Post-hybridization; washes twice in 3×SSC containing 1% SDS (1×SSC is 0.15M NaCl, 0.015M NaCitrate) and twice in 0.1×SSC containing 0.1% SDS; all washes were at 650° C. for 15 min. Autoradiography of washed filters allowed the detection of selectively hybridized probes. Hybridization of the probe to a specific target DNA indicated a high degree of similarity between the nucleotide sequence of these two DNAs. Species-specific DNA fragments selected from various bacterial genomic libraries ranging in size from 0.25 to 5.0 kbp were isolated for 10 common bacterial pathogens (Table 6) based on hybridization to chromosomal DNAs from a variety of bacteria performed as described above. All of the bacterial species tested (66 species listed in Table 5) were likely to be pathogens associated with common infections or potential contaminants which can be isolated from clinical specimens. A DNA fragment probe was considered specific only when it hybridized solely to the pathogen from which it was isolated. DNA fragment probes found to be specific were subsequently tested for their ubiquity (i.e. ubiquitous probes recognized most isolates of the target species) by hybridization to bacterial DNAs from approximately 10 to 80 clinical isolates of the species of interest (Table 6). The DNAs were denatured, fixed onto nylon membranes and hybridized as described above.[0024]
• Sequencing of the Species-Specific Fragment Orobes[0025]
• The nucleotide sequence of the totality or of a portion of the species-specific DNA fragments isolated (Table 6) was determined using the dideoxynucleotide termination sequencing method which was performed using Sequenase (USB Biochemicals) or T7 DNA polymerase (Pharmacia). These nucleotide sequences are shown in the sequence listing. Alternatively, sequences selected from data banks (GenBank and EMBL) were used as sources of oligonucleotides for diagnostic purposes for[0026]Escherichia coli, Enterococcus faecalis, Streptococcus pyogenesandPseudomonas aeruginosa.For this strategy, an array of suitable oligonucleotide primers or probes derived from a variety of genomic DNA fragments (size of more than 100 bp) selected from data banks was tested for their specificity and ubiquity in PCR and hybridization assays as described later. It is important to note that the data bank sequences were selected based on their potential of being species-specific according to available sequence information. Only data bank sequences from which species-specific oligonucleotides could be derived are included in this invention.
• Oligonucleotide probes and amplification primers derived from species-specific fragments selected from the genomic libraries or from data bank sequences were synthesized using an automated DNA synthesizer (Millipore). Prior to synthesis, all oligonucleotides (probes for hybridization and primers for DNA amplification) were evaluated for their suitability for hybridization or DNA amplification by polymerase chain reaction (PCR) by computer analysis using standard programs (e.g. Genetics Computer Group (GCG) and Oligo™ 4.0 (National Biosciences)). The potential suitability of the PCR primer pairs was also evaluated prior to the synthesis by verifying the absence of unwanted features such as long stretches of one nucleotide, a high proportion of G or C residues at the 3′ end and a 3′-terminal T residue (Persing et al, 1993. Diagnostic Molecular Microbiology: Principles and Applications, American Society for Microbiology, Washington, D.C.).[0027]
• Hybridization with Oligonucleotide Probes[0028]
• In hybridization experiments, oligonucleotides (size less than 100 nucleotides) have some advantages over DNA fragment probes for the detection of bacteria such as ease of preparation in large quantities, consistency in results from batch to batch and chemical stability. Briefly, for the hybridizations, oligonucleotides were 5′ end-labeled with the radionucleotide γ[0029]³²P(ATP) using T4 polynucleotide kinase (Pharmacia). The unincorporated radionucleotide was removed by passing the labeled single-stranded oligonucleotide through a Sephadex G50 column. Alternatively, oligonucleotides were labeled with biotin, either enzymatically at their 3′ ends or incorporated directly during synthesis at their 5′ ends, or with digoxigenin. It will be appreciated by the person skilled in the art that labeling means other than the three above labels may be used.
• The target DNA was denatured, fixed onto a solid support and hybridized as previously described for the DNA fragment probes. Conditions for pre-hybridization and hybridization were as described earlier. Post-hybridization washing conditions were as follows: twice in 3×SSC containing 1% SDS, twice in 2×SSC containing 1% SDS and twice in 1×SSC containing 1% SDS (all of these washes were at 65° C. for 15 min ), and a final wash in 0.1×SSC containing 1% SDS at 25° C. for 15 min. For probes labeled with radioactive labels the detection of hybrids was by autoradiography as described earlier. For non-radioactive labels detection may be calorimetric or by chemiluminescence.[0030]
• The oligonucleotide probes may be derived from either strand of the duplex DNA. The probes may consist of the bases A, G, C, or T or analogs. The probes may be of any suitable length and may be selected anywhere within the species-specific genomic DNA fragments selected from the genomic libraries or from data bank sequences.[0031]
• DNA Amplification[0032]
• For DNA amplification by the widely used PCR (polymerase chain reaction) method, primer pairs were derived either from the sequenced species-specific DNA fragments or from data bank sequences or, alternatively, were shortened versions of oligonucleotide probes. Prior to synthesis, the potential primer pairs were analyzed by using the program oligo™ 4.0 (National Biosciences) to verify that they are likely candidates for PCR amplifications.[0033]
• During DNA amplification by PCR, two oligonucleotide primers binding respectively to each strand of the denatured double-stranded target DNA from the bacterial genome are used to amplify exponentially in vitro the target DNA by successive thermal cycles allowing denaturation of the DNA, annealing of the primers and synthesis of new targets at each cycle (Persing et al, 1993. Diagnostic Molecular Microbiology: Principles and Applications, American Society for Microbiology, Washington, D.C.). Briefly, the PCR protocols were as follows. Clinical specimens or bacterial colonies were added directly to the 50 μL PCR reaction mixtures containing 50 mM KCl, 10 mM Tris-HCl pH 8.3, 2.5 mM MgCl[0034]_2,0.4 μM of each of the two primers, 200 μM of each of the four dNTPs and 1.25 Units of Taq DNA polymerase (Perkin Elmer). PCR reactions were then subjected to thermal cycling (3 min at 95° C. followed by 30 cycles of 1 second at 95° C. and 1 second at 55° C.) using a Perkin Elmer 480™ thermal cycler and subsequently analyzed by standard ethidium bromide-stained agarose gel electrophoresis. It is clear that other methods for the detection of specific amplification products, which may be faster and more practical for routine diagnosis, may be used. Such methods may be based on the detection of fluorescence after amplification (e.g. TaqMan™ system from Perkin Elmer or Amplisensor™ from Biotronics) or liquid hybridization with an oligonucleotide probe binding to internal sequences of the specific amplification product. These novel probes can be generated from our species-specific fragment probes. Methods based on the detection of fluorescence are particularly promising for utilization in routine diagnosis as they are, very rapid and quantitative and can be automated.
• To assure PCR efficiency, glycerol or dimethyl sulfoxide (DMSO) or other related solvents, can be used to increase the sensitivity of the PCR and to overcome problems associated with the amplification of target with a high GC content or with strong secondary structures. The concentration ranges for glycerol and DMSO are 5-15% (v/v) and 3-10% (v\v), respectively. For the PCR reaction mixture, the concentration ranges for the amplification primers and the MgCl[0035]₂are 0.1-1.0 μM and 1.5-3.5 mM, respectively. Modifications of the standard PCR protocol using external and nested primers (i.e. nested PCR) or using more than one primer pair (i.e. multiplex PCR) may also be used (Persing et al, 1993. Diagnostic Molecular Microbiology: Principles and Applications, American Society for Microbiology, Washington, D.C.). For more details about the PCR protocols and amplicon detection methods see examples 7 and 8.
• The person skilled in the art of DNA amplification knows the existence of other rapid amplification procedures such as ligase chain reaction (LCR), transcription-based amplification systems (TAS), self-sustained sequence replication (3SR), nucleic acid sequence-based amplification (NASBA), strand displacement amplification (SDA) and branched DNA (bDNA) (Persing et al, 1993. Diagnostic Molecular Microbiology: Principles and Applications, American Society for Microbiology, Washington, D.C.). The scope of this invention is not limited to the use of amplification by PCR, but rather includes the use of any rapid nucleic acid amplification methods or any other procedures which may be used to increase rapidity and sensitivity of the tests. Any oligonucleotides suitable for the amplification of nucleic acid by approaches other than PCR and derived from the species-specific fragments and from selected antibiotic resistance gene sequences included in this document are also under the scope of this invention.[0036]
• Specificity and Ubiquity Tests for Oligonucleotide Probes and Primers[0037]
• The specificity of oligonucleotide probes, derived either from the sequenced species-specific fragments or from data bank sequences, was tested by hybridization to DNAs from the array of bacterial species listed in Table 5 as previously described. Oligonucleotides found to be specific were subsequently tested for their ubiquity by hybridization to bacterial DNAs from approximately 80 isolates of the target species as described for fragment probes. Probes were considered ubiquitous when they hybridized specifically with the DNA from at least 80% of the isolates. Results for specificity and ubiquity tests with the oligonucleotide probes are summarized in Table 6. The specificity and ubiquity of the amplification primer pairs were tested directly from cultures (see example 7) of the same bacterial strains. For specificity and ubiquity tests, PCR assays were performed directly from bacterial colonies of approximately 80 isolates of the target species. Results are summarized in Table 7. All specific and ubiquitous oligonucleotide probes and amplification primers for each of the 12 bacterial species investigated are listed in Annexes I and II, respectively. Divergence in the sequenced DNA fragments can occur and, insofar as the divergence of these sequences or a part thereof does not affect the specificity of the probes or amplification primers, variant bacterial DNA is under the scope of this invention.[0038]
• Universal Bacterial Detection[0039]
• In the routine microbiology laboratory a high percentage of clinical specimens sent for bacterial identification is negative (Table 4). For example, over a 2 year period, around 80% of urine specimens received by the laboratory at the “Centre Hospitalier de 1' Université Laval (CHUL)” were negative (i.e. <10[0040]⁷CFU/L) (Table 3). Testing clinical samples with universal probes or universal amplification primers to detect the presence of bacteria prior to specific identification and screen out the numerous negative specimens is thus useful as it saves costs and may rapidly orient the clinical management of the patients. Several oligonucleotides and amplification primers were therefore synthesized from highly conserved portions of bacterial 16S or 23S ribosomal RNA gene sequences available in data banks (Annexes III and IV). In hybridization tests, a pool of seven oligonucleotides (Annex I; Table 6) hybridized strongly to DNA from all bacterial species listed in Table 5. This pool of universal probes labeled with radionucleotides or with any other modified nucleotides is consequently very useful for detection of bacteria in urine samples with a sensitivity range of ≧10⁷CFU/L. These probes can also be applied for bacterial detection in other clinical samples.
• Amplification primers also derived from the sequence of highly conserved ribosomal RNA genes were used as an alternative strategy for universal bacterial detection directly from clinical specimens (Annex IV; Table 7). The DNA amplification strategy was developed to increase the sensitivity and the rapidity of the test. This amplification test was ubiquitous since it specifically amplified DNA from 23 different bacterial species encountered in clinical specimens.[0041]
• Well-conserved bacterial genes other than ribosomal RNA genes could also be good candidates for universal bacterial detection directly from clinical specimens. Such genes may be associated with processes essential for bacterial survival (e.g. protein synthesis, DNA synthesis, cell division or DNA repair) and could therefore be highly conserved during evolution. We are working on these candidate genes to develop new rapid tests for the universal detection of bacteria directly from clinical specimens.[0042]
• Antibiotic Resistance Genes[0043]
• Antimicrobial resistance complicates treatment and often leads to therapeutic failures. Furthermore, overuse of antibiotics inevitably leads to the emergence of bacterial resistance. Our goal is to provide the clinicians, within one hour, the needed information to prescribe optimal treatments. Besides the rapid identification of negative clinical specimens with DNA-based tests for universal bacterial detection and the identification of the presence of a specific pathogen in the positive specimens with DNA-based tests for specific bacterial detection, the clinicians also need timely information about the ability of the bacterial pathogen to resist antibiotic treatments. We feel that the most efficient strategy to evaluate rapidly bacterial resistance to antimicrobials is to detect directly from the clinical specimens the most common and important antibiotic resistance genes (i.e. DNA-based tests for the detection of antibiotic resitance genes). Since the sequence from the most important and common bacterial antibiotic resistance genes are available from data banks, our strategy is to use the sequence from a portion or from the entire gene to design specific oligonucleotides which will be used as a basis for the development of rapid DNA-based tests. The sequence from the bacterial antibiotic resistance genes selected on the basis of their clinical relevance (i.e. high incidence and importance) is given in the sequence listing. Table 8 summarizes some characteristics of the selected antibiotic resistance genes.[0044]
EXAMPLES
• The following examples are intended to be illustrative of the various methods and compounds of the invention.[0045]
Example 1
• Isolation and cloning of fragments. Genomic DNAs from[0046]Escherichia colistrain ATCC 25922,Klebsiella pneumoniaestrain CK2,Pseudomonas aeruginosastrain ATCC 27853,Proteus mirabilisstrain ATCC 35657,Streptococcus pneumoniaestrain ATCC 27336,Staphylococcus aureusstrain ATCC 25923,Staphylococcus epidermidisstrain ATCC 12228,Staphylococcus saprophyticusstrain ATCC 15305,Haemophilus influenzaereference strain Rd andMoraxella catarrhalisstrain ATCC 53879 were prepared using standard procedures. It is understood that the bacterial genomic DNA may have been isolated from strains other than the ones mentioned above. (ForEnterococcus faecalisandStreptococcus pyogenesoligonucleotide sequences were derived exclusively from data banks). Each DNA was digested with a restriction enzyme which frequently cuts DNA such as Sau3AI. The resulting DNA fragments were ligated into a plasmid vector (pGEM3Zf) to create recombinant plasmids and transformed into competentE. colicells (DH5α). It is understood that the vectors and corresponding competent cells should not be limited to the ones herein above specifically examplified. The objective of obtaining recombinant plasmids and transformed cells is to provide an easily reproducible source of DNA fragments useful as probes. Therefore, insofar as the inserted fragments are specific and selective for the target bacterial DNA, any recombinant plasmids and corresponding transformed host cells are under the scope of this invention. The plasmid content of the transformed bacterial cells was analyzed using standard methods. DNA fragments from target bacteria ranging in size from 0.25 to 5.0 kbp were cut out from the vector by digestion of the recombinant plasmid with various restriction endonucleases. The insert was separated from the vector by agarose gel electrophoresis and purified in a low melting point agarose gel. Each of the purified fragments was then used for specificity tests.
• Labeling of DNA fragment probes. The label used was α[0047]³²P(dATP), a radioactive nucleotide which can be incorporated enzymatically into a double-stranded DNA molecule. The fragment of interest is first denatured by heating at 95° C. for 5 min, then a mixture of random primers is allowed to anneal to the strands of the fragments. These primers, once annealed, provide a starting point for synthesis of DNA. DNA polymerase, usually the Klenow fragment, is provided along with the four nucleotides, one of which is radioactive. When the reaction is terminated, the mixture of new DNA molecules is once again denatured to provide radioactive single-stranded DNA molecules (i.e. the probe). As mentioned earlier, other modified nucleotides may be used to label the probes.
• Specificity and ubiquity tests for the DNA fragment probes. Species-specific DNA fragments ranging in size from 0.25 to 5.0 kbp were isolated for 10 common bacterial pathogens (Table 6) based on hybridization to chromosomal DNAs from a variety of bacteria. Samples of whole cell DNA for each bacterial strain listed in Table 5 were transferred onto a nylon membrane using a dot blot apparatus, washed and denatured before being irreversibly fixed. Hybridization conditions were as described earlier. A DNA fragment probe was considered specific only when it hybridized solely to the pathogen from which it was isolated. Labeled DNA fragments hybridizing specifically only to target bacterial species (i.e. specific) were then tested for their ubiquity by hybridization to DNAs from approximately 10 to 80 isolates of the species of interest as described earlier. The conditions for pre-hybridization, hybridization and post-hybridization washes were as described earlier. After autoradiography (or other detection means appropriate for the non-radioactive label used), the specificity of each individual probe can be determined. Each probe found to be specific (i.e. hybridizing only to the DNA from the bacterial species from which it was isolated) and ubiquitous (i.e. hybridizing to most isolates of the target species) was kept for further experimentations.[0048]
Example 2
• Same as example 1 except that testing of the strains is by colony hybridization. The bacterial strains were inoculated onto a nylon membrane placed on nutrient agar. The membranes were incubated at 37° C. for two hours and then bacterial lysis and DNA denaturation were carried out according to standard procedures. DNA hybridization was performed as described earlier.[0049]
Example 3
• Same as example 1 except that bacteria were detected directly from clinical samples. Any biological samples were loaded directly onto a dot blot apparatus and cells were lysed in situ for bacterial detection. Blood samples should be heparizined in order to avoid coagulation interfering with their convenient loading on a dot blot apparatus.[0050]
Example 4
• Nucleotide sequencina of DNA fragments. The nucleotide sequence of the totality or a portion of each fragment found to be specific and ubiquitous (Example 1) was determined using the dideoxynucleotide termination sequencing method (Sanger et al., 1977, Proc. Natl. Acad. Sci. USA. 74:5463-5467). These DNA sequences are shown in the sequence listing. Oligonucleotide probes and amplification primers were selected from these nucleotide sequences, or alternatively, from selected data banks sequences and were then synthesized on an automated Biosearch synthesizer (Millipore™) using phosphoramidite chemistry.[0051]
• Labeling of oliaonucleotides. Each oligonucleotide was 5′ end-labeled with γ[0052]³²P-ATP by the T4 polynucleotide kinase (Pharmacia) as described earlier. The label could also be non-radioactive.
• Specificity test for oligonucleotide probes. All labeled oligonucleotide probes were tested for their specificity by hybridization to DNAs from a variety of Gram positive and Gram negative bacterial species as described earlier. Species-specific probes were those hybridizing only to DNA from the bacterial species from which it was isolated. Oligonucleotide probes found to be specific were submitted to ubiquity tests as follows.[0053]
• Ubiquity test for oligonucleotide probes. Specific oligonucleotide probes were then used in ubiquity tests with approximately 80 strains of the target species. Chromosomal DNAs from the isolates were transferred onto nylon membranes and hybridized with labeled oligonucleotide probes as described for specificity tests. The batteries of approximately 80 isolates constructed for each target species contain reference ATCC strains as well as a variety of clinical isolates obtained from various sources. Ubiquitous probes were those hybridizing to at least 80% of DNAs from the battery of clinical isolates of the target species. Examples of specific and ubiquitous oligonucleotide probes are listed in Annex 1.[0054]
Example 5
• Same as example 4 except that a pool of specific oligonucleotide probes is used for bacterial identification (i) to increase sensitivity and assure 100% ubiquity or (ii) to identify simultaneously more than one bacterial species. Bacterial identification could be done from isolated colonies or directly from clinical specimens.[0055]
Example 6
• PCR amplification. The technique of PCR was used to increase sensitivity and rapidity of the tests. The PCR primers used were often shorter derivatives of the extensive sets of oligonucleotides previously developed for hybridization assays (Table 6). The sets of primers were tested in PCR assays performed directly from a bacterial colony or from a bacterial suspension (see Example 7) to determine their specificity and ubiquity (Table 7). Examples of specific and ubiquitous PCR primer pairs are listed in annex II.[0056]
• Specificity and ubiquity tests for amplification primers. The specificity of all selected PCR primer pairs was tested against the battery of Gram negative and Gram positive bacteria used to test the oligonucleotide probes (Table 5). Primer pairs found specific for each species were then tested for their ubiquity to ensure that each set of primers could amplify at least 80% of DNAs from a battery of approximately 80 isolates of the target species. The batteries of isolates constructed for each species contain reference ATCC strains and various clinical isolates representative of the clinical diversity for each species.[0057]
• Standard precautions to avoid false positive PCR results should be taken. Methods to inactivate PCR amplification products such as the inactivation by uracil-N-glycosylase may be used to control PCR carryover.[0058]
Example 7
• Amplification directly from a bacterial colony or suspension. PCR assays were performed either directly from a bacterial colony or from a bacterial suspension, the latter being adjusted to a standard McFarland 0.5 (corresponds to 1.5 ×10[0059]⁸bacteria/mL). In the case of direct amplification from a colony, a portion of the colony was transferred directly to a 50 μL PCR reaction mixture (containing 50 mM KCl, 10 mM Tris pH 8.3, 2.5 mM MgCl_2,0.4 μM of each of the two primers, 200 μM of each of the four dNTPs and 1.25 Unit of Taq DNA polymerase (Perkin Elmer)) using a plastic rod. For the bacterial suspension, 4 μL of the cell suspension was added to 46 μL of the same PCR reaction mixture. For both strategies, the reaction mixture was overlaid with 50 μL of mineral oil and PCR amplifications were carried out using an initial denaturation step of 3 min. at 95° C. followed by 30 cycles consisting of a 1 second denaturation step at 95° C. and of a 1 second annealing step at 55° C. in a Perkin Elmer 480™ thermal cycler. PCR amplification products were then analyzed by standard agarose gel (2%) electrophoresis. Amplification products were visualized in agarose gels containing 2.5 μg/mL of ethidium bromide under UV at 254 nm. The entire PCR assay can be completed in approximately one hour.
• Alternatively, amplification from bacterial cultures was performed as described above but using a “hot start” protocol. In that case, an initial reaction mixture containing the target DNA, primers and dNTPs was heated at 85° C. prior to the addition of the other components of the PCR reaction mixture. The final concentration of all reagents was as described above. Subsequently, the PCR reactions were submitted to thermal cycling and analysis as described above.[0060]
Example 8
• Amplification directly from clinical specimens. For amplification from urine specimens, 4 μL of undiluted or diluted (1:10) urine was added directly to 46 μL of the above PCR reaction mixture and amplified as described earlier.[0061]
• To improve bacterial cell lysis and eliminate the PCR inhibitory effects of clinical specimens, samples were routinely diluted in lysis buffer containing detergent(s). Subsequently, the lysate was added directly to the PCR reaction mixture. Heat treatments of the lysates, prior to DNA amplification, using the thermocycler or a microwave oven could also be performed to increase the efficiency of cell lysis.[0062]
• Our strategy is to develop rapid and simple protocols to eliminate PCR inhibitory effects of clinical specimens and lyse bacterial cells to perform DNA amplification directly from a variety of biological samples. PCR has the advantage of being compatible with crude DNA preparations. For example, blood, cerebrospinal fluid and sera may be used directly in PCR assays after a brief heat treatment. We intend to use such rapid and simple strategies to develop fast protocols for DNA amplification from a variety of clinical specimens.[0063]
Example 9
• Detection of antibiotic resistance genes. The presence of specific antibiotic resistance genes which are frequently encountered and clinically relevant is identified using the PCR amplification or hybridization protocols described in previous sections. Specific oligonucleotides used as a basis for the DNA-based tests are selected from the antibiotic resistance gene sequences. These tests can be performed either directly from clinical specimens or from a bacterial colony and should complement diagnostic tests for specific bacterial identification.[0064]
Example 10
• Same as examples 7 and 8 except that assays were performed by multiplex PCR (i.e. using several pairs of primers in a single PCR reaction) to (i) reach an ubiquity of 100% for the specific target pathogen or (ii) to detect simultaneously several species of bacterial pathogens.[0065]
• For example, the detection of Escherichia coli requires three pairs of PCR primers to assure a ubiquity of 100%. Therefore, a multiplex PCR assay (using the “hot-start” protocol (Example 7)) with those three primer pairs was developed. This strategy was also used for the other bacterial pathogens for which more than one primer pair was required to reach an ubiquity of 100%.[0066]
• Multiplex PCR assays could also be used to (i) detect simultaneously several bacterial species or, alternatively, (ii) to simultaneously identify the bacterial pathogen and detect specific antibiotic resistance genes either directly from a clinical specimen or from a bacterial colony.[0067]
• For these applications, amplicon detection methods should be adapted to differentiate the various amplicons produced. Standard agarose gel electrophoresis could be used because it discriminates the amplicons based on their sizes. Another useful strategy for this purpose would be detection using a variety of fluorochromes emitting at different wavelengths which are each coupled with a specific oligonucleotide linked to a fluorescence quencher which is degraded during amplification to release the fluorochrome (e.g. TaqMan™, Perkin Elmer).[0068]
Example 11
• Detection of amplification Products. The person skilled in the art will appreciate that alternatives other than standard agarose gel electrophoresis (Example 7) may be used for the revelation of amplification products. Such methods may be based on the detection of fluorescence after amplification (e.g. Amplisensor™, Biotronics; TaqMan™) or other labels such as biotin (SHARP Signal™ system, Digene Diagnostics). These methods are quantitative and easily automated. One of the amplification primers or an internal oligonucleotide probe specific to the amplicon(s) derived from the species-specific fragment probes is coupled with the fluorochrome or with any other label. Methods based on the detection of fluorescence are particularly suitable for diagnostic tests since they are rapid and flexible as fluorochromes emitting different wavelengths are available (Perkin Elmer).[0069]
Example 12
• Species-specific, universal and antibiotic resistance gene amplification primers can be used in other rapid amplification procedures such as the ligase chain reaction (LCR), transcription-based amplification systems (TAS), self-sustained sequence replication (3SR), nucleic acid sequence-based amplification (NASBA), strand displacement amplification (SDA) and branched DNA (bDNA) or any other methods to increase the sensitivity of the test. Amplifications can be performed from an isolated bacterial colony or directly from clinical specimens. The scope of this invention is therefore not limited to the use of PCR but rather includes the use of any procedures to specifically identify bacterial DNA and which may be used to increase rapidity and sensitivity of the tests.[0070]
Example 13
• A test kit would contain sets of probes specific for each bacterium as well as a set of universal probes. The kit is provided in the form of test components, consisting of the set of universal probes labeled with non-radioactive labels as well as labeled specific probes for the detection of each bacterium of interest in specific clinical samples. The kit will also include test reagents necessary to perform the pre-hybridization, hybridization, washing steps and hybrid detection. Finally, test components for the detection of known antibiotic resistance genes (or derivatives therefrom) will be included. Of course, the kit will include standard samples to be used as negative and positive controls for each hybridization test.[0071]
• Components to be included in the kits will be adapted to each specimen type and to detect pathogens commonly encountered in that type of specimen. Reagents for the universal detection of bacteria will also be included. Based on the sites of infection, the following kits for the specific detection of pathogens may be developed:[0072]
• A kit for the universal detection of bacterial pathogens from most clinical specimens which contains sets of probes specific for highly conserved regions of the bacterial genomes.[0073]
• A kit for the detection of bacterial pathogens retrieved from urine samples, which contains eight specific test components (sets of probes for the detection of[0074]Escherichia coli, Enterococcus faecalis, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa, Staphylococcus saprophyticus, Staphylococcus aureusandStaphylococcus epidermidis).
• A kit for the detection of respiratory pathogens which contains seven specific test components (sets of probes for detecting[0075]Streptococcus pneumoniae, Moraxella catarrhalis, Haemophilus influenzae, Klebsiella pneumoniae, Pseudomonas aeruginosa, Streptococcus pyogenesandStaphylococcus aureus).
• A kit for the detection of pathogens retrieved from blood samples, which contains eleven specific test components (sets of probes for the detection of[0076]Streptococcus pneumoniae, Moraxella catarrhalis, Haemophilus influenzae, Proteus mirabilis, Klebsiella pneumoniae, Pseudomonas aeruginosa, Escherichia coli, Enterococcus faecalis, Staphylococcus aureus, Streptococcus pyogenesandStaphylococcus epidermidis).
• A kit for the detection of pathogens causing meningitis, which contains four specific test components (sets of probes for the detection of[0077]Haemophilus influenzae, Streptococcus pneumoniae, Escherichia coliandPseudomonas aeruginosa).
• A kit for the detection of clinically important antibiotic resistance genes which contains sets of probes for the specific detection of at least one of the 19 following genes associated with bacterial resistance : bla[0078]_tem, bla_rob, bla_shv, aadB, aacC1, aacC2, aacC3, aacA4, mecA, vanA, vanH, vanX, satA, aacA-aphD, vat, vga, msrA, sul and int.
• Other kits adapted for the detection of pathogens from skin, abdominal wound or any other clinically relevant kits will be developed.[0079]
Example 14
• Same as example 13 except that the test kits contain all reagents and controls to perform DNA amplification assays. Diagnostic kits will be adapted for amplification by PCR (or other amplification methods) performed directly either from clinical specimens or from a bacterial colony. Components required for universal bacterial detection, bacterial identification and antibiotic resistance genes detection will be included.[0080]
• Amplification assays could be performed either in tubes or in microtitration plates having multiple wells. For assays in plates, the wells will be coated with the specific amplification primers and control DNAs and the detection of amplification products will be automated. Reagents and amplification primers for universal bacterial detection will be included in kits for tests performed directly from clinical specimens. Components required for bacterial identification and antibiotic resistance gene detection will be included in kits for testing directly from colonies as well as in kits for testing directly from clinical specimens.[0081]
• The kits will be adapted for use with each type of specimen as described in example 13 for hybridization-based diagnostic kits.[0082]
Example 15
• It is understood that the use of the probes and amplification primers described in this invention for bacterial detection and identification is not limited to clinical microbiology applications. In fact, we feel that other sectors could also benefit from these new technologies. For example, these tests could be used by industries for quality control of food, water, pharmaceutical products or other products requiring microbiological control. These tests could also be applied to detect and identify bacteria in biological samples from organisms other than humans (e.g. other primates, mammals, farm animals and live stocks). These diagnostic tools could also be very useful for research purposes including clinical trials and epidemiological studies.[0083]

  TABLE 1
  Distribution of urinary isolates from positive urine samples
  (≧107 CFU/L) at the Centre Hospitalier
  de l′Université Laval (CHUL) for the
  1992-1994 period.

  % of isolates

  	Nov 92	April 93	July 93	Jan 94
  Organisms	n = 267a	n = 265	n = 238	n = 281

  Escherichia coli	53.2	51.7	53.8	54.1
  Enterococcus faecalis	13.8	12.4	11.7	11.4
  Klebsiella pneumoniae	6.4	6.4	5.5	5.3
  Staphylococcus epidermidis	7.1	7.9	3.0	6.4
  Proteus mirabilis	2.6	3.4	3.8	2.5
  Pseudomonas aeruginosa	3.7	3.0	5.0	2.9
  Staphylococcus saprophyticus	3.0	1.9	5.4	1.4
  Othersb	10.2	13.3	11.8	16.0

• [0084]

  TABLE 2
  Distribution of uncommonaurinary isolates from positive urine
  samples ( ≧107CFU/L) at the Centre Hospitalier de
  I′Université Laval (CHUL) for the 1992-1994 period.

  % of isolates

  Organismsa	Nov 92	April 93	July 93	Jan 94

  Staphylococcus aureus	0.4	1.1	1.3	1.4
  Staphylococcus spp.	2.2	4.9	1.7	6.0
  Micrococcus spp.	0.0	0.0	0.4	0.7
  Enterococcus faecium	0.4	0.4	1.3	1.4
  Citrobacter spp.	1.4	0.8	0.4	0.7
  Enterobacter spp.	1.5	1.1	1.3	1.4
  Klebsiella oxytoca	1.1	1.5	2.5	1.8
  Serratia spp.	0.8	0.0	0.5	0.0
  Proteus spp.	0.4	0.4	0.0	1.1
  Morganella and Providencia	0.4	0.8	0.4	0.0
  Hafnia alvei	0.8	0.0	0.0	0.0
  NFBb(Stenotrophomonas,	0.0	0.4	1.3	1.1
  Acinetobacter)
  Candida spp.	0.8	1.9	0.7	0.4

• [0085]

  TABLE 3
  Distribution of positivea(bacterial count ≧107CFU/L)
  and negative (bacterial count <107CFU/L) urine specimens tested
  at the Centre Hospitalier de l′Université Laval (CHUL) for
  the 1992-1994 period.

  Number of isolates (%)

  Specimens	Nov 92	April 93	July 93	Jan 94
  received:	1383(100)	1338(100)	1139(100)	1345(100)
  positive:	267(19.3)	265(19.8)	238(20.9)	281(20.9)
  negative:	1116(80.7)	1073(80.2)	901(79.1)	1064(79.1)

• [0086]

  TABLE 4
  Distribution of positive and negative clinical specimens tested
  in the Microbiology Laboratory of the CHUL.

  	No. of	% of	% of
  Clinical	samples	positive	negative
  specimensa	tested	specimens	specimens

  Urine	17,981	19.4	80.6
  Haemoculture/marrow	10,010	6.9	93.1
  Sputum	1,266	68.4	31.6
  Superficial pus	1,136	72.3	27.7
  Cerebrospinal fluid	553	1.0	99.0
  Synovial fluid-articular	523	2.7	97.3
  Bronch./Trach./Amyg./Throat	502	56.6	43.4
  Deep pus	473	56.8	43.2
  Ears	289	47.1	52.9
  Pleural and pericardial fluid	132	1.0	99.0
  Peritonial fluid	101	28.6	71.4

• [0087]

  TABLE 5
  Bacterial species (66) used for testing the specificity of DNA fragment
  probes, oligonucleotide probes and PCR primers.

  	Number of		Number of
  Bacterial species	strains	Bacterial species	strains
  Gram negative:	tested	Gram positive:	tested
  Proteus mirabilis	5	Streptococcus pneumoniae	7
  Klebsiella pneumoniae	5	Streptococcus salivarius	2
  Pseudomonas aeruginosa	5	Streptococcus viridans	2
  Escherichia coli	5	Streptococcus pyogenes	2
  Moraxella catarrhalis	5	Staphylococcus aureus	2
  Proteus vulgaris	2	Staphylococcus epidermidis	2
  Morganella morganii	2	Staphylococcus saprophyticus	5
  Enterobacter cloacae	2	Micrococcus species	2
  Providencia stuartii	1	Corynebacterium species	2
  Providencia species	1	Streptococcusgroupe B	2
  Enterobacter agglomerans	2	Staphylococcus simulans	2
  Providencia rettgeri	2	Staphylococcus ludgunensis	1
  Neisseria mucosa	1	Staphylococcus capitis	2
  Providencia alcalifaciens	1	Staphylococcus haemolyticus	2
  Providencia rustigianii	1	Staphylococcus hominis	2
  Burkholderia cepacia	2	Enterococcus faecalis	2
  Enterobacter aerogenes	2	Enterococcus faecium	1
  Stenotrophomonas maltophilia	2	Staphylococcus warneri	1
  Pseudomonas fluorescens	1	Enterococcus durans	1
  Comamonas acidovorans	2	Streptococcus bovis	1
  Pseudomonas putida	2	Diphteroids	2
  Haemophilus influenzae	5	Lactobacillus acidophilus	1
  Haemophilus parainfluenzae	2
  Bordetella pertussis	2
  Haemophilus parahaemolyticus	2
  Haemophilus haemolyticus	2
  Haemophilus aegyptius	1
  Kingella indologenes	1
  Moraxella atlantae	1
  Neisseria caviae	1
  Neisseria subflava	1
  Moraxella urethralis	1
  Shigella sonnei	1
  Shigella flexneri	1
  Klebsiella oxytoca	2
  Serratia marcescens	2
  Salmonella typhimurium	1
  Yersinia enterocolitica	1
  Acinetobacter calcoaceticus	1
  Acinetobacter lwoffi	1
  Haftnia alvei	2
  Citrobacter diversus	1
  Citrobacter freundii	1
  Salmonella species	1

• [0088]

  TABLE 6
  Species-specific DNA fragment and oligonucleotide probes for hybridization.

  Number of fragment probesb

  Number of oligonucleotide probes

  Organismsa	Tested	Specific	Ubiquitousc	Synthesized	Specific	Ubiquitousc
  E. colid	—	—	—	20	12	9f
  E. coli	14	2	2e	—	—	—
  K. pneumoniaed	—	—	—	15	1	1
  K. pneumoniae	33	3	3	18	12	8
  P. mirabilisd	—	—	—	3	3	2
  P. mirabilis	14	3	3e	15	8	7
  P. aeruginosad	—	—	—	26	13	9
  P. aeruginosa	6	2	2e	6	0	0
  S. saprophyticus	7	4	4	20	9	7
  H. influenzaed	—	—	—	16	2	2
  H. influenzae	1	1	1	20	1	1
  S. pneumoniaed	—	—	—	6	1	1
  S. pneumoniae	19	2	2	4	1	1
  M. catarrhalis	2	2	2	9	8	8
  S. epidermidis	62	1	1	—	—	—
  S. aureus	30	1	1	—	—	—
  Universal probesd	—	—	—	7	—	7g

• [0089]

  TABLE 7
  PCR amplification for bacterial pathogens commonly encountered in
  urine, sputum, blood, cerebrospinal fluid and other specimens.

  Primer paira

  Amplicon

  DNA amplification

  Organism	# (SEQ ID NO)	size (bp)	Ubiquityb	from coloniesc	from specimensd
  E. coli	le(55-56)	107	75/80	+	+
  	2e(46-47)	297	77/80	+	+
  	3 (42-43)	102	78/80	+	+
  	4 (131-132)	134	73/80	+	+
  	1 + 3 + 4	—	80/80	+	+
  E.faecalis	1e(38-39)	200	71/80	+	+
  	2e(40-41)	121	79/80	+	+
  	1 + 2	—	80/80	+	+
  K. pneumoniae	1 (67-68)	198	76/80	+	+
  	2 (61-62)	143	67/80	+	+
  	3h(135-136)	148	78/80	+	N.T.i
  	4 (137-138)	116	69/80	+	N.T.
  	1 + 2 + 3	—	80/80	+	N.T.
  P. mirabilis	1 (74-75)	167	73/80	+	N.T.
  	2 (133-134)	123	80/80	+	N.T.
  P. aeruginosa	1e(83-84)	139	79/80	+	N.T.
  	2e(85-86)	223	80/80	+	N.T.
  S. saprophyticus	1 (98-99)	126	79/80	+	+
  	2 (139-140)	190	80/80	+	N.T.
  M. catarrhalis	1 (112-113)	157	79/80	+	N.T.
  	2 (118-119)	118	80/80	+	N.T.
  	3 (160-119)	137	80/80	+	N.T.
  H. influenzae	1e(154-155)	217	80/80	+	N.T.
  S. pneumoniae	1e(156-157)	134	80/80	+	N.T.
  	2e(158-159)	197	74/80	+	N.T.
  	3 (78-79)	175	67/80	+	N.T.
  S. epidermidis	1 (147-148)	175	80/80	+	N.T.
  	2 (145-146)	125	80/80	+	N.T.
  S. aureus	1 (152-153)	108	80/80	+	N.T.
  	2 (149-150)	151	80/80	+	N.T.
  	3 (149-151)	176	80/80	+	N.T.
  S. pyogenesf	1e(141-142)	213	80/80	+	N.T.
  	2e(143-144)	157	24/24	+	N.T.
  Universal	1e(126-127)	241	194/195g	+	+

• a All primer pairs are specific in PCR assays since no amplification was observed with DNA from 66 different species of both Gram positive and Gram negative bacteria other than the species of interest (Table 5).[0090]
• b The ubiquity was normally tested on 80 strains of the species of interest. All retained primer pairs amplified at least 90% of the isolates. When combinations of primers were used, an ubiquity of 100% was reached.[0091]
• c For all primer pairs and multiplex combinations, PCR amplifications directly performed from a bacterial colony were 100% species-specific.[0092]
• d PCR assays performed directly from urine specimens.[0093]
• e Primer pairs derived from data bank sequences. Primer pairs with no “e” are derived from our species-specific fragments.[0094]
• f For S. pyogenes, primer pair #1 is specific for Group A Streptococci (GAS). Primer pair #2 is specific for the GAS-producing exotoxin A gene (SpeA).[0095]
• g Ubiquity tested on 195 isolates from 23 species representative of bacterial pathogens commonly encountered in clinical specimens.[0096]
• h Optimizations are in progress to eliminate non-specific amplification observed with some bacterial species other than the target species.[0097]
• N.T.: not tested.[0098]

  TABLE 8
  Selected antibiotic resistance genes for diagnostic purposes.

  Genes	Antibiotics	Bacteriaa	SEQ ID NO
  (blatem) TEM-1	β-lactams	Enterobacteriaceae,	161
  		Pseudomonadaceae,
  		Haemophilus, Neisseria
  (blarob) ROB-1	β-lactams	Haemophilus, Pasteurella	162
  (blashv) SHV-1	β-lactams	Klebsiella and other	163
  		Enterobacteriaceae
  aadB, aacC1, aacC2,	Aminoglycosides	Enterobacteriaceae,	164, 165, 166
  aacC3, aacA4		Pseudomonadaceae	167, 168
  mecA	β-lactams	Staphylococci	169
  vanH, vanA, vanX	Vancomycin	Enterococci	170
  satA	Macrolides	Enterococci	173
  aacA-aphD	Aminoglycosides	Enterococci, Staphylococci	174
  vat	Macrolides	Staphylococci	175
  vga	Macrolides	Staphylococci	176
  msrA	Erythromycin	Staphylococci	177
  Int and Sul	β-lactams, trimethoprim,	Enterobacteriaceae,	171, 172
  conserved sequences	aminoglycosides, antiseptic,	Pseudomonadaceae
  	chloramphenicol

• [0099]

  Annex I: Specific and ubiquitous oligonucleotides probes
  for hybridization

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Bacterial species:Escherichia coli

  44	5′-CAC CCG CTT GCG TGG CAA GCT GCC C	5a	213-237
  45	5′-CGT TTG TGG ATT CCA GTT CCA TCC G	5a	489-513
  48	5′-TGA AGC ACT GGC CGA AAT GCT GCG T	6a	759-783
  49	5′-GAT GTA CAG GAT TCG TTG AAG GCT T	6a	898-922
  50	5′-TAG CGA AGG CGT AGC AGA AAC TAA C	7a	1264-1288
  51	5′-GCA ACC CGA ACT CAA CGC CGG ATT T	7a	1227-1251
  52	5′-ATA CAC AAG GGT CGC ATC TGC GGC C	7a	1313-1337
  53	5′-TGC GTA TGC ATT GCA GAC CTT GTG GC	7a	111-136
  54	5′-GCT TTC ACT GGA TAT CGC GCT TGG G	7a	373-397

  Bacterial species:Proteus mirabilis

  70b	5′-TGG TTC ACT GAC TTT GCG ATG TTT C	12	23-47
  71	5′-TCG AGG ATG GCA TGC ACT AGA AAA T	12	53-77
  72b	5′-CGC TGA TTA GGT TTC GCT AAA ATC TTA TTA	12	80-109
  73	5′-TTG ATC CTC ATT TTA TTA ATC ACA TGA CCA	12	174-203

• [0100]

  Annex I: Specific and ubiquitous oligonucleotides probes
  for hybridization

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Bacterial species:Proteus mirabilis

  76	5′-CCG CCT TTA GCA TTA ATT GGT GTT TAT AGT	13	246-275
  77	5′-CCT ATT GCA GAT ACC TTA AAT GTC TTG GGC	13	291-320
  80b	5′-TTG AGT GAT GAT TTC ACT GAC TCC C	14	18-42
  81	5′-GTG AGA CAG TGA TGG TGA GGA CAC A	15a	1185-1209
  82	5′-TGG TTG TCA TGC TGT TTG TGT GAA AAT	15a	1224-1230

  Bacterial species:Klebsiella pneumoniae

  57	5′-GTG GTG TCG TTC AGG GGT TTC AC	8	45-67
  58	5′-GCG ATA TTC ACA CCC TAC GCA GCC A	9	161-185
  59b	5′-GTC GAA AAT GCC GGA AGA GGT ATA CG	9	203-228
  60b	5′-ACT GAG CTG CAG ACC GGT AAA ACT CA	9	233-258
  63b	5′-CGT GAT GGA TAT TCT TAA CGA AGG GC	10	250-275
  64b	5′-ACC AAA CTG TTG AGC CGC CTG GA	10	201-223
  65	5′-GTG ATC GCC CCT CAT CTG CTA CT	10	77-99
  66	5′-CGC CCT TCG TTA AGA ATA TCC ATC AC	10	249-274
  69	5′-CAG GAA GAT GCT GCA CCG GTT GTT G	11a	296-320

• [0101]

  Annex I: Specific and ubiquitous oligonucleotides probes
  for hybridization

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Bacterial species:Pseudomonas aeruginosa

  87	5′-AAT GCG GCT GTA CCT CGG CGC TGG T	18a	2985-3009
  88	5′-GGC GGA GGG CCA GTT GCA CCT GCC A	18a	2929-2953
  89	5′-AGC CCT GCT CCT CGG CAG CCT CTG C	18a	2821-2845
  90	5′-TGG CTT TTG CAA CCG CGT TCA GGT T	18a	1079-1103
  91	5′-GCG CCC GCG AGG GCA TGC TTC GAT G	19a	705-729
  92	5′-ACC TGG GCG CCA ACT ACA AGT TCT A	19a	668-692
  93	5′-GGC TAC GCT GCC GGG CTG CAG GCC G	19a	505-529
  94	5′-CCG ATC TAG ACC ATC GAG ATG GGC G	20a	1211-1235
  95	5′-GAG CGC GGC TAT GTG TTC GTC GGC T	20a	2111-2135

  Bacterial species:Streptococcus pneumoniae

  120	5′-TCT GTG CTA GAG ACT GCC CCA TTT C	30	423-447
  121	5′-CGA TGT CTT GAT TGA GCA GGG TTA T	31a	1198-1222

• [0102]

  Annex I: Specific and ubiquitous oligonucleotides probes
  for hybridization

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Bacterial species:Staphylococcus saprophyticus

  96	5′-CGT TTT TAC CCT TAC CTT TTC GTA CTA CC	21	45-73
  97b	5′-TCA GGC AGA GGT AGT ACG AAA AGG TAA GGG	21	53-82
  100	5′-CAC CAA GTT TGA CAC GTG AAG ATT CAT	22	89-115
  101b	5′-ATG AGT GAA GCG GAG TCA GAT TAT GTG CAG	23	105-134
  102	5′-CGC TCA TTA CGT ACA GTG ACA ATC G	24	20-44
  103	5′-CTG GTT AGC TTG ACT CTT AAC AAT CTT GTC	24	61-90
  104b	5′-GAC GCG ATT GTC ACT GTA CGT AAT GAG CGA	24	19-48

  Bacterial species:Moraxella catarrhalis

  108	5′-GCC CCA AAA CAA TGA AAC ATA TGG T-3′	28	81-105
  109	5′-CTG CAG ATT TTG GAA TCA TAT CGC C-3′	28	126-130
  110	5′-TGG TTT GAC CAG TAT TTA ACG CCA T-3′	28	165-189
  111	5′-CAA CGG CAC CTG ATG TAC CTT GTA C-3′	28	232-256
  114	5′-TTA CAA CCT GCA CCA CAA GTC ATC A-3′	29	97-121
  115	5′-GTA CAA ACA AGC CGT CAG CGA CTT A-3′	29	139-163
  116	5′-CAA TCT GCG TGT GTG CGT TCA CT-3′	29	178-200
  117	5′-GCT ACT TTG TCA GCT TTA GCC ATT CA-3′	29	287-312

• [0103]

  Annex I:
  Specific and ubiquitous oligonucleotides probes
  for hybridization

  Originating DNA fragment

  		SEQ ID	Nucleotide
  SEQ ID NO	Nucleotide sequence	NO	position
  Bacterial species:	Haemophilus influenzae
  105b	5′-GCG TCA GAA AAA GTA GGC GAA ATG AAA G	25	138-165
  106b	5′-AGC GGC TCT ATC TTG TAA TGA CAC A	26a	770-794
  107b	5′-GAA ACG TGA ACT CCC CTC TAT ATA A	27a	5184-5208
  	Universal probesc
  122b	5′-ATC CCA CCT TAG GCG GCT GGC TCC A	—	—
  123	5′-ACG TCA AGT CAT CAT GGC CCT TAC GAG TAG G	—	—
  124b	5′-GTG TGA CGG GCG GTG TGT ACA AGG C	—	—
  125b	5′-GAG TTG CAG ACT CCA ATC CGG ACT ACG A	—	—
  128b	5′-CCC TAT ACA TCA CCT TGC GGT TTA GCA GAG AG	—	—
  129	5′-GGG GGG ACC ATC CTC CAA GGC TAA ATA C	—	—
  130b	5′-CGT CCA CTT TCG TGT TTG CAG AGT GCT GTG TT	—	—

• [0104]

  Annex II: Specific and ubiquitous primers for DNA amplification

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Bacterial species:Escherichia coli

  42	5′-GCT TTC CAG CGT CAT ATT G	4	177-195
  43b	5′-GAT CTC GAC AAA ATG GTG A	4	260-278
  46	5′-TCA CCC GCT TGC GTG GC	5a	212-228
  47b	5′-GGA ACT GGA ATC CAC AAA C	5a	490-508
  55	5′-GCA ACC CGA ACT CAA CGC C	7a	1227-1245
  56b	5′-GCA GAT GCG ACC CTT GTG T	7a	1315-1333
  131	5′-CAG GAG TAC GGT GAT TTT TA	3	60-79
  132b	5′-ATT TCT GGT TTG GTC ATA CA	3	174-193

  Bacterial species:Enterococcus faecalis

  38	5′-GCA ATA CAG GGA AAA ATG TC	1a	69-88
  39b	5′-CTT CAT CAA ACA ATT AAC TC	1a	249-268
  40	5′-GAA CAG AAG AAG CCA AAA AA	2a	569-588
  41b	5′-GCA ATC CCA AAT AAT ACG GT	2a	670-689

• [0105]

  Annex II: Specific and ubiquitous primers for DNA amplification

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Bacterial species:Klebsiella pneumoniae

  61	5′-GAC AGT CAG TTC GTC AGC C	9	37-55
  62b	5′-CGT AGG GTG TGA ATA TCG C	9	161-179
  67	5′-TCG CCC CTC ATC TGC TAC T	10	81-99
  68b	5′-GAT CGT GAT GGA TAT TCT T	10	260-278
  135	5′-GCA GCG TGG TGT CGT TCA	8	40-57
  136b	5′-AGC TGG CAA CGG CTG GTC	8	170-187
  137	5′-ATT CAC ACC CTA CGC AGC CA	9	166-185
  138b	5′-ATC CGG CAG CAT CTC TTT GT	9	262-281

  Bacterial species:Proteus mirabilis

  74	5′-GAA ACA TCG CAA AGT CAG T	12	23-41
  75b	5′-ATA AAA TGA GGA TCA AGT TC	12	170-189
  133	5′-CGG GAG TCA GTG AAA TCA TC	14	17-36
  134b	5′-CTA AAA TCG CCA CAC CTC TT	14	120-139

• [0106]

  Annex II: Specific and ubiquitous primers for DNA amplification

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Bacterial species:Staphylococcus saprophyticus

  98	5′-CGT TTT TAC CCT TAC CTT TTC GTA CT	21	45-70
  99b	5′-ATC GAT CAT CAC ATT CCA TTT GTT TTT A	21	143-170
  139	5′-CTG GTT AGC TTG ACT CTT AAC AAT C	24	61-85
  140b	5′-TCT TAA CGA TAG AAT GGA GCA ACT G	24	226-250

  Bacterial species:Pseudomonas aeruginosa

  83	5′-CGA GCG GGT GGT GTT CAT C	16a	554-572
  84b	5′-CAA GTC GTG GTG GGA GGG A	16a	674-692
  85	5′-TCG CTG TTC ATC AAG ACC C	17a	1423-1441
  86b	5′-CCG AGA ACC AGA CTT CAT C	17a	1627-1645

  Bacterial species:Moraxella catarrhalis

  112	5′-GGC ACC TGA TGT ACC TTG	28	235-252
  113b	5′-AAC AGC TCA CAC GCA TT	28	375-391
  118	5′-TGT TTT GAG CTT TTT ATT TTT TGA	29	41-64
  119b	5′-CGC TGA CGG CTT GTT TGT ACC A	29	137-158
  160	5′-GCT CAA ATC AGG GTC AGC	29	22-39
  119b	5′-CGC TGA CGG CTT GTT TGT ACG A	29	137-158

• [0107]

  Annex II: Specific and ubiquitous primers for DNA amplification

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Bacterial species: Staphylococcus epidermidis

  145	5′-ATC AAA AAG TTG GCG AAC CTT TTC A	36	21-45
  146b	5′-CAA AAG AGC GTG GAG AAA AGT ATC A	36	121-145
  147	5′-TCT CTT TTA ATT TCA TCT TCA ATT CCA TAG	36	448-477
  148b	5′-AAA CAC AAT TAC AGT CTG GTT ATC CAT ATC	36	593-622

  Bacterial species: Staphylococcus aureus

  149b	5′-CTT CAT TTT ACG GTG ACT TCT TAG AAG ATT	37	409-438
  150	5′-TCA ACT GTA GCT TCT TTA TCC ATA CGT TGA	37	288-317
  149b	5′-CTT CAT TTT ACG GTG ACT TCT TAG AAG ATT	37	409-438
  151	5′-ATA TTT TAG CTT TTC AGT TTC TAT ATC AAC	37	263-292
  152	5′-AAT CTT TGT CGG TAC ACG ATA TTC TTC ACG	37	5-34
  153b	5′-CGT AAT GAG ATT TCA GTA GAT AAT ACA ACA	37	83-112

• [0108]

  Annex II: Specific and ubiquitous primers for DNA amplification

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Bacterial species:Haemophilus influenzae

  154	5′-TTT AAC GAT CCT TTT ACT CCT TTT G	27a	5074-5098
  155b	5′-ACT GCT GTT GTA AAG AGG TTA AAA T	27a	5266-5290

  Bacterial species:Streptococcus pneumoniae

  78	5′-AGT AAA ATG AAA TAA GAA CAG GAC AG	34	164-189
  79b	5′-AAA ACA GGA TAG GAG AAC GGG AAA A	34	314-338
  156	5′-ATT TGG TGA CGG GTG ACT TT	31a	1401-1420
  157b	5′-GCT GAG GAT TTG TTC TTC TT	31a	1515-1534
  158	5′-GAG CGG TTT CTA TGA TTG TA	35a	1342-1361
  159b	5′-ATC TTT CCT TTC TTG TTC TT	35a	1519-1538

  Bacterial species:Streptococcus pyogenes

  141	5′-TGA AAA TTC TTG TAA CAG GC	32a	286-305
  142b	5′-GGC CAC CAG CTT GCC CAA TA	32a	479-498
  143	5′-ATA TTT TCT TTA TGA GGG TG	33a	966-985
  144b	5′-ATC CTT AAA TAA AGT TGC CA	33a	1103-1122

• [0109]

  Annex II: Specific and ubiquitous primers for DNA amplification

  Originating DNA fragment

  SEQ ID NO	Nucleotide Sequence	SEQ ID NO	Nucleotide position

  Universal primersc

  126	5′-GGA GGA AGG TGG GGA TGA CG	—	—
  127b	5′-ATG GTG TGA CGG GCG GTG TG	—	—

• [0110]

  Annex III
  Selection of Universal Probes by Alignment of the
  Sequences of Bacterial 16S and 23S Ribosomal RNA Genes.

  Reverse strand of SEQ ID NO: 122	TGGAGCC AGCCGCCTA GGTGGGAT

  	1461      1510
  Streptococcus salivarius	TGAGGTAACC TTTTGGAGCC AGCCGCCTAA GGTGGGATAG ATGANNGGGG
  Proteus vulgaris	TAGCTTAACC TTCGGGAGGG CGCTTACCAC TTTGTGATTC ATGACTGGGG
  Pseudomonas aeruginosa	TAGTCTAACC GCAAGGGGGA CGGTTACCAC GGAGTGATTC ATGACTGGGG
  Neisseria gonorrhoeae	TAGGGTAACC GCAAGGAGTC CGCTTACCAC GGTATGCTTC ATGACTGGGG
  Streptococcus lactis	TTGCCTAACC GCAAGGAGGG CGCTTCCTAA GGTAAGACCG ATGACNNGGG

• [0111]

  Annex III. Selection of universal probes by alignment of the
  sequences of bacterial 165 and 23S ribosomal RNA genes.

  SEQ ID NO: 123	ACGTCAAGTC ATCATGGC CCTTACGAGT AGG
  	1251                                              1300
  Haemophilus influenzae	GGTNGGGATG ACGTCAAGTC ..ATCATGGC CCTTACGAGT AGGGCTACAC
  Neisseria gonorrhoeae	GGTGGGGATG ACGTCAAGTC ..CTCATGGC CCTTATGACC AGGGCTTCAC
  Pseudomonas cepacia	GGTNGGGATG ACGTCAAGTC ..CTCATGGC CCTTATGGGT AGGGCTTCAC
  Serratia marcescens	GGTGGGGATG ACGTCAAGTC ..ATCATGGC CCTTACGAGT AGGGCTACAC
  Escherichia coli	GGTGGGGATG ACGTCAAGTC ..ATCATGGC CCTTACGACC AGGGCTACAC
  Proteus vulgaris	GGTGGGGATG ACGTTAAGTC GTATCATGGC CCTTACGAGT AGGGCTACAC
  Pseudomonas aeruginosa	GGTGGGGATG ACGTCAAGTC ..ATCATGGC CCTTACGGCN AGGGCTACAC
  Clostridium perfringens	GGTGGGGATG ACGTNNAATC ..ATCATGCC CNTTATGTGT AGGGCTACAC
  Mycoplasma hominis	GGTGGGGATG ACGTCAAATC ..ATCATGCC TCTTACGAGT GGGGCCACAC
  Helicobacter pylori	GGTGGGGACG ACGTCAAGTC ..ATCATGGC CCTTACGCCT AGGGCTACAC
  Mycoplasma pneumoniae	GGAAGGGATG ACGTCAAATC ..ATCATGCC CCTTATGTCT AGGGCTGCAA

• [0112]

  Annex III. Selection of universal probes by alignment of the
  sequences of bacterial 16S and 23S ribosomal RNA genes.

  Reverse of the probe SEQ ID NO: 124	GCCTTGTACA CACCGCCCGT CACAC
  	1451                                   1490
  Escherichia coli	ACGTTCCCGG GCCTTGTACA CACCGCCCGT CACACCATGG
  Neisseria gonorrhoeae	ACGTTCCCNG NNCTTGTACA CACCGCCCGT CACACCATGG
  Pseudomonas cepacia	ACGTTCCCGG GTCTTGTACA CACNGCCCGT CACACCATGG
  Serratia marcescens	ACGTTCCCGG GCCTTGTACA CACCGCCCGT CACACCATGG
  Proteus vulgaris	ACGTTCCCGG GCCTTGTACA CACCGCCCGT CACACCATGG
  Haemophilus influenzae	ACGTTCCCGG GCNTTGTACA CACCGCCCGT CACACCATGG
  Pseudomonas aeruginosa	ACGTTCCCGG GCCTTGTACA CACCGCCCGT CACACCATGG
  Clostridium perfringens	ACGTTCCCNG GTCTTGTACA CACCGCNCGT CACACCATGA
  Mycoplasma hominis	ACGTTCTCGG GTCTTGTACA CACCGCCCGT CACACCATGG
  Helicobacter pylori	ACGTTCCCGG GTCTTGTACT CACCGCCCGT CACACCATGG
  Mycoplasma pneumoniae	ACGTTCTCGG GTCTTGTACA CACCGCCCGT CAAACTATGA

• [0113]

  Annex III. Selection of universal probes by alignment of the
  sequences of bacterial 16S and 23S ribosomal RNA genes.

  Reverse strand of SEQ ID NO 125:	TCG TAGTCCGGAT TGGAGTCTGC AACTC
  	1361                                   1400
  Escherichia coli	AAGTGCGTCG TAGTCCGGAT TGGAGTCTGC AACTCGACTC
  Neisseria gonorrhoeae	AAACCGATCG TAGTCCGGAT TGCACTCTGC AACTCGAGTG
  Pseudomonas cepacia	AAACCGATCG TAGTCCGGAT TGCACTCTGC AACTCGAGTG
  Serratia marcescens	AAGTATGTCG TAGTCCGGAT TGGAGTCTGC AACTCGACTC
  Proteus vulgaris	AAGTCTGTCG TAGTCCGGAT TGGAGTCTGC AACTCGACTC
  Haemophilus influenzae	AAGTACGTCT AAGTCCGGAT TGGAGTCTGC AACTCGACTC
  Pseudomonas aeruginosa	AAACCGATCG TAGTCCGGAT CGCAGTCTGC AACTCGACTG
  Clostridium perfringens	AAACCAGTCT CAGTTCGGAT TGTAGGCTGA AACTCGCCTA
  Mycoplasma hominis	AAGCCGATCT CAGTTCGGAT TGGAGTCTGC AATTCGACTC
  Helicobacter pylori	ACACC..TCT CAGTTCGGAT TGTAGGCTGC AACTCGCCTG
  Mycloplasma pneumoniae	AAGTTGGTCT CAGTTCGGAT TGAGGGCTGC AATTCGTCCT

• [0114]

  Annex III. Selection of universal probes by alignment of the
  sequences of bacterial 16S and 23S ribosomal RNA genes.

  Reverse strand of SEQ ID NO: 128	CT CTCTGCTAAA CCGCAAGGTG ATGTATAGGG
  	1991                                              2040
  Lactobacillus lactis	AAACACAGCT CTCTGCTAAA CCGCAAGGTG ATGTATAGGG GGTGACGCCT
  Escherichia coli	AAACACAGCA CTGTGCAAAC ACGAAAGTGG ACGTATACGG TGTGACGCCT
  Pseudomonas aeruginosa	AAACACAGCA CTCTGCAAAC ACGAAAGTGG ACGTATAGGG TGTGACGCCT
  Pseudomonas cepacia	AAACACAGCA CTCTGCAAAC ACGAAAGTGG ACGTATAGGG TGTGACGCCT
  Bacillus stearothermophilus	AAACACAGGT CTCTGCGAAG TCGTAAGGCG ACGTATAGGG GCTGACACCT
  Micrococcus luteus	AAACACAGGT CCATGCGAAG TCGTAAGACG ATGTATATGG ACTGACTCCT
  SEQ ID NO: 129	GGGGGGACC ATCCTCCAAG GCTAAATAC
  	481                                                530
  Escherichia coli	TGTCTGAATA TGGGGGGACC ATCCTCCAAG GCTAAATACT CCTGACTGAC
  Pseudomonas aeruginosa	TGTCTGAACA TGGGGGGACC ATCCTCCAAG GCTAAATACT ACTGACTGAC
  Pseudomonas cepacia	TGTCTGAAGA TGGGGGGACC ATCCTCCAAG GCTAAATACT CGTGATCGAC
  Lactobacillus lactis	AGTTTGAATC CGGGAGGACC ATCTCCCAAC CCTAAATACT CCTTAGTGAC
  Micrococcus luteus	CGTGTGAATC TGCCAGGACC ACCTGGTAAG CCTGAATACT ACCTGTTGAC

• [0115]

  Annex III. Selection of universal probes by alignment of the
  sequences of bacterial 16S and 23S ribosomal RNA genes.

  Reverse strand of SEQ ID NO: 130	AACACAGCA CTCTGCAAAC ACGAAAGTGG ACG
  	1981                                              2030
  Pseudomonas aeruginosa	TGTTTATTAA AAACACAGCA CTCTGCAAAC ACGAAAGTGG ACGTATAGGG
  Escherichia coli	TGTTTATTAA AAACACAGCA CTGTGCAAAC ACGAAAGTGG ACGTATACGG
  Pseudomonas cepacia	TGTTTAATAA AAACACAGCA CTCTGCAAAC ACGAAAGTGG ACGTATAGGG
  Bacillus stearothermophilus	TGTTTATCAA AAACACAGGT CTCTGCGAAG TCGTAAGGCG ACGTATAGGG
  Lactobacillus lactis	TGTTTATCAA AAACACAGCT CTCTGCTAAA CCACAAGGTG ATGTATAGGG
  Micrococcus luteus	TGTTTATCAA AAACACAGGT CCATGCGAAG TCGTAAGACG ATGTATATGG

• [0116]

  Annex IV. Selection of the universal PCR primers by alignment of the bacterial
  16S ribosomal RNA gene

  SEQ ID NO: 126	GGAGGAA GGTGGGGATG ACG
  Reverse strand of SEQ ID NO: 127	CA CACCGCCCGT CACACCAT
  	1241                        1270......1461                        1490
  Escherichia coli	ACTGGAGGAA GGTGGGGATG ACGTCAAGTC......GCCTTGTACA CACCGCCCGT CACACCATGG
  Neisseria gonorrhoeae	GCCGGAGGAA GGTGGGGATG ACGTCAAGTC......NNCTTGTACA CACCGCCCGT CACACCATGG
  Pseudomonas cepacia	ACCGGAGGAA GGTNGGGATG ACGTCAAGTC......GTCTTGTACA CACNGCCCGT CACACCATGG
  Seratia marcescens	ACTGGAGGAA GGTGGGGATG ACGTCAAGTC......GCCTTGTACA CACCGCCCGT CACACCATGG
  Proteus vulgaris	ACCGGAGGAA GGTGGGGATG ACGTTAAGTC......GCCTTGTACA CACCGCCCGT CACACCATGG
  Haemophilus influenzae	ACTGGAGGAA GGTNGGGATG ACGTCAAGTC......GCNTTGTACA CACCGCCCGT CACACCATGG
  Legionella pneumophila	ACCGGAGGAA GGCGGGGATG ACGTCAAGTC......GCCTTGTACA CACCGCCCGT CACACCATGG
  Pseudomonas aeruginos	ACCGGAGGAA GGTGGGGATG ACGTCAAGTC......GCCTTGTACA CACCGCCCGT CACACCATGG
  Clostridium perfringens	CCAGGAGGAA GGTGGGGATG ACGTNNAATC......GTCTTGTACA CACCGCNCGT CACACCATGA
  Mycoplasma hominis	CTGGGAGGAA GGTGGGGATG ACGTCAAATC......GTCTTGTACA CACCGCCCGT CACACCATGG
  Helicobacter pylori	GGAGGAGGAA GGTGGGGACG ACGTCAAGTC......GTCTTGTACT CACCGCCCGT CACACCATGG
  Mycoplasma pneumoniae	ATTGGAGGAA GGAAGGGATG ACGTCAAATC......GTCTTGTACA CACCGCCCGT CAAACTATGA

• [0117]
• 117711817DNAEnterococcus faecalis 1 acagtaaaaa agttgttaac gaatgaattt gttaacaact tttttgctat ggtattgagt 60 tatgaggggc aatacaggga aaaatgtcgg ctgattaagg aatttagata gtgccggtta 120 gtagttgtct ataatgaaaa tagcaacaaa tatttacgca gggaaagggg cggtcgttta 180 acgggaaaaa ttagggagga taaagcaata cttttgttgg gaaaagaaat aaaaggaaac 240 tggggaagga gttaattgtt tgatgaaggg aaataaaatt ttatacattt taggtacagg 300 catctttgtt ggaagttcat gtctattttc ttcacttttt gtagccgcag aagaacaagt 360 ttattcagaa agtgaagttt caacagtttt atcgaagttg gaaaaggagg caatttctga 420 ggcagctgct gaacaatata cggttgtaga tcgaaaagaa gacgcgtggg ggatgaagca 480 tcttaagtta gaaaagcaaa cggaaggcgt tactgttgat tcagataatg tgattattca 540 tttagataaa aacggtgcag taacaagtgt tacaggaaat ccagttgatc aagttgtgaa 600 aattcaatcg gttgatgcaa tcggtgaaga aggagttaaa aaaattgttg cttctgataa 660 tccagaaact aaagatcttg tctttttagc tattgacaaa cgtgtaaata atgaagggca 720 attattttat aaagtcagag taacttcttc accaactggt gaccccgtat cattggttta 780 taaagtgaac gctacagatg gaacaattat ggaaaaacaa gatttaacgg aacatgtcgg 840 tagtgaagta acgttaaaaa actcttttca agtaacgttt aatgtaccag ttgaaaaaag 900 caatacggga attgctttac acggaacgga taacacaggg gtttaccatg cagtagttga 960 tggcaaaaat aattattcta ttattcaagc gccatcacta gcgacattaa atcagaatgc 1020 tattgacgcc tatacgcatg gaaaatttgt gaaaacatat tatgaagatc atttccaacg 1080 acacagtatt gatgatcgag ggatgcccat cttgtcagtt gttgatgaac aacatccaga 1140 tgcttatgac aatgcttttt gggatggaaa agcaatgcgt tatggtgaaa caagtacacc 1200 aacaggaaaa acgtatgctt cctctttaga tgtagttggt catgaaatga cacatggtgt 1260 gacggaacat actgccggtt tagaatattt aggacaatca ggtgccttga atgaatctta 1320 ttctgatttg atgggttata ttatttcggg tgcatctaat ccagaaattg gtgcggatac 1380 tcagagtgtt gaccgaaaaa caggtattcg aaatttacaa acgccaagta aacacggaca 1440 accagaaacc atggctcaat acgacgatcg agcacggtat aaaggaacgc cttattatga 1500 tcaaggcggt gttcattata acagtggaat tattaatcgg attggttaca ccattatcca 1560 gaacttaggc attgaaaaag cacagactat tttctacagc tcgttagtaa attacttaac 1620 acctaaagca caattcagtg atgctcgtga tgcgatgctt gctgctgcaa aagttcaata 1680 tggcgatgaa gcagcttcag tggtgtcagc agcctttaac tctgctggaa tcggagctaa 1740 agaagacatt caggtaaacc aaccaagtga atctgttctg gtcaatgaat gaaaaaaatt 1800 ccccaattaa ataaaaa 181722275DNAEnterococcus faecalis 2 ggtaccaaag aaaaaaacga acgccacaac caacagcctc taaagcaaca cctgcttctg 60 aaattgaggg agatttagca aatgtcaatg agattctttt ggttcacgat gatcgtgtcg 120 ggtcagcaac gatgggaatg aaagtcttag aagaaatttt agataaagag aaaatttcaa 180 tgccgattcg aaaaattaat attaatgaat taactcaaca aacacaggct ttaattgtca 240 caaaagctga actaacggaa caagcacgta aaaaagcacc gaaagcgaca cacttatcag 300 taaaaagtta tggttaatcc ccaaaaatat gaaacagtgg gtttcgctct taaaagaaag 360 tgcctagaga ggaagaaaac aatggaaaat cttacgaata tttcaattga attaaatcaa 420 cagtttaata caaaagaaga agctattcgc ttttccggcc agaaactagt cgaggcaggc 480 tgtgttgagc ccgcttatat cgaagcaatg attgaaagag accaattgct atctgcccat 540 atggggaatt ttattgccat tcctcatgga acagaagaag ccaaaaaatt agtgaaaaaa 600 tcaggaatct gtgtagtgca agtcccagag ggcgttaatt ttggcaccga agaagatgaa 660 aaaattgcta ccgtattatt tgggattgcc ggagtcggtg aagaacattt gcaattagtc 720 caacaaattg cactttattg tagtgatatg gataacgtgg tgcaacttgc cgatgcatta 780 agtaaagaag aaataacaga aaatttagcc attgcttaaa ggagagaata agaatgaacg 840 cagtacattt tggagcagga aatattggac gcggctttat tggcgaaatt ttagctaaaa 900 cgggtttcat attaccgttt gtggatgtta atggaaacca tcatcaagcg ttaaaagaac 960 gtaaaagtta tacaattgaa ttggccgatg cctcacatca acaaattaac gttgaaaatg 1020 tgaccgggtt aaataacatg acagaaccag aaaaagtagt agaagcaatt gcggaagccg 1080 atttagtcac gacggcaatt ggtcctaata ttttaccaag aattgctgaa ttaattgctc 1140 aaggaattga tgcacgtgcc gaagcaaatt gtcaaaacgg cccgctggat attatcgctt 1200 gtgaaaatat gattggtggt tcaacctttt tagcagaaga agtggccata atatttgaaa 1260 aacccagctt atctgaacaa tggattggtt ttcctgatgc ggcagttgat cggattgttc 1320 cattacaaaa acataaagat ccactttttg ttcaagttga gcctttttgt gaatgggtca 1380 ttgatgatac caaccgaaaa gccaaagaga ttcagttaga aggcgtcatt acttgtcgat 1440 tagagccgta tattgaacga aaattattta gtgtaaccag tggccatgct acagttgcct 1500 atacaggggc gttgttaggc tatcaaacca ttgacgaagc gatgcaggac gccttagtgg 1560 tagcgcaact caaatcagtt ttgcaggaaa ccggtaaact tttagtggcc aaatggaatt 1620 ttgatgaaca agaacatgca gcctatattg aaaaaattat caaccgtttc caaaataaat 1680 atatttcaga tgctattaca cgtgtagcac ggacaccaat cagaaaatta ggtgcgcaag 1740 aacggtttat tcgaccaatc cgtgaattac aggaacgcaa tctagtgtcg gccgcattta 1800 tagcaatgat tggtattgtc tttaattatc atgatccaga agatgaacaa agccgtcaat 1860 tacaggaaat gcttgaccaa gaaagtgttg atacagtgga tcgctgaagt aacgggcatt 1920 gaagatccag aaacggttaa aaatattaaa caaaacgtag aactgctatg cgcgaccaca 1980 agtagcataa ttaacaaaat ccttctacca agatacttca catttcttaa ttaaagaaaa 2040 aacaaccgcg cctcacctga gccgaccccc aaaagttaga cctagaaatc taacttttgg 2100 aggttttttt gtatggcaaa atacagtttt gaaatttaaa cttaaacttg ttcatgacta 2160 cttatatggt caaggaggtc taaggtttct cgcaaagaag tatgggttta aagatagtct 2220 caaataagca aatggataaa tgcctataaa gaacttggtg aagaaggggg gatcc 22753227DNAEscherichia coli 3 gatccgccat gggttgtttt ccgattgagg attttataga tggtttctgg cgacctgcac 60 aggagtacgg tgatttttaa ttattgcaat tgcacaagag tcagttctcc cccaaagaca 120 gcaccggtat caatataatg caggttgcca atatccacgc gatggcgcaa aggtgtatga 180 ccaaaccaga aatgatcggc cacctgcatc gccagttcgc gagtcgg 2274278DNAEscherichia coli 4 gatctaaatc aaattaattg gttaaagata accacagcgg ggccgacata aactctgaca 60 agaagttaac aaccatataa cctgcacagg acgcgaacat gtcttctcat ccgtatgtca 120 cccagcaaaa taccccgctg gcggacgaca ccactctgat gtccactacc gatctcgctt 180 tccagcgtca tattggggcg cgctacgttg gggcgtgggc gtaattggtc aatcaggcgc 240 ggggtcagcg gataaacatt caccattttg tcgagatc 27851596DNAEscherichia coli 5 atggctgaca ttctgctgct cgataatatc gactctttta cgtacaacct ggcagatcag 60 ttgcgcagca atgggcataa cgtggtgatt taccgcaacc atataccggc gcaaacctta 120 attgaacgct tggcgaccat gagtaatccg gtgctgatgc tttctcctgg ccccggtgtg 180 ccgagcgaag ccggttgtat gccggaactc ctcacccgct tgcgtggcaa gctgcccatt 240 attggcattt gcctcggaca tcaggcgatt gtcgaagctt acgggggcta tgtcggtcag 300 gcgggcgaaa ttctccacgg taaagcctcc agcattgaac atgacggtca ggcgatgttt 360 gccggattaa caaacccgct gccggtggcg cgttatcact cgctggttgg cagtaacatt 420 ccggccggtt taaccatcaa cgcccatttt aatggcatgg tgatggcagt acgtcacgat 480 gcggatcgcg tttgtggatt ccagttccat ccggaatcca ttctcaccac ccagggcgct 540 cgcctgctgg aacaaacgct ggcctgggcg cagcataaac tagagccagc caacacgctg 600 caaccgattc tggaaaaact gtatcaggcg cagacgctta gccaacaaga aagccaccag 660 ctgttttcag cggtggtgcg tggcgagctg aagccggaac aactggcggc ggcgctggtg 720 agcatgaaaa ttcgcggtga gcacccgaac gagatcgccg gggcagcaac cgcgctactg 780 gaaaacgcag cgccgttccc gcgcccggat tatctgtttg ctgatatcgt cggtactggc 840 ggtgacggca gcaacagtat caatatttct accgccagtg cgtttgtcgc cgcggcctgt 900 gggctgaaag tggcgaaaca cggcaaccgt agcgtctcca gtaaatctgg ttcgtccgat 960 ctgctggcgg cgttcggtat taatcttgat atgaacgccg ataaatcgcg ccaggcgctg 1020 gatgagttag gtgtatgttt cctctttgcg ccgaagtatc acaccggatt ccgccacgcg 1080 atgccggttc gccagcaact gaaaacccgc accctgttca atgtgctggg gccattgatt 1140 aacccggcgc atccgccgct ggcgttaatt ggtgtttata gtccggaact ggtgctgccg 1200 attgccgaaa ccttgcgcgt gctggggtat caacgcgcgg cggtggtgca cagcggcggg 1260 atggatgaag tttcattaca cgcgccgaca atcgttgccg aactgcatga cggcgaaatt 1320 aaaagctatc agctcaccgc agaagacttt ggcctgacac cctaccacca ggagcaactg 1380 gcaggcggaa caccggaaga aaaccgtgac attttaacac gtttgttaca aggtaaaggc 1440 gacgccgccc atgaagcagc cgtcgctgcg aacgtcgcca tgttaatgcg cctgcatggc 1500 catgaagatc tgcaagccaa tgcgcaaacc gttcttgagg tactgcgcag tggttccgct 1560 tacgacagag tcaccgcact ggcggcacga gggtaa 159662703DNAEscherichia coli 6 gacgacttag ttttgacgga atcagcatag ttaatcactt cactgtggaa aatgaggaaa 60 tattattttt tttgcgcttc gtaattaatg gttataaggt cggccagaaa cctttctaat 120 gcaagcgatg acgttttttt atgtgtctga atttgcactg tgtcacaatt ccaaatcttt 180 attaacaact cacctaaaac gacgctgatc cagcgtgaat actggtttcc cttatgttca 240 tcagattcat ttaagcaagg gtttcttctt cattcctgat gaaagtgcca tctaaaaaga 300 tgatcttaat aaatctatta agaatgagat ggagcacact ggatatttta cttatgaaac 360 tgtttcactc ctttacttaa tttatagagt taccttccgc tttttgaaaa tacgcaacgg 420 ccattttttg cacttagata cagattttct gcgctgtatt gcattgattt gatgctaatc 480 ctgtggtttg cactagcttt aagtggttga gatcacattt ccttgctcat ccccgcaact 540 cctccctgcc taatcccccg caggatgagg aaggtcaaca tcgagcctgg caaactagcg 600 ataacgttgt gttgaaaatc taagaaaagt ggaactccta tgtcacaacc tatttttaac 660 gataagcaat ttcaggaagc gctttcacgt cagtggcagc gttatggctt aaattctgcg 720 gctgaaatga ctcctcgcca gtggtggcta gcagtgagtg aagcactggc cgaaatgctg 780 cgtgctcagc cattcgccaa gccggtggcg aatcagcgac atgttaacta catctcaatg 840 gagtttttga ttggtcgcct gacgggcaac aacctgttga atctcggctg gtatcaggat 900 gtacaggatt cgttgaaggc ttatgacatc aatctgacgg acctgctgga agaagagatc 960 gacccggcgc tgggtaacgg tggtctggga cgtctggcgg cgtgcttcct cgactcaatg 1020 gcaactgtcg gtcagtctgc gacgggttac ggtctgaact atcaatatgg tttgttccgc 1080 cagtcttttg tcgatggcaa acaggttgaa gcgccggatg actggcatcg cagtaactac 1140 ccgtggttcc gccacaacga agcactggat gtgcaggtag ggattggcgg taaagtgacg 1200 aaagacggac gctgggagcc ggagtttacc attaccggtc aagcgtggga tctccccgtt 1260 gtcggctatc gtaatggcgt ggcgcagccg ctgcgtctgt ggcaggcgac gcacgcgcat 1320 ccgtttgatc tgactaaatt taacgacggt gatttcttgc gtgccgaaca gcagggcatc 1380 aatgcggaaa aactgaccaa agttctctat ccaaacgaca accatactgc cggtaaaaag 1440 ctgcgcctga tgcagcaata cttccagtgt gcctgttcgg tagcggatat tttgcgtcgc 1500 catcatctgg cggggcgtga actgcacgaa ctggcggatt actaagttat tcagctgaac 1560 gatacccacc caactatcgc gattccagaa ctgctgcgcg tgctgatcga tgagcaccag 1620 atgagctggg atgacgcttg ggccattacc agcaaaactt tcgcttacac caaccatacc 1680 ctgatgccag aagcgctgga acgctgggat gtgaaactgg tgaaaggctt actgccgcgc 1740 cacatgcaga ttattaacga aattaatact cgctttaaaa cgctggtaga gaaaacctgg 1800 ccgggcgatg aaaaagtgtg ggccaaactg gcggtggtgc acgacaaaca agtgcatatg 1860 gcgaacctgt gtgtggttgg cggtttcgcg gtgaacggtg ttgcggcgct gcactcggat 1920 ctggtggtga aagatctgtt cccggaatat caccagctat ggccgaacaa attccataac 1980 gtcaccaacg gtattacccc acgtcgctgg atcaaacagt gcaacccggc actggcggct 2040 ctgttggata aatcactgca aaaagagtgg gctaacgatc tcgatcagct gatcaatctg 2100 gttaaattgg ctgatgatgc gaaattccgt cagctttatc gcgtgatcaa gcaggcgaat 2160 aaagtccgtc tggcggagtt tgtgaaagtt cgtaccggta ttgacatcaa tccacaggcg 2220 attttcgata ttcagatcaa acgtttgcac gagtacaaac gccagcacct gaatctgctg 2280 cgtattctgg cgttgtacaa agaaattcgt gaaaacccgc aggctgatcg cgtaccgcgc 2340 gtcttcctct tcggcgcgaa agcggcaccg ggctactacc tggctaagaa tattatcttt 2400 gcgatcaaca aagtggctga cgtgatcaac aacgatccgc tggttggcga taagttgaag 2460 gtggtgttcc tgccggatta ttgcgtttcg gcggcggaaa aactgatccc ggcggcggat 2520 atctccgaac aaatttcgac tgcaggtaaa gaagcttccg gtaccggcaa tatgaaactg 2580 gcgctcaatg gtgcgcttac tgtcggtacg ctggatgggg cgaacgttga aatcgccgag 2640 aaagtcggtg aagaaaatat ctttattttt ggtcatacgg tcaaacaagt gaaggcaatc 2700 gac 270371391DNAEscherichia coli 7 agagaagcct gtcggcaccg tctggtttgc ttttgccact gcccgcggtg aaggcattac 60 ccggcgggat gcttcagcgg cgaccgtgat gcggtgcgtc gtcaggctac tgcgtatgca 120 ttgcagacct tgtggcaaca atttctacaa aacacttgat actgtatgag catacagtat 180 aattgcttca acagaacata ttgactatcc ggtattaccc ggcatgacag gagtaaaaat 240 ggctatcgac gaaaacaaac agaaagcgtt ggcggcagca ctgggccaga ttgagaaaca 300 atttggtaaa ggctccatca tgcgcctggg tgaagaccgt tccatggatg tggaaaccat 360 ctctaccggt tcgctttcac tggatatcgc gcttggggca ggtggtctgc cgatgggccg 420 tatcgtcgaa atctacggac cggaatcttc cggtaaaacc acgctgacgc tgcaggtgat 480 cgccgcagcg cagcgtgaag gtaaaacctg tgcgtttatc gatgctgaac acgcgctgga 540 cccaatctac gcacgtaaac tgggcgtcga tatcgacaac ctgctgtgct cccagccgga 600 caccggcgag caggcactgg aaatctgtga cgccctggcg cgttctggcg cagtagacgt 660 tatcgtcgtt gactccgtgg cggcactgac gccgaaagcg gaaatcgaag gcgaaatcgg 720 cgactctcac atgggccttg cggcacgtat gatgagccag gcgatgcgta agctggcggg 780 taacctgaag cagtccaaca cgctgctgat cttcatcaac cagatccgta tgaaaattgg 840 tgtgatgttc ggtaacccgg aaaccactac cggtggtaac gcgctgaaat tctacgcctc 900 tgttcgtctc gacatccgtc gtatcggcgc ggtgaaagag ggcgaaaacg tggtgggtag 960 cgaaacccgc gtgaaagtgg tgaagaacaa aatcgctgcg ccgtttaaac aggctgaatt 1020 ccagatcctc tacggcgaag gtatcaactt ctacggcgaa ctggttgacc tgggcgtaaa 1080 agagaagctg atcgagaaag caggcgcgtg gtacagctac aaaggtgaga agatcggtca 1140 gggtaaagcg aatgcgactg cctggctgaa agataacccg gaaaccgcga aagagatcga 1200 gaagaaagta cgtgagttgc tgctgagcaa cccgaactca acgccggatt tctctgtaga 1260 tgatagcgaa ggcgtagcag aaactaacga agatttttaa tcgtcttgtt tgatacacaa 1320 gggtcgcatc tgcggccctt ttgctttttt aagttgtaag gatatgccat gacagaatca 1380 acatcccgtc g 13918238DNAKlebsiella pneumoniae 8 tcgccaggaa ggcggcattc ggctgggtca gagtgacctg cagcgtggtg tcgttcagcg 60 ctttcacccc caacgtctcg ggtccctttt gcccgagggc aatctcgcgg gcgttggcga 120 tatgcatatt gccagggtag ctcgcgtagg gggaggctgt tgccggcgag accagccgtt 180 gccagctcca gacgatatcc tgcgctgtaa tggccgtgcc gtcagaccag gtcagacc 2389385DNAKlebsiella pneumoniae 9 cagcgtaatg cgccgcggca taacggcgcc actatcgaca gtcagttcgt cagcctgcag 60 cctgggctga atctgggacc atggcgcctg ccgaactaca gcacctatag ccacagcgat 120 aacaacagcc gctgggagtc ggtttactcc tatcttgccc gcgatattca caccctacgc 180 agccagctgg tggtcggtaa tacgtatacc tcttccggca ttttcgacag tttgagtttt 240 accggtctgc agctcagttc gacaaagaga tgctgccgga tagcctgcat gctttgcgcc 300 gacgattcga gggatcgcgc gcaccaccgc ggaggtctcg gtttatcaga atggttacag 360 catttataaa accaccgtcg ctacc 38510462DNAKlebsiella pneumoniae 10 ctctatattc aggacgaaca tatctggacc tctggcgggg tcagttccgg ctttgatcgc 60 cctgcacccg cagcgggtga tcgcccctca tctgctactg cggcgctgca acaggcgacg 120 atcgatgacg ttattcctgg ccagcaaaca gcagaccaat taaggtctga tagtggctct 180 cttcctccgg cgcgcgacgg tccaggcggc tcaacagttt ggtgcatagc gctttgcggt 240 tgagatgacg cccttcgtta agaatatcca tcacgatctc cgtccatgga gagtagcgtt 300 tattccagaa tagggttttt caggatctca tggatctgcg cctgcttatc gctattttgt 360 aaccagatcg cataaagtgg acgggataac gtagcgctgt ccatgaccgt atgtaaccca 420 tgcttctctt tcgcccagcg agcaggtagc caacagcagc cg 46211730DNAKlebsiella pneumoniae 11 gctgaccgct aaactgggtt acccgatcac tgacgatctg gacatctaca cccgtctggg 60 cggcatggtt tggcgcgctg actccaaagg caactacgct tcaaccggcg tttcccgtag 120 cgaacacgac actggcgttt ccccagtatt tgctggcggc gtagagtggg ctgttactcg 180 tgacatcgct acccgtctgg aataccagtg ggttaacaac atcggcgacg cgggcactgt 240 gggtacccgt cctgataacg gcatgctgag cctgggcgtt tcctaccgct tcggtcagga 300 agatgctgca ccggttgttg ctccggctcc ggctccggct ccggaagtgg ctaccaagca 360 cttcaccctg aagtctgacg ttctgttcaa cttcaacaaa gctaccctga aaccggaagg 420 tcagcaggct ctggatcagc tgtacactca gctgagcaac atggatccga aagacggttc 480 cgctgttgtt ctgggctaca ccgaccgcat cggttccgaa gcttacaacc agcagctgtc 540 tgagaaacgt gctcagtccg ttgttgacta cctggttgct aaaggcatcc cggctggcaa 600 aatctccgct cgcggcatgg gtgaatccaa cccggttact ggcaacacct gtgacaacgt 660 gaaagctcgc gctgccctga tcgattgcct ggctccggat cgtcgtgtag agatcgaagt 720 taaaggtatc 73012225DNAProteus mirabilis 12 cgctactgtt taaatctcat ttgaaacatc gcaaagtcag tgaaccacat attcgaggat 60 ggcatgcact agaaaatatt aataagattt tagcgaaacc taatcagcgc aatatcgctt 120 aattatttta ggtatgttct cttctatcct acagtcacga ggcagtgtcg aacttgatcc 180 tcattttatt aatcacatga ccaatggtat aagcgtcgtc acata 22513402DNAProteus mirabilis 13 acattttaaa taggaagcca cctgataaca tccccgcagt tggatcatca gatttatagc 60 ggcatttggt atccgctaga taaaagcagt ccaacgatcc cgccaattgt tagatgaaat 120 tggactattc tttttatttg ctccgcttta tcacagtggt tttcgctttg ccgcccctgt 180 gcgccaacag ctaagaacac gcacgctctt taatgtgtta ggcccattaa ttaatccagc 240 gcgttccgcc tttagcatta attggtgttt atagtcctga attattaatg cctattgcag 300 ataccttaaa tgtcttgggc tacaaacgtg cggcagtggt ccatagtggt ggaatggatg 360 aagtgtcatt acatgctccc acacaagtgg ctgagttaca ca 40214157DNAProteus mirabilis 14 ctgaaacgca tttatgcggg agtcagtgaa atcatcactc aattttcacc cgatgtattt 60 tctgttgaac aagtctttat ggcaaaaaat gcagactcag cattaaaatt aggccaagca 120 agaggtgtgg cgattttagc ggcagtcaat aatgatc 157151348DNAProteus mirabilis 15 tttctcttta aaatcaattc ttaaagaaat tattaataat taacttgata ctgtatgatt 60 atacagtata atgagtttca acaagcaaaa tcatatacgt tttaatggta gtgacccatc 120 tttatgcttc actgcccaga gggagataac atggctattg atgaaaacaa acaaaaagca 180 ttggccgcag cacttggtca aattgaaaag caatttggta aaggttctat catgcgtctg 240 ggcgaagacc gttccatgaa cgtagaaact atctctacag gatctttatc attagacgtt 300 gctttaggtg caggtggatt gccacgtggc cgtattgttg aaatctatgg ccctgaatct 360 tctggtaaaa caaccttgac tctacaagtt attgcctctg ctcagcgtga aggaaaaatt 420 tgtgcattta ttgatgctga acatgcatta gacccaattt atgctcaaaa gctaggtgtc 480 gatatcgata atctactctg ctctcaacct gacacaggtg aacaagctct ggaaatttgt 540 gatgcattat ctcgctctgg tgcggtcgat gttattgtcg tggactccgt ggcagcatta 600 acaccaaaag ctgaaattga aggtgaaatt ggtgattcac acgttggttt agccgcacgt 660 atgatgagcc aagctatgcg taaactagcg ggtaacctta aaaactctaa tacactgctg 720 attttcatta accaaattcg tatgaaaatc ggtgttatgt ttggtaaccc agaaaccacg 780 accggtggta atgcgcttaa attctatgct tctgttcgtt tagacattcg tcgcattggc 840 tctgtcaaaa atggtgatga agtcattggt agtgagactc gcgttaaagt tgttaaaaat 900 aaagtggctg caccgtttaa acaagctgaa ttccaaatta tgtacggtga aggtattaat 960 acctatggcg aactgattga tttaggtgtt aaacataagt tagtagagaa agcaggtgct 1020 tggtatagct acaatggcga aaaaattggt caaggtaaag ctaacgcaac caattactta 1080 aaagaacatc ctgaaatgta caatgagtta aacactaaat tgcgtgaaat gttgttaaat 1140 catgctggtg aattcacaag tgctgcggat tttgcaggtg aagagtcaga cagtgatgct 1200 gacgacacaa aagagtaatt agctggttgt catgctgttt gtgtgaaaat agaccttaaa 1260 tcattggcta ttatcacgac agcatcccat agaataactt gtttgtataa attttattca 1320 gatggcaaag gaagccttaa aaaagctt 1348162167DNAPseudomonas aeruginosa 16 ggtaccgctg gccgagcatc tgctcgatca ccaccagccg ggcgacggga actgcacgat 60 ctacctggcg agcctggagc acgagcgggt tcgcttcgta cggcgctgag cgacagtcac 120 aggagaggaa acggatggga tcgcaccagg agcggccgct gatcggcctg ctgttctccg 180 aaaccggcgt caccgccgat atcgagcgct cgcacgcgta tggcgcattg ctcgcggtcg 240 agcaactgaa ccgcgagggc ggcgtcggcg gtcgcccgat cgaaacgctg tcccaggacc 300 ccggcggcga cccggaccgc tatcggctgt gcgccgagga cttcattcgc aaccgggggg 360 tacggttcct cgtgggctgc tacatgtcgc acacgcgcaa ggcggtgatg ccggtggtcg 420 agcgcgccga cgcgctgctc tgctacccga ccccctacga gggcttcgag tattcgccga 480 acatcgtcta cggcggtccg gcgccgaacc agaacagtgc gccgctggcg gcgtacctga 540 ttcgccacta cggcgagcgg gtggtgttca tcggctcgga ctacatctat ccgcgggaaa 600 gcaaccatgt gatgcgccac ctgtatcgcc agcacggcgg cacggtgctc gaggaaatct 660 acattccgct gtatccctcc gacgacgact tgcagcgcgc cgtcgagcgc atctaccagg 720 cgcgcgccga cgtggtcttc tccaccgtgg tgggcaccgg caccgccgag ctgtatcgcg 780 ccatcgcccg tcgctacggc gacggcaggc ggccgccgat cgccagcctg accaccagcg 840 aggcggaggt ggcgaagatg gagagtgacg tggcagaggg gcaggtggtg gtcgcgcctt 900 acttctccag catcgatacg cccgccagcc gggccttcgt ccaggcctgc catggtttct 960 tcccggagaa cgcgaccatc accgcctggg ccgaggcggc ctactggcag accttgttgc 1020 tcggccgcgc cgcgcaggcc gcaggcaact ggcgggtgga agacgtgcag cggcacctgt 1080 acgacatcga catcgacgcg ccacaggggc cggtccgggt ggagcgccag aacaaccaca 1140 gccgcctgtc ttcgcgcatc gcggaaatcg atgcgcgcgg cgtgttccag gtccgctggc 1200 agtcgcccga accgattcgc cccgaccctt atgtcgtcgt gcataacctc gacgactggt 1260 ccgccagcat gggcggggga ccgctcccat gagcgccaac tcgctgctcg gcagcctgcg 1320 cgagttgcag gtgctggtcc tcaacccgcc gggggaggtc agcgacgccc tggtcttgca 1380 gctgatccgc atcggttgtt cggtgcgcca gtgctggccg ccgccggaag ccttcgacgt 1440 gccggtggac gtggtcttca ccagcatttt ccagaatggc caccacgacg agatcgctgc 1500 gctgctcgcc gccgggactc cgcgcactac cctggtggcg ctggtggagt acgaaagccc 1560 cgcggtgctc tcgcagatca tcgagctgga gtgccacggc gtgatcaccc agccgctcga 1620 tgcccaccgg gtgctgcctg tgctggtatc ggcgcggcgc atcagcgagg aaatggcgaa 1680 gctgaagcag aagaccgagc agctccagga ccgcatcgcc ggccaggccc ggatcaacca 1740 ggccaaggtg ttgctgatgc agcgccatgg ctgggacgag cgcgaggcgc accagcacct 1800 gtcgcgggaa gcgatgaagc ggcgcgagcc gatcctgaag atcgctcagg agttgctggg 1860 aaacgagccg tccgcctgag cgatccgggc cgaccagaac aataacaaga ggggtatcgt 1920 catcatgctg ggactggttc tgctgtacgt tggcgcggtg ctgtttctca atgccgtctg 1980 gttgctgggc aagatcagcg gtcgggaggt ggcggtgatc aacttcctgg tcggcgtgct 2040 gagcgcctgc gtcgcgttct acctgatctt ttccgcagca gccgggcagg gctcgctgaa 2100 ggccggagcg ctgaccctgc tattcgcttt tacctatctg tgggtggccg ccaaccagtt 2160 cctcgag 2167171872DNAPseudomonas aeruginosa 17 gaattcccgg gagttcccga cgcagccacc cccaaaacac tgctaaggga gcgcctcgca 60 gggctcctga ggagatagac catgccattt ggcaagccac tggtgggcac cttgctcgcc 120 tcgctgacgc tgctgggcct ggccaccgct cacgccaagg acgacatgaa agccgccgag 180 caataccagg gtgccgcttc cgccgtcgat cccgctcacg tggtgcgcac caacggcgct 240 cccgacatga gtgaaagcga gttcaacgag gccaagcaga tctacttcca acgctgcgcc 300 ggttgccacg gcgtcctgcg caagggcgcc accggcaagc cgctgacccc ggacatcacc 360 cagcaacgcg gccagcaata cctggaagcg ctgatcacct acggcacccc gctgggcatg 420 ccgaactggg gcagctccgg cgagctgagc aaggaacaga tcaccctgat ggccaagtac 480 atccagcaca ccccgccgca accgccggag tggggcatgc cggagatgcg cgaatcgtgg 540 aaggtgctgg tgaagccgga ggaccggccg aagaaacagc tcaacgacct cgacctgccc 600 aacctgttct cggtgaccct gcgcgacgcc gggcagatcg ccctggtcga cggcgacagc 660 aaaaagatcg tcaaggtcat cgataccggc tatgccgtgc atatctcgcg gatgtccgct 720 tccggccgct acctgctggt gatcggccgc gacgcgcgga tcgacatgat cgacctgtgg 780 gccaaggagc cgaccaaggt cgccgagatc aagatcggca tcgaggcgcg ctcggtggaa 840 agctccaagt tcaagggcta cgaggaccgc tacaccatcg ccggcgccta ctggccgccg 900 cagttcgcga tcatggacgg cgagaccctg gaaccgaagc agatcgtctc cacccgcggc 960 atgaccgtag acacccagac ctaccacccg gaaccgcgcg tggcggcgat catcgcctcc 1020 cacgagcacc ccgagttcat cgtcaacgtg aaggagaccg gcaaggtcct gctggtcaac 1080 tacaaggata tcgacaacct caccgtcacc agcatcggtg cggcgccgtt cctccacgac 1140 ggcggctggg acagcagcca ccgctacttc atgaccgccg ccaacaactc caacaaggtt 1200 gccgtgatcg actccaagga ccgtcgcctg tcggccctgg tcgacgtcgg caagaccccg 1260 cacccggggc gtggcgccaa cttcgtgcat cccaagtacg gcccggtgtg gagcaccagc 1320 cacctgggcg acggcagcat ctcgctgatc ggcaccgatc cgaagaacca tccgcagtac 1380 gcctggaaga aagtcgccga actacagggc cagggcggcg gctcgctgtt catcaagacc 1440 catccgaagt cctcgcacct ctacgtcgac accaccttca accccgacgc caggatcagc 1500 cagagcgtcg cggtgttcga cctgaagaac ctcgacgcca agtaccaggt gctgccgatc 1560 gccgaatggg ccgatctcgg cgaaggcgcc aagcgggtgg tgcagcccga gtacaacaag 1620 cgcggcgatg aagtctggtt ctcggtgtgg aacggcaaga acgacagctc cgcgctggtg 1680 gtggtggacg acaagaccct gaagctcaag gccgtggtca aggacccgcg gctgatcacc 1740 ccgaccggta agttcaacgt ctacaacacc cagcacgacg tgtactgaga cccgcgtgcg 1800 gggcacgccc cgcacgctcc cccctacgag gaaccgtgat gaaaccgtac gcactgcttt 1860 cgctgctcgc ca 1872183451DNAPseudomonas aeruginosa 18 tcgagacggg aagccactct ctacgagaag acagaagccc ctcacagagg cctctgtcta 60 cgcctactaa agctcggctt attcatatgt atttatattc tttcaataga tcactcagcg 120 ctattttaag ttcaccctct gtaagttcac ctgggcgctc tttctttcct tcggtaaagc 180 tgtcggccag accaaacatt aaactcaagc atctcccaag cgatgcatca tcttgggcca 240 gcatccctga atcgcgcgtc ggacctccaa gtcttaaaaa attcttcgct gaaggttttc 300 ccatcaatcg atgaggctaa tagcttcttt gcaatatcta tcatttccat gctcacctta 360 aagcacctca tttttcatgt aaaaattgta ttgatccgtg ccagactcaa tcctccaccc 420 agaaacaaac atcccatcct ctccaatgat aacaacaata ttagtcctgg cattgtaatg 480 tacttttgag tttacttcgg agtggtaagt ccctttttct acggttgcag gatcagcaag 540 gtgctcaaga attttatccc taaactctgc aagcgttcca ttgttggcgc ttttttcacc 600 cagcccaaaa tcatatttgt ggctatcaaa ttttttctgt agttgcctcc gtgtgaagat 660 accactatca agaggactac tgagcattac ataaacaggt ttgactccag aatccgccgg 720 gaaaatcacg atcagatcgt ttaggtccag tagcattccc ggataggact ccgggccggt 780 cttcaacggt gtgagggccg ctccctcata taccggcacc ggcttcggta tgaccggagt 840 ggtactcgaa gggttctggt ttcctggagg actcgccggc gtccaagtca ggatcagtgg 900 cggcgcttct gcgaccgtag agggaaccgt aacctcgtac agtcctgttg cggcgttata 960 ggccccatcc ggaccggaac gctttcggaa cgctcacacc atcggtctga ccaccgaaag 1020 gtcgtcgtgt tgcctcgcgc ctcgttggtc aggcgcatcg gcagatcgac ggtaccgctg 1080 gcttttgcaa ccgcgttcag gtttacgctt gggggaagcc ccaatttagc ggcatccatg 1140 cccagggcgt aacgaacgct atcgggcgtt tggtcctgcc attgctcggc agtccgggag 1200 agtaggtcag actggcaagc cacggccatc accgaggtgc tgaagccagg accgccagga 1260 cggcaatcgc atcggagatc gcttgagcaa gggatgcggc gcctgtgcga cctggatcag 1320 accccgctgc ggcggtggcg cacccgctgc cattggctgg catggcataa gtattggcag 1380 ccctgatcgc cgcttgacga gcgatttcct tgcgccttgc cgtttcggcg ttcagcttgt 1440 ccagccgtgc ttgcaggctg gcgatttcat ccactaggta ggacatcggc gttgtaggtt 1500 gccttttgtt tctccagtgc attgggtgcc ttggcaatca aggcattgtt tgcagtctgc 1560 aattcttctt attgcgatcg cctgcgtaag gagttgagta gcgcgttcaa gccactgctc 1620 tggcgttgga ttggtcagtt gaggcaaagc attcccagcc tggtcaagct cggactgcac 1680 ttttttctcg acatttgcct tcctggcctt gtagtccgcc tccacctcag cagcggctcg 1740 ctgggcttct gcttccaatg accgggcttt attctccagc tcttgagacg tttgtttcaa 1800 gatagcgatt tgcgccttat agatatcggc gctgtacgct ttggccagct cactcatatg 1860 gcgatccagg aactctccat agaattttcg gctggccagc aactgactct ggtacatcga 1920 ctctgacttc tgaggaaagt ctgaagccgt ataaagattg gccgggcgat cctcaatgac 1980 ctttagcgat tttgctttgg catccatgag tgcatcaacg atactctttt catcgcggat 2040 gtcattggca ctgaccgctt tacctggcaa ccccgcttca ctcttgagtt catcaacctc 2100 cttcagggtt tcatttttca ggtttttctt gagttctgaa tgggacttat caagcgtact 2160 tcttagcttc ctgtactcct gcattccagt accgacatac ggacttggtc ctggtgggac 2220 aaatggtgga gtaccgtagc ttgatcgagc aggaatatac tggattatgt cacgcccacc 2280 accctgcaca tgtgtaataa ccatcgaacc aggttcgtaa tcattgacag ccatagatcg 2340 cccctacatt aatttgaaag tgtaatgtat tgagcgactc ccacctagag aaccctctcc 2400 cagtcaataa gccccaatgc atcggcaata cactgcaatc aacttcaata tcccgtgttt 2460 agatgatcca gaaggtgcgc tctctcgcct cttataatcg cgcctgcgtc aaacggtcat 2520 ttccttaacg cacacctcat ctaccccggc cagtcacgga agccgcatac cttcggttca 2580 ttaacgaact cccactttca aaattcatcc atgccgcccc ttcgcgagct tccggacaaa 2640 gccacgctga ttgcgagccc agcgtttttg attgcaagcc gctgcagctg gtcaggccgt 2700 ttccgcaacg cttgaagtcc tggccgatat accggcaggg ccagccatcg ttcgacgaat 2760 aaagccacct cagccatgat gccctttcca tccccagcgg aaccccgaca tggacgccaa 2820 agccctgctc ctcggcagcc tctgcctggc cgccccattc gccgacgcgg cgacgctcga 2880 caatgctctc tccgcctgcc tcgccgcccg gctcggtgca ccgcacacgg cggagggcca 2940 gttgcacctg ccactcaccc ttgaggcccg gcgctccacc ggcgaatgcg gctgtacctc 3000 ggcgctggtg cgatatcggc tgctggccag gggcgccagc gccgacagcc tcgtgcttca 3060 agagggctgc tcgatagtcg ccaggacacg ccgcgcacgc tgaccctggc ggcggacgcc 3120 ggcttggcga gcggccgcga actggtcgtc accctgggtt gtcaggcgcc tgactgacag 3180 gccgggctgc caccaccagg ccgagatgga cgccctgcat gtatcctccg atcggcaagc 3240 ctcccgttcg cacattcacc actctgcaat ccagttcata aatcccataa aagccctctt 3300 ccgctccccg ccagcctccc cgcatcccgc accctagacg ccccgccgct ctccgccggc 3360 tcgcccgaca agaaaaacca accgctcgat cagcctcatc cttcacccat cacaggagcc 3420 atcgcgatgc acctgatacc ccattggatc c 345119744DNAPseudomonas aeruginosa 19 gggttcagca agcgttcagg ggcggttcag taccctgtcc gtactctgca agccgtgaac 60 gacacgactc tcgcagaacg gagaaacacc atgaaagcac tcaagactct cttcatcgcc 120 accgccctgc tgggttccgc cgccggcgtc caggccgccg acaacttcgt cggcctgacc 180 tggggcgaga ccagcaacaa catccagaaa tccaagtcgc tgaaccgcaa cctgaacagc 240 ccgaacctcg acaaggtgat cgacaacacc ggcacctggg gcatccgcgc cggccagcag 300 ttcgagcagg gccgctacta cgcgacctac gagaacatct ccgacaccag cagcggcaac 360 aagctgcgcc agcagaacct gctcggcagc tacgacgcct tcctgccgat cggcgacaac 420 aacaccaagc tgttcggcgg tgccaccctc ggcctggtca agctggaaca ggacggcaag 480 ggcttcaagc gcgacagcga tgtcggctac gctgccgggc tgcaggccgg tatcctgcag 540 gagctgagca agaatgcctc gatcgaaggc ggctatcgtt acctgcgcac caacgccagc 600 accgagatga ccccgcatgg cggcaacaag ctgggctccc tggacctgca cagcagctcg 660 caattctacc tgggcgccaa ctacaagttc taaatgaccg cgcagcgccc gcgagggcat 720 gcttcgatgg ccgggccgga aggt 744202760DNAPseudomonas aeruginosa 20 ctgcagctgg tcaggccgtt tccgcaacgc ttgaagtcct ggccgatata ccggcagggc 60 cagccatcgt tcgacgaata aagccacctc agccatgatg ccctttccat ccccagcgga 120 accccgacat ggacgccaaa gccctgctcc tcggcagcct ctgcctggcc gccccattcg 180 ccgacgcggc gacgctcgac aatgctctct ccgcctgcct cgccgcccgg ctcggtgcac 240 cgcacacggc ggagggccag ttgcacctgc cactcaccct tgaggcccgg cgctccaccg 300 gcgaatgcgg ctgtacctcg gcgctggtgc gatatcggct gctggccagg ggcgccagcg 360 ccgacagcct cgtgcttcaa gagggctgct cgatagtcgc caggacacgc cgcgcacgct 420 gaccctggcg gcggacgccg gcttggcgag cggccgcgaa ctggtcgtca ccctgggttg 480 tcaggcgcct gactgacagg ccgggctgcc accaccaggc cgagatggac gccctgcatg 540 tatcctccga tcggcaagcc tcccgttcgc acattcacca ctctgcaatc cagttcataa 600 atcccataaa agccctcttc cgctccccgc cagcctcccc gcatcccgca ccctagacgc 660 cccgccgctc tccgccggct cgcccgacaa gaaaaaccaa ccgctcgatc agcctcatcc 720 ttcacccatc acaggagcca tcgcgatgca cctgataccc cattggatcc ccctggtcgc 780 cagcctcggc ctgctcgccg gcggctcgtc cgcgtccgcc gccgaggaag ccttcgacct 840 ctggaacgaa tgcgccaaag cctgcgtgct cgacctcaag gacggcgtgc gttccagccg 900 catgagcgtc gacccggcca tcgccgacac caacggccag ggcgtgctgc actactccat 960 ggtcctggag ggcggcaacg acgcgctcaa gctggccatc gacaacgccc tcagcatcac 1020 cagcgacggc ctgaccatcc gcctcgaagg cggcgtcgag ccgaacaagc cggtgcgcta 1080 cagctacacg cgccaggcgc gcggcagttg gtcgctgaac tggctggtac cgatcggcca 1140 cgagaagccc tcgaacatca aggtgttcat ccacgaactg aacgccggca accagctcag 1200 ccacatgtcg ccgatctaca ccatcgagat gggcgacgag ttgctggcga agctggcgcg 1260 cgatgccacc ttcttcgtca gggcgcacga gagcaacgag atgcagccga cgctcgccat 1320 cagccatgcc ggggtcagcg tggtcatggc ccagacccag ccgcgccggg aaaagcgctg 1380 gagcgaatgg gccagcggca aggtgttgtg cctgctcgac ccgctggacg gggtctacaa 1440 ctacctcgcc cagcaacgct gcaacctcga cgatacctgg gaaggcaaga tctaccgggt 1500 gctcgccggc aacccggcga agcatgacct ggacatcaaa cccacggtca tcagtcatcg 1560 cctgcacttt cccgagggcg gcagcctggc cgcgctgacc gcgcaccagg cttgccacct 1620 gccgctggag actttcaccc gtcatcgcca gccgcgcggc tgggaacaac tggagcagtg 1680 cggctatccg gtgcagcggc tggtcgccct ctacctggcg gcgcggctgt cgtggaacca 1740 ggtcgaccag gtgatccgca acgccctggc cagccccggc agcggcggcg acctgggcga 1800 agcgatccgc gagcagccgg agcaggcccg tctggccctg accctggccg ccgccgagag 1860 cgagcgcttc gtccggcagg gcaccggcaa cgacgaggcc ggcgcggcca acgccgacgt 1920 ggtgagcctg acctgcccgg tcgccgccgg tgaatgcgcg ggcccggcgg acagcggcga 1980 cgccctgctg gagcgcaact atcccactgg cgcggagttc ctcggcgacg gcggcgacgt 2040 cagcttcagc acccgcggca cgcagaactg gacggtggag cggctgctcc aggcgcaccg 2100 ccaactggag gagcgcggct atgtgttcgt cggctaccac ggcaccttcc tcgaagcggc 2160 gcaaagcatc gtcttcggcg gggtgcgcgc gcgcagccag gacctcgacg cgatctggcg 2220 cggtttctat atcgccggcg atccggcgct ggcctacggc tacgcccagg accaggaacc 2280 cgacgcacgc ggccggatcc gcaacggtgc cctgctgcgg gtctatgtgc cgcgctcgag 2340 cctgccgggc ttctaccgca ccagcctgac cctggccgcg ccggaggcgg cgggcgaggt 2400 cgaacggctg atcggccatc cgctgccgct gcgcctggac gccatcaccg gccccgagga 2460 ggaaggcggg cgcctggaga ccattctcgg ctggccgctg gccgagcgca ccgtggtgat 2520 tccctcggcg atccccaccg acccgcgcaa cgtcggcggc gacctcgacc cgtccagcat 2580 ccccgacaag gaacaggcga tcagcgccct gccggactac gccagccagc ccggcaaacc 2640 gccgcgcgag gacctgaagt aactgccgcg accggccggc tcccttcgca ggagccggcc 2700 ttctcggggc ctggccatac atcaggtttt cctgatgcca gcccaatcga atatgaattc 276021172DNAStaphylococcus saprophyticus 21 ttgatgaaat gcatcgatta ataaattttc atgtacgatt aaaacgtttt tacccttacc 60 ttttcgtact acctctgcct gaagttgacc acctttaaag tgattcgttg aaatccatta 120 tgctcattat taatacgatc tataaaaaca aatggaatgt gatgatcgat ga 17222155DNAStaphylococcus saprophyticus 22 gttccattga ctctgtatca cctgttgtaa cgaacatcca tatgtcctga aactccaacc 60 acaggtttga ccacttccaa tttcagacca ccaagtttga cacgtgaaga ttcatcttct 120 aatatttcgg aattaatatc atattattta aatag 15523145DNAStaphylococcus saprophyticus 23 acatagaaaa actcaaaaga tttacttttt tcaaatggaa aataagggta cacacgatat 60 ttcccgtcat cttcagttac cggtacaaca tcctctttat taacctgcac ataatctgac 120 tccgcttcac tcatcaaact actaa 14524266DNAStaphylococcus saprophyticus 24 tttcactgga attacatttc gctcattacg tacagtgaca atcgcgtcag atagtttctt 60 ctggttagct tgactcttaa caatcttgtc taaattttgt ttaattcttt gattcgtact 120 agaaatttta cttctaattc cttgtaattc ataacttgca ttatcatata aatcataagt 180 atcacatttt tgatgaatac tttgatataa atctgacaat acaggcagtt gctccattct 240 atcgttaaga atagggtaat taatag 26625845DNAHaemophilus influenzae 25 tgttaaattt ctttaacagg gattttgtta tttaaattaa acctattatt ttgtcgcttc 60 tttcactgca tctactgctt gagttgcttt ttctgaaacc gcctctttca tttcacttgc 120 tttttctgat gctgcttctt tcatttcgcc tactttttct gacgctgctt ctgttgctga 180 tttaattact tctttcgcat cttccacttt ctctgctact ttatttttca cgtctgtaga 240 aagctgctgt gctttttcct ttacttcagt cattgtatta gctgcagcat cttttgtttc 300 tgatgcgact gatgctacag tttgcttcgt atcctcaact ttttgttttg cttcttgctt 360 atcaaaacaa cctgtcacga ctaaagctga acctaaaacc aatgctaatg ttaatttttt 420 cattattttc tccatagaat aatttgattg ttacaaagcc ctattacttt gatgcagttt 480 agtttacggg aattttcata aaaagaaaaa cagtaatagt aaaactttac ctttctttaa 540 aaagattact ttataaaaaa acatctaaga tattgatttt taatagatta taaaaaacca 600 ataaaaattt tattttttgt aaaaaaaaag aatagtttat tttaaataaa ttacaggaga 660 tgcttgatgc atcaatattt ctgatttatt accatcccat aataattgag caatagttgc 720 aggataaaat gatattggat ttcgttttcc atacagttca gcaacaattt ctcccactaa 780 gggcaaatgg gaaacaatta atacagattt aacgccctcg tcttttagca cttctaaata 840 atcaa 845261598DNAHaemophilus influenzae 26 gaatagagtt gcactcaata gattcgggct ttataattgc ccagattttt atttataaca 60 aagggttcca aatgaaaaaa tttaatcaat ctctattagc aactgcaatg ttgttggctg 120 caggtggtgc aaatgcggca gcgtttcaat tggcggaagt ttctacttca ggtcttggtc 180 gtgcctatgc gggtgaagcg gcgattgcag ataatgcttc tgtcgtggca actaacccag 240 ctttgatgag tttatttaaa acggcacagt tttccacagg tggcgtttat attgattcta 300 gaattaatat gaatggtgat gtaacttctt atgctcagat aataacaaat cagattggaa 360 tgaaagcaat aaaggacggc tcagcttcac agcgtaatgt tgttcccggt gcttttgtgc 420 caaatcttta tttcgttgcg ccagtgaatg ataaattcgc gctgggtgct ggaatgaatg 480 tcaatttcgg tctaaaaagt gaatatgacg atagttatga tgctggtgta tttggtggaa 540 aaactgactt gagtgctatc aacttaaatt taagtggtgc ttatcgagta acagaaggtt 600 tgagcctagg tttaggggta aatgcggttt atgctaaagc ccaagttgaa cggaatgctg 660 gtcttattgc ggatagtgtt aaggataacc aaataacaag cgcactctca acacagcaag 720 aaccattcag agatcttaag aagtatttgc cctctaagga caaatctgtt gtgtcattac 780 aagatagagc cgcttggggc tttggctgga atgcaggtgt aatgtatcaa tttaatgaag 840 ctaacagaat tggtttagcc tatcattcta aagtggacat tgattttgct gaccgcactg 900 ctactagttt agaagcaaat gtcatcaaag aaggtaaaaa aggtaattta acctttacat 960 tgccagatta cttagaactt tctggtttcc atcaattaac tgacaaactt gcagtgcatt 1020 atagttataa atatacccat tggagtcgtt taacaaaatt acatgccagc ttcgaagatg 1080 gtaaaaaagc ttttgataaa gaattacaat acagtaataa ctctcgtgtt gcattagggg 1140 caagttataa tctttatgaa aaattgacct tacgtgcggg tattgcttac gatcaagcgg 1200 catctcgtca tcaccgtagt gctgcaattc cagataccga tcgcacttgg tatagtttag 1260 gtgcaaccta taaattcacg ccgaatttat ctgttgatct tggctatgct tacttaaaag 1320 gcaaaaaagt tcactttaaa gaagtaaaaa caataggtga caaacgtaca ttgacattga 1380 atacaactgc aaattatact tctcaagcac acgcaaatct ttacggtttg aatttaaatt 1440 atagtttcta atccgttaaa aaatttagca taataaagca caattccaca ctaagtgtgc 1500 ttttctttta taaaacaagg cgaaaaatga ccgcacttta ttacacttat tacccctcgc 1560 cagtcggacg gcttttgatt ttatctgacg gcgaaaca 1598279100DNAHaemophilus influenzae 27 gtcaaaaatt gcgtgcattc tagcgaaaaa atgggctttt gggaactgtg ggatttattt 60 aaaatcttag aaaatcttac cgcactttta agctataaag tgcggtgaaa tttagtggcg 120 tttataatgg agaattactc tggtgtaatc cattcgactg tccagcttcc agtaccttct 180 ggaactaatg tttttgtgag ataaggcaaa atttctttca tttgggtttc taatgtccaa 240 ggtggattaa ttaccaccat accgctcgca gtcattcctc gttgatcgct atctgggcga 300 acggcgagtt caatttttag aatttttcta attcccgttg cttctaaacc cttaaaaata 360 cgtttagttt gttggcgtaa tacaacagga taccaaatcg cataagtgcc agtggcaaaa 420 cgtttatagc cctcttcaat ggctttaaca acgagatcat aatcatcttt taattcataa 480 ggcggatcga tgagtactaa gcctcggcgt tcttttggcg gaagcgttgc tttgacttgt 540 tgaaagccat tgtcacattt tacggtgaca tttttgtcgt cgctaaaatt attgcgaaga 600 attggataat cgctaggatg aagctcggtc aatagtgcgc gatcttgtga gcgcaacaat 660 tccgcggcaa ttaatggaga acccgcgtaa taacgtagtt ctttgccacc ataattgagt 720 tttttgatca tttttacata acgagcaata tcttcgggta aatctgtttg atcccacagg 780 cgtccaatac cttctttata ttcccccgtt ttttctgatt catttgagga taaacgataa 840 cgccccacac cagagtgcgt atccaaataa aaaaagcctt tttctttgag tttaagattt 900 tccaaaatga gcattaaaac aatatgtttc aagacatcgg catgattgcc agcgtgaaat 960 gagtgatgat aactcagcat aatatattcc ttatatattc cttatttgtt taataacgaa 1020 ggcgagccaa ttgactcgcc cgattacaca ctaaagtgcg gtcattttta gaagagttct 1080 tgtggttgcg tcgctggcgt attgccttca ttatttaagc gttgctgtaa ctcagtagga 1140 acataataac cacgctcttg catttccgaa agataggtac gtgtcggttc tgttcccgca 1200 ataaaatatt ctttgcgccc accgtttgga gaaagcaaac ctgtcaaagt atcaatgttt 1260 ttttccacaa tttttggcgg tagcgacaat ttacgttctg gcttatcact caaagccgtt 1320 ttcatataag tgatccaagc aggcattgct gtttttgctc ctgcttctcc acgcccaagt 1380 actcgtttgt tatcatcaaa cccgacataa gttgtggtta ctaagtttgc accaaatccc 1440 gcataccaag ccacttttga actgttggta gtacctgttt taccgcctat atcgctacgt 1500 ttaatgcttt gtgcaatacg ccagctggtg cctttccagt ctaaaccttg ttcgccataa 1560 attgccgtat ttaaggcact acgaatgaga aaagcaagtt cgccactaat gacacgtggc 1620 gcatattcta ttttcgacga agcatttttt gcagcagcca ttaaatcaat cgcatcttct 1680 ttaagtgcgg tcatatttga ttgtaattct ggcagttcag gcacagtttc aggttgttga 1740 tctaattctt cgccattggt gctgtcatct gttggtttta aggcattctc gcctaaagga 1800 atattggcaa agccgttgat tttgtctttg gtttcgccat aaattacagg tatatcatta 1860 cattcaatgc aagcaatttt agggtttgca ataaataagt ctttacccgt gttatcttga 1920 attttttcaa tgatataagg ttcaatgagg aagccaccat tatcaaacac cgcataagct 1980 cgcgccattt ctaatggtgt gaaagaggct gcgccaagtg ctaaggcttc actggcaaaa 2040 tattgatcac gtttaaaacc aaaacgttgt aaaaattctg ctgtgaaatc aatacctgcc 2100 gtttggatag cacgaatagc aattatattt ttggattgac ctaatcctac gcgtaaacgc 2160 atcgggccat cataacgatc aggcgagttt ttcggttgcc acattttttg tcccggtttt 2220 tgaatagaaa tcgggctgtc ttgtaatacg cttgaaagtg ttaagccttt ttctaatgct 2280 gccgcgtaaa taaatggttt gatagaagaa cccacttgaa ctaaagactg tgtggctcga 2340 ttgaatttac tttgttcata gctaaagcca ccgaccactg cttcaatcgc accattatct 2400 gaattaagag aaactaatgc tgaatttgct gcgggaattt gtcctaattg ccattcccca 2460 ttagcacgct gatgaatcca aatttgctcg ccgactttca caggattgct tctgcctgtc 2520 caacgcattg cattggttga taaggtcatt ttttccccag aagcgagcaa tatatcagca 2580 ccgcctttta caattccaat cactgccgca ggaataaatg gctctgaatc aggtagtttg 2640 cgtagaaaac cgacaatgcg atcattgtcc caagcggctt catttttttg ccataatggc 2700 gcgccaccgc gataaccgtg acgcatatcg taatcaatca agttattacg cacagctttt 2760 tgggcttcag cttggtcttt tgaaagtaca gtggtaaata ctttataacc actggtgtaa 2820 gcattttctt cgccaaaacg acgcaccatt tcttgacgca ccatttcagt gacataatcg 2880 gctcgaaatt caaattttgc gccgtgatag ctcgccacaa tcggctcttt caatgcagca 2940 tcatattctt ctttgctgat gtatttttca tctaacatac ggcttagcac cacattgcgg 3000 cgttcttctg aacgttttaa agaataaagc gggttcattg ttgaaggtgc tttaggtaaa 3060 ccagcaataa tcgccatttc cgataaggtc aattcattca atgatttacc gaaataggtt 3120 tgtgctgccg ctgcaacacc ataagaacga tagcctaaaa agattttgtt taaataaagc 3180 tctaatattt cttgtttgtt gagagtattt tcgatttcta ccgcaagcac ggcttcacga 3240 gctttacgaa taatggtttt ttctgaggtt aagaaaaagt tacgcgctaa ttgttgagta 3300 atcgtacttg cgccttgtga tgcaccgcca ttactcactg cgacaaacaa tgcacgggca 3360 atgccgatag ggtctaatcc gtgatgatcg taaaaacgac tgtcttccgt cgctaaaaat 3420 gcgtcaatta agcgttgtgg cacatcggct aatttcactg gaatacggcg ttgctcaccc 3480 acttcgccaa ttaatttacc gtcagccgta taaatctgca ttggttgctg taattcaacg 3540 gtttttaatg tttctactga gggcaattca gattttaagt ggaaatacaa cattccgcct 3600 gctactaaac ctaaaataca taaagttaat agggtgttta atattaattt tgcgatccgc 3660 atcgtaaaat tctcgcttcg ttaatgaata ttcttgtcaa gagacctatg atttggctgt 3720 taagtataaa agattcagcc tttaaagaat aggaaagaat atgcaattct ccctgaaaaa 3780 ttaccgcact ttacaaatcg gcattcatcg taagcagagt tattttgatt ttgtgtggtt 3840 tgatgatctc gaacagccac aaagttatca aatctttgtt aatgatcgtt attttaaaaa 3900 tcgtttttta caacagctaa aaacacaata tcaagggaaa acctttcctt tgcagtttgt 3960 agcaagcatt cccgcccact taacttggtc gaaagtatta atgttgccac aagtgttaaa 4020 tgcgcaagaa tgtcatcaac aatgtaaatt tgtgattgaa aaagagctgc ctattttttt 4080 agaagaattg tggtttgatt atcgttctac cccgttaaag caaggttttc gattagaggt 4140 tactgcaatt cgtaaaagta gcgctcaaac ttatttgcaa gattttcagc catttaatat 4200 taatatattg gatgttgcgt caaatgctgt tttgcgtgca tttcaatatc tgttgaatga 4260 acaagtgcgg tcagaaaata ccttattttt atttcaagaa gatgactatt gcttggcgat 4320 ttgtgaaaga tctcagcaat cacaaatttt acaatctcac gaaaatttga ccgcacttta 4380 tgaacaattt accgaacgtt ttgaaggaca acttgaacaa gtttttgttt atcaaattcc 4440 ctcaagtcat acaccattac ccgaaaactg gcagcgagta gaaacagaac tcccttttat 4500 tgcgctgggc aacgcgctat ggcaaaaaga tttacatcaa caaaaagtgg gtggttaaat 4560 gtcgatgaat ttattgcctt ggcgtactta tcaacatcaa aagcgtttac gtcgtttagc 4620 tttttatatc gctttattta tcttgcttgc tattaattta atgttggctt ttagcaattt 4680 gattgaacaa cagaaacaaa atttgcaggc acagcaaaag tcgtttgaac aacttaatca 4740 acagcttcat aaaactacca tgcaaattga tcagttacgc attgcggtga aagttggtga 4800 agttttgaca tctattccca acgagcaagt aaaaaagagt ttacaacagc taagtgaatt 4860 accttttcaa caaggagaac tgaataaatt taaacaagat gccaataact taagcttgga 4920 aggtaacgcg caagatcaaa cagaatttga actgattcat caatttttaa agaaacattt 4980 tcccaatgtg aaattaagtc aggttcaacc tgaacaagat acattgtttt ttcactttga 5040 tgtggaacaa ggggcggaaa aatgaaagct ttttttaacg atccttttac tccttttgga 5100 aaatggctaa gtcagccttt ttatgtgcac ggtttaacct ttttattgct attaagtgcg 5160 gtgatttttc gccccgtttt agattatata gaggggagtt cacgtttcca tgaaattgaa 5220 aatgagttag cggtgaaacg ttcagaattg ttgcatcaac agaaaatttt aacctcttta 5280 caacagcagt cggaaagtcg aaaactttct ccagaactgg ctgcacaaat tattcctttg 5340 aataaacaaa ttcaacgttt agctgcgcgt aacggtttat ctcagcattt acgttgggaa 5400 atggggcaaa agcctatttt gcatttacag cttacaggtc attttgaaaa aacgaagaca 5460 tttttatccg cacttttggc taattcgtca cagctttctg taagtcggtt gcaatttatg 5520 aaacccgaag acggcccatt gcaaaccgag atcatttttc agctagataa ggaaacaaaa 5580 tgaaacattg gtttttcctg attatattat tttttatgaa ttgcagttgg ggacaagatc 5640 ctttcgataa aacacagcgt aaccgttctc agtttgataa cgcacaaaca gtaatggagc 5700 aaacagaaat aatttcctca gatgtgccta ataatctatg cggagcggat gaaaatcgcc 5760 aagcggctga aattcctttg aacgctttaa aattggtggg ggtagtgatt tctaaagata 5820 aagcctttgc cttgttgcaa gatcaaggtt tgcaagttta cagcgtttta gagggcgttg 5880 atgtggctca agagggctat attgtagaaa aaatcaacca aaacaatgtt caatttatgc 5940 gtaagctagg agagcaatgt gatagtagtg aatggaaaaa attaagtttt taaaggaaga 6000 ttatgaagaa atatttttta aagtgcggtt attttttagt atgtttttgt ttgccattaa 6060 tcgtttttgc taatcctaaa acagataacg aacgtttttt tattcgttta tcgcaagcac 6120 ctttagctca aacactggag caattagctt ttcaacaaga tgtgaattta gtgattggag 6180 atatattgga aaacaagatc tctttgaaat taaacaatat tgatatgcca cgtttgctac 6240 aaataatcgc aaaaagtaag catcttactt tgaataaaga tgatgggatt tattatttaa 6300 acggcagtca atctggcaaa ggtcaggttg caggaaatct tacgacaaat gaaccgcact 6360 tagtgagtca cacggtaaaa ctccattttg ctaaagcttc tgaattaatg aaatccttaa 6420 caacaggaag tggctctttg ctttctcccg ctgggagcat tacctttgat gatcgcagta 6480 atttgctggt tattcaggat gaacctcgtt ctgtgcaaaa tatcaaaaaa ctgattgctg 6540 aaatggataa gcctattgaa cagatcgcta ttgaagcgcg aattgtgaca attacggatg 6600 agagtttgaa agaacttggc gttcggtggg ggatttttaa tccaactgaa aatgcaagac 6660 gagttgcggg cagccttaca ggcaatagct ttgaaaatat tgcggataat cttaatgtaa 6720 attttgcgac aacgacgaca cctgctggct ctatagcatt acaagtcgcc aaaattaatg 6780 ggcgattgct tgatttagaa ttgagtgcgt tggagcgtga aaataatgta gaaattattg 6840 caagccctcg cttactcact accaataaga aaagtgcgag cattaaacag gggacagaaa 6900 ttccttacat cgtgagtaat actcgtaacg atacgcaatc tgtggaattt cgtgaggcgg 6960 tgcttggttt ggaagtgacg ccacatattt ctaaagataa caatatctta cttgatttat 7020 tggtaagtca aaattcccct ggttctcgtg tcgcttatgg acaaaatgag gtggtttcta 7080 ttgataaaca agaaattaat actcaggttt ttgccaaaga tggggaaacc attgtgcttg 7140 gcggcgtatt tcacgataca atcacgaaaa gcgaagataa agtgccattg cttggcgata 7200 tacccgttat taaacgatta tttagcaaag aaagtgaacg acatcaaaaa cgtgagctag 7260 tgattttcgt cacgccacat attttaaaag caggagaaaa cgttagaggc gttgaaacaa 7320 aaaagtgagg gtaaaaaata actttttaaa tgatgaattt ttttaatttt cgctgtatcc 7380 actgtcgtgg caatcttcat atcgcaaaaa atgggttatg ttcaggttgc caaaaacaaa 7440 ttaaatcttt tccttattgc ggtcattgtg gttcggaatt gcaatattat gcgcagcatt 7500 gtgggaattg tcttaaacaa gaaccaagtt gggataagat ggtcattatt gggcattata 7560 ttgaacctct ttcgatattg attcagcgtt ttaaatttca aaatcaattt tggattgacc 7620 gcactttagc tcggctttta tatcttgcgg tacgtgatgc taaacgaacg catcaactta 7680 aattgccaga ggcaatcatt ccagtgcctt tatatcattt tcgtcagtgg cgacggggtt 7740 ataatcaggc agatttatta tctcagcaat taagtcgttg gctggatatt cctaatttga 7800 acaatatcgt aaagcgtgtg aaacacacct atactcaacg tggtttgagt gcaaaagatc 7860 gtcgtcagaa tttaaaaaat gccttttctc ttgctgtttc gaaaaatgaa tttccttatc 7920 gtcgtgttgc gttggtggat gatgtgatta ctactggttc tacactcaat gaaatctcaa 7980 aattgttgcg aaaattaggt gtggaggaga ttcaagtgtg ggggctggca cgagcttaat 8040 ataaagcact ggaaaaaaaa gcgcgataag cgtattattc ccgatacttt ctctcaagta 8100 tttaggacat aattatggaa caagcaaccc agcaaatcgc tatttctgat gccgcacaag 8160 cgcattttcg aaaactttta gacacccaag aagaaggaac gcatattcgt attttcgcgg 8220 ttaatcctgg tacgcctaat gcggaatgtg gcgtatctta ttgccccccg aatgccgtgg 8280 aagaaagcga tattgaaatg aaatataata ctttttctgc atttattgat gaagtgagtt 8340 tgcctttctt agaagaagca gaaattgatt atgttaccga agagcttggt gcgcaactga 8400 ccttaaaagc accgaatgcc aaaatgcgta aggtggctga tgatgcgcca ttgattgaac 8460 gtgttgaata tgtaattcaa actcaaatta acccacagct tgcaaatcac ggtggacgta 8520 taaccttaat tgaaattact gaagatggtt acgcagtttt acaatttggt ggtggctgta 8580 acggttgttc aatggtggat gttacgttaa aagatggggt agaaaaacaa cttgttagct 8640 tattcccgaa tgaattaaaa ggtgcaaaag atataactga gcatcaacgt ggcgaacatt 8700 cttattatta gtgagttata aaagaagatt tataatgacc gcacttttga aagtgcggtt 8760 atttttatgg agaaaaaatg aaaatacttc aacaagatga ttttggttat tggttgctta 8820 cacaaggttc taatctgtat ttagtgaata atgaattgcc ttttggtatc gctaaagata 8880 ttgatttgga aggattgcag gcaatgcaaa ttggggaatg gaaaaattat ccgttgtggc 8940 ttgtggctga gcaagaaagt gatgaacgag aatatgtgag tttgagtaac ttgctttcac 9000 tgccagagga tgaattccat atattaagcc gaggtgtgga aattaatcat tttctgaaaa 9060 cccataaatt ctgtggaaag tgcggtcata aaacacaaca 910028525DNAMoraxella catarrhalis 28 aaaaatcgac tgccgtcatt ttcaaccacc acatagctca tattcgcaag ccaatgtatt 60 gaccgttggg aataataaca gccccaaaac aatgaaacat atggtgatga gccaaacata 120 ctttcctgca gattttggaa tcatatcgcc atcagcacca gtatggtttg accagtattt 180 aacgccatag acatgtgtaa aaaaattaaa taacggtgca agcatgagac caacggcacc 240 tgatgtacct tgtacgatga cctcacctgc tgtggcaacc ataccaagtc cattgcctgt 300 gatatttttg cgaaaagaca aacttaccac acagaccaag ccgatgattg agatgacaaa 360 ataaaaccaa tccaaatgcg tgtgagctgt tgtggtccaa aatccagtaa atagtgcaat 420 aaatccgcaa acaaaccaaa gtagcaccca gcttgttgtc caatcttttt taccaaagcc 480 tgtgatgtta tctaaaatat caattttcat cagattttcc ctaat 52529466DNAMoraxella catarrhalis 29 taatgataac cagtcaagca agctcaaatc agggtcagcc tgttttgagc tttttatttt 60 ttgatcatca tgcttaagat tcactctgcc atttttttac aacctgcacc acaagtcatc 120 atcgcatttg caaaaatggt acaaacaagc cgtcagcgac ttaaacaaaa aaaggctcaa 180 tctgcgtgtg tgcgttcact tttacaaatc accatgcacc gctttgacat tgttggtgaa 240 tttcatgacc atgcacaccc ttattatatt aactcaaata aaatacgcta ctttgtcagc 300 tttagccatt cagataatca agtcgctctc atcatcagct taacaccttg tgccattgac 360 atagaagtta acgatattaa atacagtgtg gttgaacgat actttcatcc caatgaaatt 420 tatctactta ctcaatttag ctctactgat aggcaacagc ttatta 46630631DNAStreptococcus pneumoniae 30 gatctttgat tttcattgag tattactctc tcttgtcact tctttctatt ttaccataaa 60 gtccagcctt tgaagaactt ttactagaag acaaggggct tctgtctcta tttgccatct 120 taggcatcaa aaaagagggg tcatccctct ttacgaattc aatgctacta gggtatccaa 180 atactggttg ttgatgactg ccaaaatata ggtatctgct ttcaagaggt catctggtcc 240 aaattcaaca tccaatgggg aattttcctg ctctcggaaa cccaaaatat tcagattgta 300 tttgccacgg aggtctaatt tacttcagac tttgacctgc ccaagactga ggaattttca 360 tctccacgat agacacattt ttatccaact gaaagacatc aacactatta tgaaaagaat 420 ggtctgtgct agagactgcc ccatttcata ctctggcgag ataaccgagt cagctccaat 480 cttttctagc actttcttag cggtctgact tttgacctta gcaataacag tcggtacccc 540 caaactctta cagtgcataa ccgcaagcac actcgactcc agattttcac ctgtcgcgac 600 tacaacggta tcgcaggtat caatccctgc t 631313754DNAStreptococcus pneumoniae 31 ccaatatttt ggtcagcata gtgttctttt tcagtggtaa cagcttgcaa tacttgagca 60 gaaatggcag atttatcaag gaaaaagtta acgtaaggtc ctgttgcgac aactttttca 120 aaggcttggc tgttcatttt ttcagccagt tcagccgcaa tcatttgtgg tgctttacgt 180 tcgacttttg caagagaaaa agcagggaaa gcaatgtctc ccatttctga gtttttaggg 240 gtttccagta actttaaaat agcctcttgg tccaggctat caatgatgct agataattcg 300 ctagcaatca attcttttgt attcattaag agctcctttt tggacttttc tactatttta 360 tcacaatttt aaagaaagaa gaaaaaattt ttgaaatctc ctgttttttt ggtataatat 420 ggttataaat atagttataa atatagttat aaatatgcac gcaagaggat tttatgagaa 480 aaagagatcg tcatcagtta ataaaaaaaa tgattactga ggagaaatta agtacacaaa 540 aagaaattca agatcggttg gaggcgcaca atgtttgtgt gacgcagaca accttgtctc 600 gtgatttgcg cgaaatcggc ttgaccaagg tcaagaaaaa tgatatggtg tattatgtac 660 tagtaaatga gacagaaaag attgatttgg tggaattttt gtctcatcat ttagaaggtg 720 ttgcaagagc agagtttacc ttggtgcttc ataccaaatt gggagaagcc tctgttttgg 780 caaatattgt agatgtaaac aaggatgaat ggattttagg aacagttgct ggtgccaata 840 ccttattggt tatttgtcga gatcagcacg ttgccaaact catggaagat cgtttgctag 900 atttgatgaa agataagtaa ggtcttggga gttgctctca agacttattt ttgaaaagga 960 gagacagaaa atggcgatag aaaagctatc acccggcatg caacagtatg tggatattaa 1020 aaagcaatat ccagatgctt ttttgctctt tcggatgggt gatttttatg aattatttta 1080 tgaggatgcg gtcaatgctg cgcagattct ggaaatttcc ttaacgagtc gcaacaagaa 1140 tgccgacaat ccgatcccta tggcgggtgt tccctatcat tctgcccaac agtatatcga 1200 tgtcttgatt gagcagggtt ataaggtggc tatcgcagag cagatggaag atcctaaaca 1260 agcagttggg gttgttaaac gagaggttgt tcaggtcatt acgccaggga cagtggtcga 1320 tagcagtaag ccggacagtc agaataattt tttggtttcc atagaccgcg aaggcaatca 1380 atttggccta gcttatatgg atttggtgac gggtgacttt tatgtgacag gtcttttgga 1440 tttcacgctg gtttgtgggg aaatccgtaa cctcaaggct cgagaagtgg tgttgggtta 1500 tgacttgtct gaggaagaag aacaaatcct cagccgccag atgaatctgg tactctctta 1560 tgaaaaagaa agctttgaag accttcattt attggatttg cgattggcaa cggtggagca 1620 aacggcatct agtaagctgc tccagtatgt tcatcggact cagatgaggg aattgaacca 1680 cctcaaacct gttatccgct acgaaattaa ggatttcttg cagatggatt atgcgaccaa 1740 ggctagtctg gatttggttg agaatgctcg ctcaggtaag aaacaaggca gtcttttctg 1800 gcttttggat gaaaccaaaa cggctatggg gatgcgtctc ttgcgttctt ggattcatcg 1860 ccccttgatt gataaggaac gaatcgtcca acgtcaagaa gtagtgcagg tctttctcga 1920 ccatttcttt gagcgtagtg acttgacaga cagtctcaag ggtgtttatg acattgagcg 1980 cttggctagt cgtgtttctt ttggcaaaac caatccaaag gatctcttgc agttggcgac 2040 taccttgtct agtgtgccac ggattcgtgc gattttagaa gggatggagc aacctactct 2100 agcctatctc atcgcacaac tggatgcaat ccctgagttg gagagtttga ttagcgcagc 2160 gattgctcct gaagctcctc atgtgattac agatggggga attatccgga ctggatttga 2220 tgagacttta gacaagtatc gttgcgttct cagagaaggg actagctgga ttgctgagat 2280 tgaggctaag gagcgagaaa actctggtat cagcacgctc aagattgact acaataaaaa 2340 ggatggctac tattttcatg tgaccaattc gcaactggga aatgtgccag cccacttttt 2400 ccgcaaggcg acgctgaaaa actcagaacg ctttggaacc gaagaattag cccgtatcga 2460 gggagatatg cttgaggcgc gtgagaagtc agccaacctc gaatacgaaa tatttatgcg 2520 cattcgtgaa gaggtcggca agtacatcca gcgtttacaa gctctagccc aaggaattgc 2580 gacggttgat gtcttacaga gtctggcggt tgtggctgaa acccagcatt tgattcgacc 2640 tgagtttggt gacgattcac aaattgatat ccggaaaggg cgccatgctg tcgttgaaaa 2700 ggttatgggg gctcagacct atattccaaa tacgattcag atggcagaag ataccagtat 2760 tcaattggtt acagggccaa acatgagtgg gaagtctacc tatatgcgtc agttagccat 2820 gacggcggtt atggcccagc tgggttccta tgttcctgct gaaagcgccc atttaccgat 2880 ttttgatgcg atttttaccc gtatcggagc agcagatgac ttggtttcgg gtcagtcaac 2940 ctttatggtg gagatgatgg aggccaataa tgccatttcg catgcgacca agaactctct 3000 cattctcttt gatgaattgg gacgtggaac tgcaacttat gacgggatgg ctcttgctca 3060 gtccatcatc gaatatatcc atgagcacat cggagctaag accctctttg cgacccacta 3120 ccatgagttg actagtctgg agtctagttt acaacacttg gtcaatgtcc acgtggcaac 3180 tttggagcag gatgggcagg tcaccttcct tcacaagatt gaaccgggac cagctgataa 3240 atcctacggt atccatgttg ccaagattgc tggcttgcca gcagaccttt tagcaagggc 3300 ggataagatt ttgactcagc tagagaatca aggaacagag agtcctcctc ccatgagaca 3360 aactagtgct gtcactgaac agatttcact ctttgatagg gcagaagagc atcctatcct 3420 agcagaatta gctaaactgg atgtgtataa tatgacacct atgcaggtta tgaatgtctt 3480 agtagagtta aaacagaaac tataaaacca agactcacta gttaatctag ctgtatcaag 3540 gagacttctt tgacaattct ccactttttt gctagaataa catcacacaa acagaatgaa 3600 aagggctgac gcattgtcgc tcccttttgt ctatttttta aggagaaagt atgctgattc 3660 agaaaataaa aacctacaag tggcaggccc tgcttcgctc ctgatgacag gcttgatggt 3720 tgctagttca cttctgcaac cgcgttatct gcag 3754321337DNAStreptococcus pyogenes 32 aacaaaataa aagaacttac ctattttcca tccaaaatgt ttagcaatca tcatctgcaa 60 ggcaacgtat tgcatggcat tgatgtgatg agcaactaat atgtcattag aacgttgcgt 120 caaactagca tctaaataaa gatcgaaatg cagttatcaa aaatgcaagc tcctatcggc 180 ccttgtttta attattactc acattgcctt aatgtattta cttgcttatt attaactttt 240 ttgctaagtt agtagcgtca gttattcatt gaaaggacat tattatgaaa attcttgtaa 300 caggctttga tccctttggc ggcgaagcta ttaatcctgc ccttgaagct atcaagaaat 360 tgccagcaac cattcatgga gcagaaatca aatgtattga agttccaacg gtttttcaaa 420 aatctgccga tgtgctccag cagcatatcg aaagctttca acctgatgca gtcctttgta 480 ttgggcaagc tggtggccgg actggactaa cgccagaacg cgttgccatt aatcaagacg 540 atgctcgcat tcctgataac gaagggaatc agcctattga tacacctatt cgtgcagatg 600 gtaaagcagc ttatttttca accttgccaa tcaaagcgat ggttgctgcc attcatcagg 660 ctgggcttcc tgcttctgtt tctaatacag ctggtacctt tgtttgcaat catttgatgt 720 atcaagccct ttacttagtg gataaatatt gtccaaatgc caaagctggg tttatgcata 780 ttccctttat gatggaacag gttgttgata aacctaatac agctgccatg aacctcgatg 840 atattacaag aggaattgag gctgctattt ttgccattgt cgatttcaaa gatcgttccg 900 atttaaaacg tgtagggggc gctactcact gactgtgacg ctactaaacc tattttaaaa 960 aaacagagat atgaactaac tctgtttttt ttgtgctaaa aatgaaagac ctagggaaac 1020 ttttcatcgg tctttctcaa ttgtcatctt aatctaatac tacttctaac atcagcgggt 1080 atagtttgcc agtaattaag aaacgttgtt gatctaaatg agcaatccca ttcaaaacat 1140 taaggtcagg gtaatgggac ttatcaagat ttaaggcttt taacaaagga ctaatatcat 1200 aggtggctac cacctttcca gaatcaggtt ggagtttgac aatagtattg gtttgccaaa 1260 tattggcata gagataacca tctacatact ctaattcgtt aagcattgag atagggacac 1320 tttctatagc aactagt 1337331837DNAStreptococcus pyogenes 33 tcatgtttga cagcttatca tcgataagct tacttttcga atcaggtcta tccttgaaac 60 aggtgcaaca tagattaggg catggagatt taccagacaa ctatgaacgt atatactcac 120 atcacgcaat cggcaattga tgacattgga actaaattca atcaatttgt tactaacaag 180 caactagatt gacaactaat tctcaacaaa cgttaattta acaacattca agtaactccc 240 accagctcca tcaatgctta ccgtaagtaa tcataactta ctaaaacctt gttacatcaa 300 ggttttttct ttttgtcttg ttcatgagtt accataactt tctatattat tgacaactaa 360 attgacaact cttcaattat ttttctgtct actcaaagtt ttcttcattt gatatagtct 420 aattccacca tcacttcttc cactctctct accgtcacaa cttcatcatc tctcactttt 480 tcgtgtggta acacataatc aaatatcttt ccgtttttac gcactatcgc tactgtgtca 540 cctaaaatat accccttatc aatcgcttct ttaaactcat ctatatataa catatttcat 600 cctcctacct atctattcgt aaaaagataa aaataactat tgtttttttt gttattttat 660 aataaaatta ttaatataag ttaatgtttt ttaaaaatat acaattttat tctatttata 720 gttagctatt ttttcattgt tagtaatatt ggtgaattgt aataaccttt ttaaatctag 780 aggagaaccc agatataaaa tggaggaata ttaatggaaa acaataaaaa agtattgaag 840 aaaatggtat tttttgtttt agtgacattt cttggactaa caatctcgca agaggtattt 900 gctcaacaag accccgatcc aagccaactt cacagatcta gtttagttaa aaaccttcaa 960 aatatatatt ttctttatga gggtgaccct gttactcacg agaatgtgaa atctgttgat 1020 caacttttat ctcacgattt aatatataat gtttcagggc caaattatga taaattaaaa 1080 actgaactta agaaccaaga gatggcaact ttatttaagg ataaaaacgt tgatatttat 1140 ggtgtagaat attaccatct ctgttattta tgtgaaaatg cagaaaggag tgcatgtatc 1200 tacggagggg taacaaatca tgaagggaat catttagaaa ttcctaaaaa gatagtcgtt 1260 aaagtatcaa tcgatggtat ccaaagccta tcatttgata ttgaaacaaa taaaaaaatg 1320 gtaactgctc aagaattaga ctataaagtt agaaaatatc ttacagataa taagcaacta 1380 tatactaatg gaccttctaa atatgaaact ggatatataa agttcatacc taagaataaa 1440 gaaagttttt ggtttgattt tttccctgaa ccagaattta ctcaatctaa atatcttatg 1500 atatataaag ataatgaaac gcttgactca aacacaagcc aaattgaagt ctacctaaca 1560 accaagtaac tttttgcttt tggcaacctt acctactgct ggatttagaa attttattgc 1620 aattctttta ttaatgtaaa aaccgctcat ttgatgagcg gttttgtctt atctaaagga 1680 gctttacctc ctaatgctgc aaaattttaa atgttggatt tttgtatttg tctattgtat 1740 ttgatgggta atcccatttt tcgacagaca tcgtcgtgcc acctctaaca ccaaaatcat 1800 agacaggagc ttgtagctta gcaactattt tatcgtc 183734841DNAStreptococcus pneumoniae 34 gatcaatatg tccaagaaac cacatgttcc taagacaaga gctaacagac tggccgtcaa 60 taatagtatt gttctttttt tcatcattac tccttaacta gtgtttaact gattaattag 120 ccagtaaata gtttatcttt atttacacta tctgttaaga tatagtaaaa tgaaataaga 180 acaggacagt caaatcgatt tctaacaatg ttttagaagt agaggtatac tattctaatt 240 tcaatctact atattttgca cattttcata aaaaaaatga gaactagaac tcacattctg 300 ctctcatttt tcgttttccc gttctcctat cctgttttta ggagttagaa aatgctgcta 360 cctttactta ctctccttta ataaagccaa tagtttttca gcttctgcca taatagtatt 420 gttgtcctgg gtgccaaata gtaaattatt ttttaatcct gtgagagtct ctttggcatt 480 ggacttgata attggattct ggatttttcc aagtaaatct tcagcctctc tcagttttct 540 taacctttca gtctcgacct gaggttcttc tgattcctct ggtgattctt ctggtgattc 600 ttcttctggt tcctctgttg gttttggaga ctctggtttc tcgctttgcg gtttctcttc 660 tcgaggggtt tcttcctcag gtttttctgt ctgaggtttc tcctcgtttg gtttttccgt 720 ttgattggta tcagcttgac catttttgtt tctttgaaca tggtcgctag cgttaccaaa 780 accattatct gaatgcgacg ttcgtttgga tgttcgacat agtacttgac agtcgccaaa 840 a 841354500DNAStreptococcus pneumoniae 35 gatcaggaca gtcaaatcga tttctaacaa tgttttagaa gtagatgtgt actattctag 60 tttcaatcta ttatatttat agaatttttt gttgctagat ttgtcaaatt gcttaaaata 120 atttttttca gaaagcaaaa gccgatacct atcgagtagg gtagttcttg ctatcgtcag 180 gcttgtctgt aggtgttaac acttttcaaa aatctcttca aacaacgtca gctttgcctt 240 gccgtatata tgttactgac ttcgtcagtt ctatctgcca cctcaaaacg gtgttttgag 300 ctgacttcgt cagttctatc cacaacctca aaacagtgtt ttgagctgac ttcgtcagtt 360 ctatccacaa cctcaaaaca gtgttttgag ctgactttgt cagtcttatc tacaacctca 420 aaacagtgtt ttgagcatca tgcggctagc ttcttagttt gctctttgat tttcattgag 480 tataaaaaca gatgagtttc tgttttcttt ttatggacta taaatgttca gctgaaacta 540 ctttcaagga cattattata taaaagaatt ttttgaaact aaaatctact atattacact 600 atattgaaag cgttttaaaa atgaggtata ataaatttac taacacttat aaaaagtgat 660 agaatctatc tttatgtata tttaaagata gattgctgta aaaatagtag tagctatgcg 720 aaataacaga tagagagaag ggattgaagc ttagaaaagg ggaataatat gatatttaag 780 gcattcaaga caaaaaagca gagaaaaaga caagttgaac tacttttgac agtttttttc 840 gacagttttc tgattgattt atttcttcac ttatttggga ttgtcccctt taagctggat 900 aagattctga ttgtgagctt gattatattt cccattattt ctacaagtat ttatgcttat 960 gaaaagctat ttgaaaaagt gttcgataag gattgagcag gaagtatggt gtaaatagca 1020 taagctgatg tccatcattt gcttataaag agatatttta gtttaattgc agcggtgtcc 1080 tggtagataa actagattgg caggagtctg attggagaaa ggagagggga aatttggcac 1140 caatttgaga tagtttgttt agttcatttt tgtcatttaa atgaactgta gtaaaagaaa 1200 gttaataaaa gacaaactaa gtgcattttc tggaataaat gtcttatttc agaaatcggg 1260 atatagatat agagaggaac agtatgaatc ggagtgttca agaacgtaag tgtcgttata 1320 gcattaggaa actatcggta ggagcggttt ctatgattgt aggagcagtg gtatttggaa 1380 cgtctcctgt tttagctcaa gaaggggcaa gtgagcaacc tctggcaaat gaaactcaac 1440 tttcggggga gagctcaacc ctaactgata cagaaaagag ccagccttct tcagagactg 1500 aactttctgg caataagcaa gaacaagaaa ggaaagataa gcaagaagaa aaaattccaa 1560 gagattacta tgcacgagat ttggaaaatg tcgaaacagt gatagaaaaa gaagatgttg 1620 aaaccaatgc ttcaaatggt cagagagttg atttatcaag tgaactagat aaactaaaga 1680 aacttgaaaa cgcaacagtt cacatggagt ttaagccaga tgccaaggcc ccagcattct 1740 ataatctctt ttctgtgtca agtgctacta aaaaagatga gtacttcact atggcagttt 1800 acaataatac tgctactcta gaggggcgtg gttcggatgg gaaacagttt tacaataatt 1860 acaacgatgc acccttaaaa gttaaaccag gtcagtggaa ttctgtgact ttcacagttg 1920 aaaaaccgac agcagaacta cctaaaggcc gagtgcgcct ctacgtaaac ggggtattat 1980 ctcgaacaag tctgagatct ggcaatttca ttaaagatat gccagatgta acgcatgtgc 2040 aaatcggagc aaccaagcgt gccaacaata cggtttgggg gtcaaatcta cagattcgga 2100 atctcactgt gtataatcgt gctttaacac cagaagaggt acaaaaacgt agtcaacttt 2160 ttaaacgctc agatttagaa aaaaaactac ctgaaggagc ggctttaaca gagaaaacgg 2220 acatattcga aagcgggcgt aacggtaaac caaataaaga tggaatcaag agttatcgta 2280 ttccagcact tctcaagaca gataaaggaa ctttgatcgc aggtgcagat gaacgccgtc 2340 tccattcgag tgactggggt gatatcggta tggtcatcag acgtagtgaa gataatggta 2400 aaacttgggg tgaccgagta accattacca acttacgtga caatccaaaa gcttctgacc 2460 catcgatcgg ttcaccagtg aatatcgata tggtgttggt tcaagatcct gaaaccaaac 2520 gaatcttttc tatctatgac atgttcccag aagggaaggg aatctttgga atgtcttcac 2580 aaaaagaaga agcctacaaa aaaatcgatg gaaaaaccta tcaaatcctc tatcgtgaag 2640 gagaaaaggg agcttatacc attcgagaaa atggtactgt ctatacacca gatggtaagg 2700 cgacagacta tcgcgttgtt gtagatcctg ttaaaccagc ctatagcgac aagggggatc 2760 tatacaaggg taaccaatta ctaggcaata tctacttcac aacaaacaaa acttctccat 2820 ttagaattgc caaggatagc tatctatgga tgtcctacag tgatgacgac gggaagacat 2880 ggtcagcgcc tcaagatatt actccgatgg tcaaagccga ttggatgaaa ttcttgggtg 2940 taggtcctgg aacaggaatt gtacttcgga atgggcctca caagggacgg attttgatac 3000 cggtttatac gactaataat gtatctcact taaatggctc gcaatcttct cgtatcatct 3060 attcagatga tcatggaaaa acttggcatg ctggagaagc ggtcaacgat aaccgtcagg 3120 tagacggtca aaagatccac tcttctacga tgaacaatag acgtgcgcaa aatacagaat 3180 caacggtggt acaactaaac aatggagatg ttaaactctt tatgcgtggt ttgactggag 3240 atcttcaggt tgctacaagt aaagacggag gagtgacttg ggagaaggat atcaaacgtt 3300 atccacaggt taaagatgtc tatgttcaaa tgtctgctat ccatacgatg cacgaaggaa 3360 aagaatacat catcctcagt aatgcaggtg gaccgaaacg tgaaaatggg atggtccact 3420 tggcacgtgt cgaagaaaat ggtgagttga cttggctcaa acacaatcca attcaaaaag 3480 gagagtttgc ctataattcg ctccaagaat taggaaatgg ggagtatggc atcttgtatg 3540 aacatactga aaaaggacaa aatgcctata ccctatcatt tagaaaattt aattgggact 3600 ttttgagcaa agatctgatt tctcctaccg aagcgaaagt gaagcgaact agagagatgg 3660 gcaaaggagt tattggcttg gagttcgact cagaagtatt ggtcaacaag gctccaaccc 3720 ttcaattggc aaatggtaaa acagcacgct tcatgaccca gtatgataca aaaaccctcc 3780 tatttacagt ggattcagag gatatgggtc aaaaagttac aggtttggca gaaggtgcaa 3840 ttgaaagtat gcataattta ccagtctctg tggcgggcac taagctttcg aatggaatga 3900 acggaagtga agctgctgtt catgaagtgc cagaatacac aggcccatta gggacatccg 3960 gcgaagagcc agctccaaca gtcgagaagc cagaatacac aggcccacta gggacatccg 4020 gcgaagagcc agccccgaca gtcgagaagc cagaatacac aggcccacta gggacagctg 4080 gtgaagaagc agctccaaca gtcgagaagc cagaatttac agggggagtt aatggtacag 4140 agccagctgt tcatgaaatc gcagagtata agggatctga ttcgcttgta actcttacta 4200 caaaagaaga ttatacttac aaagctcctc ttgctcagca ggcacttcct gaaacaggaa 4260 acaaggagag tgacctccta gcttcactag gactaacagc tttcttcctt ggtctgttta 4320 cgctagggaa aaagagagaa caataagaga agaattctaa acatttgatt ttgtaaaaat 4380 agaaggagat agcaggtttt caagcctgct atcttttttt gatgacattc aggctgatac 4440 gaaatcataa gaggtctgaa actactttca gagtagtctg ttctataaaa tatagtagat 450036705DNAStaphylococcus epidermidis 36 gatccaagct tatcgatatc atcaaaaagt tggcgaacct tttcaaattt tggttcaaat 60 tcttgagatg tatagaattc aaaatattta ccatttgcat agtctgattg ctcaaagtct 120 tgatactttt ctccacgctc ttttgcaatt tccattgaac gttcgatgga ataatagttc 180 ataatcataa agaatatatt agcaaagtct tttgcttctt cagattcata gccaatttta 240 tttttagcta gataaccatg taagttcatt actcctagtc caacagaatg tagttcacta 300 ttcgcttttt ttacacctgg tgcattttga atatttgctt catcacttac aactgtaaga 360 gcatccatac ctgtgaacac agaatctctg aatttacctg attccataac attcactata 420 ttcaatgagc ctaagttaca tgaaatatct cttttaattt catcttcaat tccatagtcg 480 ttaattactg atgtctcttg taattggaaa atttcagtac ataaattact cattttaatt 540 tgcccaatat ttgaattcgc atgtactttg tttgcattat ctttaaacat aagatatgga 600 taaccagact gtaattgtgt ttgtgcaatc atatttaaca tttcacgtgc gtcttttttc 660 tttttatcga tttcgaaccc ggggtaccga attcctcgag tctag 70537442DNAStaphylococcus aureus 37 gatcaatctt tgtcggtaca cgatattctt cacgactaaa taaacgctca ttcgcgattt 60 tataaatgaa tgttgataac aatgttgtat tatctactga aatctcatta cgttgcatcg 120 gaaacattgt gttctgtatg taaaagccgt cttgataatc tttagtagta ccgaagctgg 180 tcatacgaga gttatatttt ccagccaaaa cgatattttt ataatcatta cgtgaaaaag 240 gtttcccttc attatcacac aaatatttta gcttttcagt ttctatatca actgtagctt 300 ctttatccat acgttgaata attgtacgat tctgacgcac catcttttgc acacctttaa 360 tgttatttgt tttaaaagca tgaataagtt tttcaacaca acgatgtgaa tcttctaaga 420 agtcaccgta aaatgaagga tc 4423820DNAEnterococcus faecalis 38 gcaatacagg gaaaaatgtc 203920DNAEnterococcus faecalis 39 cttcatcaaa caattaactc 204020DNAEnterococcus faecalis 40 gaacagaaga agccaaaaaa 204120DNAEnterococcus faecalis 41 gcaatcccaa ataatacggt 204219DNAEscherichia coli 42 gctttccagc gtcatattg 194319DNAEscherichia coli 43 gatctcgaca aaatggtga 194425DNAEscherichia coli 44 cacccgcttg cgtggcaagc tgccc 254525DNAEscherichia coli 45 cgtttgtgga ttccagttcc atccg 254617DNAEscherichia coli 46 tcacccgctt gcgtggc 174719DNAEscherichia coli 47 ggaactggaa tccacaaac 194825DNAEscherichia coli 48 tgaagcactg gccgaaatgc tgcgt 254925DNAEscherichia coli 49 gatgtacagg attcgttgaa ggctt 255025DNAEscherichia coli 50 tagcgaaggc gtagcagaaa ctaac 255125DNAEscherichia coli 51 gcaacccgaa ctcaacgccg gattt 255225DNAEscherichia coli 52 atacacaagg gtcgcatctg cggcc 255326DNAEscherichia coli 53 tgcgtatgca ttgcagacct tgtggc 265425DNAEscherichia coli 54 gctttcactg gatatcgcgc ttggg 255519DNAEscherichia coli 55 gcaacccgaa ctcaacgcc 195619DNAEscherichia coli 56 gcagatgcga cccttgtgt 195723DNAKlebsiella pneumoniae 57 gtggtgtcgt tcagcgcttt cac 235825DNAKlebsiella pneumoniae 58 gcgatattca caccctacgc agcca 255926DNAKlebsiella pneumoniae 59 gtcgaaaatg ccggaagagg tatacg 266026DNAKlebsiella pneumoniae 60 actgagctgc agaccggtaa aactca 266119DNAKlebsiella pneumoniae 61 gacagtcagt tcgtcagcc 196219DNAKlebsiella pneumoniae 62 cgtagggtgt gaatatcgc 196326DNAKlebsiella pneumoniae 63 cgtgatggat attcttaacg aagggc 266423DNAKlebsiella pneumoniae 64 accaaactgt tgagccgcct gga 236523DNAKlebsiella pneumoniae 65 gtgatcgccc ctcatctgct act 236626DNAKlebsiella pneumoniae 66 cgcccttcgt taagaatatc catcac 266719DNAKlebsiella pneumoniae 67 tcgcccctca tctgctact 196819DNAKlebsiella pneumoniae 68 gatcgtgatg gatattctt 196925DNAKlebsiella pneumoniae 69 caggaagatg ctgcaccggt tgttg 257025DNAProteus mirabilis 70 tggttcactg actttgcgat gtttc 257125DNAProteus mirabilis 71 tcgaggatgg catgcactag aaaat 257230DNAProteus mirabilis 72 cgctgattag gtttcgctaa aatcttatta 307330DNAProteus mirabilis 73 ttgatcctca ttttattaat cacatgacca 307419DNAProteus mirabilis 74 gaaacatcgc aaagtcagt 197520DNAProteus mirabilis 75 ataaaatgag gatcaagttc 207630DNAProteus mirabilis 76 ccgcctttag cattaattgg tgtttatagt 307730DNAProteus mirabilis 77 cctattgcag ataccttaaa tgtcttgggc 307826DNAStreptococcus pneumoniae 78 agtaaaatga aataagaaca ggacag 267925DNAStreptococcus pneumoniae 79 aaaacaggat aggagaacgg gaaaa 258025DNAProteus mirabilis 80 ttgagtgatg atttcactga ctccc 258125DNAProteus mirabilis 81 gtcagacagt gatgctgacg acaca 258227DNAProteus mirabilis 82 tggttgtcat gctgtttgtg tgaaaat 278319DNAPseudomonas aeruginosa 83 cgagcgggtg gtgttcatc 198419DNAPseudomonas aeruginosa 84 caagtcgtcg tcggaggga 198519DNAPseudomonas aeruginosa 85 tcgctgttca tcaagaccc 198619DNAPseudomonas aeruginosa 86 ccgagaacca gacttcatc 198725DNAPseudomonas aeruginosa 87 aatgcggctg tacctcggcg ctggt 258825DNAPseudomonas aeruginosa 88 ggcggagggc cagttgcacc tgcca 258925DNAPseudomonas aeruginosa 89 agccctgctc ctcggcagcc tctgc 259025DNAPseudomonas aeruginosa 90 tggcttttgc aaccgcgttc aggtt 259125DNAPseudomonas aeruginosa 91 gcgcccgcga gggcatgctt cgatg 259225DNAPseudomonas aeruginosa 92 acctgggcgc caactacaag ttcta 259325DNAPseudomonas aeruginosa 93 ggctacgctg ccgggctgca ggccg 259425DNAPseudomonas aeruginosa 94 ccgatctaca ccatcgagat gggcg 259525DNAPseudomonas aeruginosa 95 gagcgcggct atgtgttcgt cggct 259629DNAStaphylococcus saprophyticus 96 cgtttttacc cttacctttt cgtactacc 299730DNAStaphylococcus saprophyticus 97 tcaggcagag gtagtacgaa aaggtaaggg 309826DNAStaphylococcus saprophyticus 98 cgtttttacc cttacctttt cgtact 269928DNAStaphylococcus saprophyticus 99 atcgatcatc acattccatt tgttttta 2810027DNAStaphylococcus saprophyticus 100 caccaagttt gacacgtgaa gattcat 2710130DNAStaphylococcus saprophyticus 101 atgagtgaag cggagtcaga ttatgtgcag 3010225DNAStaphylococcus saprophyticus 102 cgctcattac gtacagtgac aatcg 2510330DNAStaphylococcus saprophyticus 103 ctggttagct tgactcttaa caatcttgtc 3010430DNAStaphylococcus saprophyticus 104 gacgcgattg tcactgtacg taatgagcga 3010528DNAHaemophilus influenzae 105 gcgtcagaaa aagtaggcga aatgaaag 2810625DNAHaemophilus influenzae 106 agcggctcta tcttgtaatg acaca 2510725DNAHaemophilus influenzae 107 gaaacgtgaa ctcccctcta tataa 2510825DNAMoraxella catarrhalis 108 gccccaaaac aatgaaacat atggt 2510925DNAMoraxella catarrhalis 109 ctgcagattt tggaatcata tcgcc 2511025DNAMoraxella catarrhalis 110 tggtttgacc agtatttaac gccat 2511125DNAMoraxella catarrhalis 111 caacggcacc tgatgtacct tgtac 2511218DNAMoraxella catarrhalis 112 ggcacctgat gtaccttg 1811317DNAMoraxella catarrhalis 113 aacagctcac acgcatt 1711425DNAMoraxella catarrhalis 114 ttacaacctg caccacaagt catca 2511525DNAMoraxella catarrhalis 115 gtacaaacaa gccgtcagcg actta 2511623DNAMoraxella catarrhalis 116 caatctgcgt gtgtgcgttc act 2311726DNAMoraxella catarrhalis 117 gctactttgt cagctttagc cattca 2611824DNAMoraxella catarrhalis 118 tgttttgagc tttttatttt ttga 2411922DNAMoraxella catarrhalis 119 cgctgacggc ttgtttgtac ca 2212025DNAStreptococcus pneumoniae 120 tctgtgctag agactgcccc atttc 2512125DNAStreptococcus pneumoniae 121 cgatgtcttg attgagcagg gttat 2512225DNAArtificial SequenceDescription of Artificial Sequence Oligonucleotide 122 atcccacctt aggcggctgg ctcca 2512331DNAArtificial SequenceDescription of Artificial Sequence Oligonucleotide 123 acgtcaagtc atcatggccc ttacgagtag g 3112425DNAArtificial SequenceDescription of Artificial Sequence Oligonucleotide 124 gtgtgacggg cggtgtgtac aaggc 2512528DNAArtificial SequenceDescription of Artificial Sequence Oligonucleotide 125 gagttgcaga ctccaatccg gactacga 2812620DNAArtificial SequenceDescription of Artificial Sequence Oligonucleotide 126 ggaggaaggt ggggatgacg 2012720DNAArtificial SequenceDescription of Artificial Sequence Oligonucleotide 127 atggtgtgac gggcggtgtg 2012832DNAArtificial SequenceDescription of Artificial Sequence Oligonucleotide 128 ccctatacat caccttgcgg tttagcagag ag 3212928DNAArtificial SequenceDescription of Artificial Sequence Oligonucleotide 129 ggggggacca tcctccaagg ctaaatac 2813032DNAArtificial SequenceDescription of Artificial Sequence Oligonucleotide 130 cgtccacttt cgtgtttgca gagtgctgtg tt 3213120DNAEscherichia coli 131 caggagtacg gtgattttta 2013220DNAEscherichia coli 132 atttctggtt tggtcataca 2013320DNAProteus mirabilis 133 cgggagtcag tgaaatcatc 2013420DNAProteus mirabilis 134 ctaaaatcgc cacacctctt 2013518DNAKlebsiella pneumoniae 135 gcagcgtggt gtcgttca 1813618DNAKlebsiella pneumoniae 136 agctggcaac ggctggtc 1813720DNAKlebsiella pneumoniae 137 attcacaccc tacgcagcca 2013820DNAKlebsiella pneumoniae 138 atccggcagc atctctttgt 2013925DNAStaphylococcus saprophyticus 139 ctggttagct tgactcttaa caatc 2514025DNAStaphylococcus saprophyticus 140 tcttaacgat agaatggagc aactg 2514120DNAStreptococcus pyogenes 141 tgaaaattct tgtaacaggc 2014220DNAStreptococcus pyogenes 142 ggccaccagc ttgcccaata 2014320DNAStreptococcus pyogenes 143 atattttctt tatgagggtg 2014420DNAStreptococcus pyogenes 144 atccttaaat aaagttgcca 2014525DNAStaphylococcus epidermidis 145 atcaaaaagt tggcgaacct tttca 2514625DNAStaphylococcus epidermidis 146 caaaagagcg tggagaaaag tatca 2514730DNAStaphylococcus epidermidis 147 tctcttttaa tttcatcttc aattccatag 3014830DNAStaphylococcus epidermidis 148 aaacacaatt acagtctggt tatccatatc 3014930DNAStaphylococcus aureus 149 cttcatttta cggtgacttc ttagaagatt 3015030DNAStaphylococcus aureus 150 tcaactgtag cttctttatc catacgttga 3015130DNAStaphylococcus aureus 151 atattttagc ttttcagttt ctatatcaac 3015230DNAStaphylococcus aureus 152 aatctttgtc ggtacacgat attcttcacg 3015330DNAStaphylococcus aureus 153 cgtaatgaga tttcagtaga taatacaaca 3015425DNAHaemophilus influenzae 154 tttaacgatc cttttactcc ttttg 2515525DNAHaemophilus influenzae 155 actgctgttg taaagaggtt aaaat 2515620DNAStreptococcus pneumoniae 156 atttggtgac gggtgacttt 2015720DNAStreptococcus pneumoniae 157 gctgaggatt tgttcttctt 2015820DNAStreptococcus pneumoniae 158 gagcggtttc tatgattgta 2015920DNAStreptococcus pneumoniae 159 atctttcctt tcttgttctt 2016018DNAMoraxella catarrhalis 160 gctcaaatca gggtcagc 18161861DNAEscherichia coli 161 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct 60 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca 120 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc 180 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc 240 cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg 300 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta 360 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc 420 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt 480 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg 540 cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct 600 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc 660 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct 720 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac 780 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc 840 tcactgatta agcattggta a 861162918DNAPasteurella haemolytica 162 atgttaaata agttaaaaat cggcacatta ttattgctga cattaacggc ttgttcgccc 60 aattctgttc attcggtaac gtctaatccg cagcctgcta gtgcgcctgt gcaacaatca 120 gccacacaag ccacctttca acagactttg gcgaatttgg aacagcagta tcaagcccga 180 attggcgttt atgtatggga tacagaaacg ggacattctt tgtcttatcg tgcagatgaa 240 cgctttgctt atgcgtccac tttcaaggcg ttgttggctg gggcggtgtt gcaatcgctg 300 cctgaaaaag atttaaatcg taccatttca tatagccaaa aagatttggt tagttattct 360 cccgaaaccc aaaaatacgt tggcaaaggc atgacgattg cccaattatg tgaagcagcc 420 gtgcggttta gcgacaacag cgcgaccaat ttgctgctca aagaattggg tggcgtggaa 480 caatatcaac gtattttgcg acaattaggc gataacgtaa cccataccaa tcggctagaa 540 cccgatttaa atcaagccaa acccaacgat attcgtgata cgagtacacc caaacaaatg 600 gcgatgaatt taaatgcgta tttattgggc aacacattaa ccgaatcgca aaaaacgatt 660 ttgtggaatt ggttggacaa taacgcaaca ggcaatccat tgattcgcgc tgctacgcca 720 acatcgtgga aagtgtacga taaaagcggg gcgggtaaat atggtgtacg caatgatatt 780 gcggtggttc gcataccaaa tcgcaaaccg attgtgatgg caatcatgag tacgcaattt 840 accgaagaag ccaaattcaa caataaatta gtagaagatg cagcaaagca agtatttcat 900 actttacagc tcaactaa 918163864DNAKlebsiella pneumoniae 163 atgcgttata ttcgcctgtg tattatctcc ctgttagcca ccctgccgct ggcggtacac 60 gccagcccgc agccgcttga gcaaattaaa ctaagcgaaa gccagctgtc gggccgcgta 120 ggcatgatag aaatggatct ggccagcggc cgcacgctga ccgcctggcg cgccgatgaa 180 cgctttccca tgatgagcac ctttaaagta gtgctctgcg gcgcagtgct ggcgcgggtg 240 gatgccggtg acgaacagct ggagcgaaag atccactatc gccagcagga tctggtggac 300 tactcgccgg tcagcgaaaa acaccttgcc gacgcaatga cggtcggcga actctgcgcc 360 gccgccatta ccatgagcga taacagcgcc gccaatctgc tactggccac cgtcggcggc 420 cccgcaggat tgactgcctt tttgcgccag atcggcgaca acgtcacccg ccttgaccgc 480 tgggaaacgg aactgaatga ggcgcttccc ggcgacgccc gcgacaccac taccccggcc 540 agcatggccg cgaccctgcg caacgttggc ctgaccagcc agcgtctgag cgcccgttcg 600 caacggcagc tgctgcagtg gatggtggac gatcgggtcg ccggaccgtt gatccgctcc 660 gtgctgccgg cgggctggtt tatcgccgat aagaccggag ctggcgagcg gggtgcgcgc 720 gggattgtcg ccctgcttgg cccgaataac aaagcagagc gcattgtggt gatttatctg 780 cgggataccc cggcgagcat ggccgagcga aatcagcaaa tcgccgggat cggcaaggcg 840 ctgtacgagc actggcaacg ctaa 864164534DNAKlebsiella pneumoniae 164 atggacacaa cgcaggtcac attgatacac aaaattctag ctgcggcaga tgagcgaaat 60 ctgccgctct ggatcggtgg gggctgggcg atcgatgcac ggctagggcg tgtaacacgc 120 aagcacgatg atattgatct gacgtttccc ggcgagaggc gcggcgagct cgaggcaata 180 gttgaaatgc tcggcgggcg cgtcatggag gagttggact atggattctt agcggagatc 240 ggggatgagt tacttgactg cgaacctgct tggtgggcag acgaagcgta tgaaatcgcg 300 gaggctccgc agggctcgtg cccagaggcg gctgagggcg tcatcgccgg gcggccagtc 360 cgttgtaaca gctgggaggc gatcatctgg gattactttt actatgccga tgaagtacca 420 ccagtggact ggcctacaaa gcacatagag tcctacaggc tcgcatgcac ctcactcggg 480 gcggaaaagg ttgaggtctt gcgtgccgct ttcaggtcgc gatatgcggc ctaa 534165465DNAUnknown OrganismDescription of Unknown Organism Enterobacteriaceae 165 atgggcatca ttcgcacatg taggctcggc cctgaccaag tcaaatccat gcgggctgct 60 cttgatcttt tcggtcgtga gttcggagac gtagccacct actcccaaca tcagccggac 120 tccgattacc tcgggaactt gctccgtagt aagacattca tcgcgcttgc tgccttcgac 180 caagaagcgg ttgttggcgc tctcgcggct tacgttctgc ccaggtttga gcagccgcgt 240 agtgagatct atatctatga tctcgcagtc tccggcgagc accggaggca gggcattgcc 300 accgcgctca tcaatctcct caagcatgag gccaacgcgc ttggtgctta tgtgatctac 360 gtgcaagcag attacggtga cgatcccgca gtggctctct atacaaagtt gggcatacgg 420 gaagaagtga tgcactttga tatcgaccca agtaccgcca cctaa 465166861DNAEscherichia coli 166 atgcatacgc ggaaggcaat aacggaggcg cttcaaaaac tcggagtcca aaccggtgac 60 ctattgatgg tgcatgcctc acttaaagcg attggtccgg tcgaaggagg agcggagacg 120 gtcgttgccg cgttacgctc cgcggttggg ccgactggca ctgtgatggg atacgcatcg 180 tgggaccgat caccctacga ggagactcgt aatggcgctc ggttggatga caaaacccgc 240 cgtacctggc cgccgttcga tcccgcaacg gccgggactt accgtgggtt cggcctgctg 300 aatcagtttc tggttcaagc ccccggcgcg cggcgcagcg cgcaccccga tgcatcgatg 360 gtcgcggttg gtccactggc tgaaacgctg acggagcctc acaagctcgg tcacgccttg 420 ggggaagggt cgcccgtcga gcggttcgtt cgccttggcg ggaaggccct gctgttgggt 480 gcgccgctaa actccgttac cgcattgcac tacgccgagg cggttgccga tatccccaac 540 aaacggcggg tgacgtatga gatgccgatg cttggaagca acggcgaagt cgcctggaaa 600 acggcatcgg attacgattc aaacggcatt ctcgattgct ttgctatcga aggaaagccg 660 gatgcggtcg aaactatagc aaatgcttac gtgaagctcg gtcgccatcg agaaggtgtc 720 gtgggctttg ctcagtgcta cctgttcgac gcgcaggaca tcgtgacgtt cggcgtcacc 780 tatcttgaga agcatttcgg aaccactccg atcgtgccag cacacgaagt cgccgagtgc 840 tcttgcgagc cttcaggtta g 861167816DNAPseudomonas aeruginosa 167 atgaccgatt tgaatatccc gcatacacac gcgcaccttg tagacgcatt tcaggcgctc 60 ggcatccgcg cggggcaggc gctcatgctg cacgcatccg ttaaagcagt gggcgcggtg 120 atgggcggcc ccaatgtgat cttgcaggcg ctcatggatg cgctcacgcc cgacggcacg 180 ctgatgatgt atgcgggatg gcaagacatc cccgacttta tcgactcgct gccggacgcg 240 ctcaaggccg tgtatcttga gcagcaccca ccctttgacc ccgccaccgc ccgcgccgtg 300 cgcgaaaaca gcgtgctagc ggaatttttg cgcacatggc cgtgcgtgca tcgcagcgca 360 aaccccgaag cctctatggt ggcggtaggc aggcaggccg ctttgctgac cgctaatcac 420 gcgctggatt atggctacgg agtcgagtcg ccgctggcta aactggtggc aatagaagga 480 tacgtgctga tgcttggcgc gccgctggat accatcacac tgctgcacca cgcggaatat 540 ctggccaaga tgcgccacaa gaacgtggtc cgctacccgt gcccgattct gcgggacggg 600 cgcaaagtgt gggtgaccgt tgaggactat gacaccggtg atccgcacga cgattatagt 660 tttgagcaaa tcgcgcgcga ttatgtggcg cagggcggcg gcacacgcgg caaagtcggt 720 gatgcggatg cttacctgtt cgccgcgcag gacctcacac ggtttgcggt gcagtggctt 780 gaatcacggt tcggtgactc agcgtcatac ggatag 816168498DNAPseudomonas aeruginosa 168 atgctctatg agtggctaaa tcgatctcat atcgtcgagt ggtggggcgg agaagaagca 60 cgcccgacac ttgctgacgt acaggaacag tacttgccaa gcgttttagc gcaagagtcc 120 gtcactccat acattgcaat gctgaatgga gagccgattg ggtatgccca gtcgtacgtt 180 gctcttggaa gcggggacgg atggtgggaa gaagaaaccg atccaggagt acgcggaata 240 gaccagttac tggcgaatgc atcacaactg ggcaaaggct tgggaaccaa gctggttcga 300 gctctggttg agttgctgtt caatgatccc gaggtcacca agatccaaac ggacccgtcg 360 ccgagcaact tgcgagcgat ccgatgctac gagaaagcgg ggtttgagag gcaaggtacc 420 gtaaccaccc cagatggtcc agccgtgtac atggttcaaa cacgccaggc attcgagcga 480 acacgcagtg atgcctaa 4981692007DNAStaphylococcus aureus 169 atgaaaaaga taaaaattgt tccacttatt ttaatagttg tagttgtcgg gtttggtata 60 tatttttatg cttcaaaaga taaagaaatt aataatacta ttgatgcaat tgaagataaa 120 aatttcaaac aagtttataa agatagcagt tatatttcta aaagcgataa tggtgaagta 180 gaaatgactg aacgtccgat aaaaatatat aatagtttag gcgttaaaga tataaacatt 240 caggatcgta aaataaaaaa agtatctaaa aataaaaaac gagtagatgc tcaatataaa 300 attaaaacaa actacggtaa cattgatcgc aacgttcaat ttaattttgt taaagaagat 360 ggtatgtgga agttagattg ggatcatagc gtcattattc caggaatgca gaaagaccaa 420 agcatacata ttgaaaattt aaaatcagaa cgtggtaaaa ttttagaccg aaacaatgtg 480 gaattggcca atacaggaac acatatgaga ttaggcatcg ttccaaagaa tgtatctaaa 540 aaagattata aagcaatcgc taaagaacta agtatttctg aagactatat caacaacaaa 600 tggatcaaaa ttgggtacaa gatgatacct tcgttccact ttaaaaccgt taaaaaaatg 660 gatgaatatt taagtgattt cgcaaaaaaa tttcatctta caactaatga aacagaaagt 720 cgtaactatc ctctagaaaa agcgacttca catctattag gttatgttgg tcccattaac 780 tctgaagaat taaaacaaaa agaatataaa ggctataaag atgatgcagt tattggtaaa 840 aagggactcg aaaaacttta cgataaaaag ctccaacatg aagatggcta tcgtgtcaca 900 atcgttgacg ataatagcaa tacaatcgca catacattaa tagagaaaaa gaaaaaagat 960 ggcaaagata ttcaactaac tattgatgct aaagttcaaa agagtattta taacaacatg 1020 aaaaatgatt atggctcagg tactgctatc caccctcaaa caggtgaatt attagcactt 1080 gtaagcacac cttcatatga cgtctatcca tttatgtatg gcatgagtaa cgaagaatat 1140 aataaattaa ccgaagataa aaaagaacct ctgctcaaca agttccagat tacaacttca 1200 ccaggttcaa ctcaaaaaat attaacagca atgattgggt taaataacaa aacattagac 1260 gataaaacaa gttataaaat cgatggtaaa ggttggcaaa aagataaatc ttggggtggt 1320 tacaacgtta caagatatga agtggtaaat ggtaatatcg acttaaaaca agcaatagaa 1380 tcatcagata acattttctt tgctagagta gcactcgaat taggcagtaa gaaatttgaa 1440 aaaggcatga aaaaactagg tgttggtgaa gatataccaa gtgattatcc attttataat 1500 gctcaaattt caaacaaaaa tttagataat gaaatattat tagctgattc aggttacgga 1560 caaggtgaaa tactgattaa cccagtacag atcctttcaa tctatagcgc attagaaaat 1620 aatggcaata ttaacgcacc tcacttatta aaagacacga aaaacaaagt ttggaagaaa 1680 aatattattt ccaaagaaaa tatcaatcta ttaaatgatg gtatgcaaca agtcgtaaat 1740 aaaacacata aagaagatat ttatagatct tatgcaaact taattggcaa atccggtact 1800 gcagaactca aaatgaaaca aggagaaagt ggcagacaaa ttgggtggtt tatatcatat 1860 gataaagata atccaaacat gatgatggct attaatgtta aagatgtaca agataaagga 1920 atggctagct acaatgccaa aatctcaggt aaagtgtatg atgagctata tgagaacggt 1980 aataaaaaat acgatataga tgaataa 20071702607DNAEnterococcus faecium 170 atgaataaca tcggcattac tgtttatgga tgtgagcagg atgaggcaga tgcattccat 60 gctctttcgc ctcgctttgg cgttatggca acgataatta acgccaacgt gtcggaatcc 120 aacgccaaat ccgcgccttt caatcaatgt atcagtgtgg gacataaatc agagatttcc 180 gcctctattc ttcttgcgct gaagagagcc ggtgtgaaat atatttctac ccgaagcatc 240 ggctgcaatc atatagatac aactgctgct aagagaatgg gcatcactgt cgacaatgtg 300 gcgtactcgc cggatagcgt tgccgattat actatgatgc taattcttat ggcagtacgc 360 aacgtaaaat cgattgtgcg ctctgtggaa aaacatgatt tcaggttgga cagcgaccgt 420 ggcaaggtac tcagcgacat gacagttggt gtggtgggaa cgggccagat aggcaaagcg 480 gttattgagc ggctgcgagg atttggatgt aaagtgttgg cttatagtcg cagccgaagt 540 atagaggtaa actatgtacc gtttgatgag ttgctgcaaa atagcgatat cgttacgctt 600 catgtgccgc tcaatacgga tacgcactat attatcagcc acgaacaaat acagagaatg 660 aagcaaggag catttcttat caatactggg cgcggtccac ttgtagatac ctatgagttg 720 gttaaagcat tagaaaacgg gaaactgggc ggtgccgcat tggatgtatt ggaaggagag 780 gaagagtttt tctactctga ttgcacccaa aaaccaattg ataatcaatt tttacttaaa 840 cttcaaagaa tgcctaacgt gataatcaca ccgcatacgg cctattatac cgagcaagcg 900 ttgcgtgata ccgttgaaaa aaccattaaa aactgtttgg attttgaaag gagacaggag 960 catgaataga ataaaagttg caatactgtt tgggggttgc tcagaggagc atgacgtatc 1020 ggtaaaatct gcaatagaga tagccgctaa cattaataaa gaaaaatacg agccgttata 1080 cattggaatt acgaaatctg gtgtatggaa aatgtgcgaa aaaccttgcg cggaatggga 1140 aaacgacaat tgctattcag ctgtactctc gccggataaa aaaatgcacg gattacttgt 1200 taaaaagaac catgaatatg aaatcaacca tgttgatgta gcattttcag ctttgcatgg 1260 caagtcaggt gaagatggat ccatacaagg tctgtttgaa ttgtccggta tcccttttgt 1320 aggctgcgat attcaaagct cagcaatttg tatggacaaa tcgttgacat acatcgttgc 1380 gaaaaatgct gggatagcta ctcccgcctt ttgggttatt aataaagatg ataggccggt 1440 ggcagctacg tttacctatc ctgtttttgt taagccggcg cgttcaggct catccttcgg 1500 tgtgaaaaaa gtcaatagcg cggacgaatt ggactacgca attgaatcgg caagacaata 1560 tgacagcaaa atcttaattg agcaggctgt ttcgggctgt gaggtcggtt gtgcggtatt 1620 gggaaacagt gccgcgttag ttgttggcga ggtggaccaa atcaggctgc agtacggaat 1680 ctttcgtatt catcaggaag tcgagccgga aaaaggctct gaaaacgcag ttataaccgt 1740 tcccgcagac ctttcagcag aggagcgagg acggatacag gaaacggcaa aaaaaatata 1800 taaagcgctc ggctgtagag gtctagcccg tgtggatatg tttttacaag ataacggccg 1860 cattgtactg aacgaagtca atactctgcc cggtttcacg tcatacagtc gttatccccg 1920 tatgatggcc gctgcaggta ttgcacttcc cgaactgatt gaccgcttga tcgtattagc 1980 gttaaagggg tgataagcat ggaaatagga tttacttttt tagatgaaat agtacacggt 2040 gttcgttggg acgctaaata tgccacttgg gataatttca ccggaaaacc ggttgacggt 2100 tatgaagtaa atcgcattgt agggacatac gagttggctg aatcgctttt gaaggcaaaa 2160 gaactggctg ctacccaagg gtacggattg cttctatggg acggttaccg tcctaagcgt 2220 gctgtaaact gttttatgca atgggctgca cagccggaaa ataacctgac aaaggaaagt 2280 tattatccca atattgaccg aactgagatg atttcaaaag gatacgtggc ttcaaaatca 2340 agccatagcc gcggcagtgc cattgatctt acgctttatc gattagacac gggtgagctt 2400 gtaccaatgg ggagccgatt tgattttatg gatgaacgct ctcatcatgc ggcaaatgga 2460 atatcatgca atgaagcgca aaatcgcaga cgtttgcgct ccatcatgga aaacagtggg 2520 tttgaagcat atagcctcga atggtggcac tatgtattaa gagacgaacc ataccccaat 2580 agctattttg atttccccgt taaataa 26071711288DNAPseudomonas aeruginosa 171 ggatccatca ggcaacgacg ggctgctgcc ggccatcagc ggacgcaggg aggactttcc 60 gcaaccggcc gttcgatgcg gcaccgatgg ccttcgcgca ggggtagtga atccgccagg 120 attgacttgc gctgccctac ctctcactag tgaggggcgg cagcgcatca agcggtgagc 180 gcactccggc accgccaact ttcagcacat gcgtgtaaat catcgtcgta gagacgtcgg 240 aatggccgag cagatcctgc acggttcgaa tgtcgtaacc gctgcggagc aaggccgtcg 300 cgaacgagtg gcggagggtg tgcggtgtgg cgggcttcgt gatgcctgct tgttctacgg 360 cacgtttgaa ggcgcgctga aaggtctggt catacatgtg atggcgacgc acgacaccgc 420 tccgtggatc ggtcgaatgc gtgtgctgcg caaaaaccca gaaccacggc caggaatgcc 480 cggcgcgcgg atacttccgc tcaagggcgt cgggaagcgc aacgccgctg cggccctcgg 540 cctggtcctt cagccaccat gcccgtgcac gcgacagctg ctcgcgcagg ctgggtgcca 600 agctctcggg taacatcaag gcccgatcct tggagccctt gccctcccgc acgatgatcg 660 tgccgtgatc gaaatccaga tccttgaccc gcagttgcaa accctcactg atccgcatgc 720 ccgttccata cagaagctgg gcgaacaaac gatgctcgcc ttccagaaaa ccgaggatgc 780 gaaccacttc atccggggtc agcaccaccg gcaagcgccg cgacggccga ggtcttccga 840 tctcctgaag ccagggcaga tccgtgcaca gcaccttgcc gtagaagaac agcaaggccg 900 ccaatgcctg acgatgcgtg gagaccgaaa ccttgcgctc gttcgccagc caggacagaa 960 atgcctcgac ttcgctgctg cccaaggttg ccgggtgacg cacaccgtgg aaacggatga 1020 aggcacgaac ccagtggaca taagcctgtt cggttcgtaa gctgtaatgc aagtagcgta 1080 tgcgctcacg caactggtcc agaaccttga ccgaacgcag cggtggtaac ggcgcagtgg 1140 cggttttcat ggcttgttat gactgttttt ttgtacagtc tatgcctcgg gcatccaagc 1200 agcaagcgcg ttacgccgtg ggtcgatgtt tgatgttatg gagcagcaac gatgttacgc 1260 agcagggcag tcgccctaaa acaaagtt 12881721650DNAPseudomonas aeruginosa 172 gttagatgca ctaagcacat aattgctcac agccaaacta tcaggtcaag tctgctttta 60 ttatttttaa gcgtgcataa taagccctac acaaattggg agatatatca tgaaaggctg 120 gctttttctt gttatcgcaa tagttggcga agtaatcgca acatccgcat taaaatctag 180 cgagggcttt actaagcttg ccccttccgc cgttgtcata atcggttatg gcatcgcatt 240 ttattttctt tctctggttc tgaaatccat ccctgtcggt gttgcttatg cagtctggtc 300 gggactcggc gtcgtcataa ttacagccat tgcctggttg cttcatgggc aaaagcttga 360 tgcgtggggc tttgtaggta tggggctcat aattgctgcc tttttgctcg cccgatcccc 420 atcgtggaag tcgctgcgga ggccgacgcc atggtgacgg tgttcggcat tctgaatctc 480 accgaggact ccttcttcga tgagagccgg cggctagacc ccgccggcgc tgtcaccgcg 540 gcgatcgaaa tgctgcgagt cggatcagac gtcgtggatg tcggaccggc cgccagccat 600 ccggacgcga ggcctgtatc gccggccgat gagatcagac gtattgcgcc gctcttagac 660 gccctgtccg atcagatgca ccgtgtttca atcgacagct tccaaccgga aacccagcgc 720 tatgcgctca agcgcggcgt gggctacctg aacgatatcc aaggatttcc tgaccctgcg 780 ctctatcccg atattgctga ggcggactgc aggctggtgg ttatgcactc agcgcagcgg 840 gatggcatcg ccacccgcac cggtcacctt cgacccgaag acgcgctcga cgagattgtg 900 cggttcttcg aggcgcgggt ttccgccttg cgacggagcg gggtcgctgc cgaccggctc 960 atcctcgatc cggggatggg atttttcttg agccccgcac cggaaacatc gctgcacgtg 1020 ctgtcgaacc ttcaaaagct gaagtcggcg ttggggcttc cgctattggt ctcggtgtcg 1080 cggaaatcct tcttgggcgc caccgttggc cttcctgtaa aggatctggg tccagcgagc 1140 cttgcggcgg aacttcacgc gatcggcaat ggcgctgact acgtccgcac ccacgcgcct 1200 ggagatctgc gaagcgcaat caccttctcg gaaaccctcg cgaaatttcg cagtcgcgac 1260 gccagagacc gagggttaga tcatgcctag cattcacctt ccggccgccc gctagcggac 1320 cctggtcagg ttccgcgaag gtgggcgcag acatgctggg ctcgtcagga tcaaactgca 1380 ctatgaggcg gcggttcata ccgcgccagg ggagcgaatg gacagcgagg agcctccgaa 1440 cgttcgggtc gcctgctcgg gtgatatcga cgaggttgtg cggctgatgc acgacgctgc 1500 ggcgtggatg tccgccaagg gaacgcccgc ctgggacgtc gcgcggatcg accggacatt 1560 cgcggagacc ttcgtcctga gatccgagct cctagtcgcg agttgcagcg acggcatcgt 1620 cggctgttgc accttgtcgg ccgaggatcc 1650173630DNAEnterococcus faecium 173 atgggtccga atcctatgaa aatgtatcct atagaaggaa acaaatcagt acaatttatc 60 aaacctattt tagaaaaatt agaaaatgtt gaggttggag aatactcata ttatgattct 120 aagaatggag aaacttttga taagcaaatt ttatatcatt atccaatctt aaacgataag 180 ttaaaaatag gtaaattttg ctcaatagga ccaggtgtaa ctattattat gaatggagca 240 aatcatagaa tggatggctc aacatatcca tttaatttat ttggtaatgg atgggagaaa 300 catatgccaa aattagatca actacctatt aagggggata caataatagg taatgatgta 360 tggataggaa aagatgttgt aattatgcca ggagtaaaaa tcggggatgg tgcaatagta 420 gctgctaatt ctgttgttgt aaaagatata gcgccataca tgttagctgg aggaaatcct 480 gctaacgaaa taaaacaaag atttgatcaa gatacaataa atcagctgct tgatataaaa 540 tggtggaatt ggccaataga cattattaat gagaatatag ataaaattct tgataatagc 600 atcattagag aagtcatatg gaaaaaatga 6301741440DNAEnterococcus faecalis 174 atgaatatag ttgaaaatga aatatgtata agaactttaa tagatgatga ttttcctttg 60 atgttaaaat ggttaactga tgaaagagta ttagaatttt atggtggtag agataaaaaa 120 tatacattag aatcattaaa aaaacattat acagagcctt gggaagatga agtttttaga 180 gtaattattg aatataacaa tgttcctatt ggatatggac aaatatataa aatgtatgat 240 gagttatata ctgattatca ttatccaaaa actgatgaga tagtctatgg tatggatcaa 300 tttataggag agccaaatta ttggagtaaa ggaattggta caagatatat taaattgatt 360 tttgaatttt tgaaaaaaga aagaaatgct aatgcagtta ttttagaccc tcataaaaat 420 aatccaagag caataagggc ataccaaaaa tctggtttta gaattattga agatttgcca 480 gaacatgaat tacacgaggg caaaaaagaa gattgttatt taatggaata tagatatgat 540 gataatgcca caaatgttaa ggcaatgaaa tatttaattg agcattactt tgataatttc 600 aaagtagata gtattgaaat aatcggtagt ggttatgata gtgtggcata tttagttaat 660 aatgaataca tttttaaaac aaaatttagt actaataaga aaaaaggtta tgcaaaagaa 720 aaagcaatat ataatttttt aaatacaaat ttagaaacta atgtaaaaat tcctaatatt 780 gaatattcgt atattagtga tgaattatct atactaggtt ataaagaaat taaaggaact 840 tttttaacac cagaaattta ttctactatg tcagaagaag aacaaaattt gttaaaacga 900 gatattgcca gttttttaag acaaatgcac ggtttagatt atacagatat tagtgaatgt 960 actattgata ataaacaaaa tgtattagaa gagtatatat tgttgcgtga aactatttat 1020 aatgatttaa ctgatataga aaaagattat atagaaagtt ttatggaaag actaaatgca 1080 acaacagttt ttgagggtaa aaagtgttta tgccataatg attttagttg taatcatcta 1140 ttgttagatg gcaataatag attaactgga ataattgatt ttggagattc tggaattata 1200 gatgaatatt gtgattttat atacttactt gaagatagtg aagaagaaat aggaacaaat 1260 tttggagaag atatattaag aatgtatgga aatatagata ttgagaaagc aaaagaatat 1320 caagatatag ttgaagaata ttatcctatt gaaactattg tttatggaat taaaaatatt 1380 aaacaggaat ttatcgaaaa tggtagaaaa gaaatttata aaaggactta taaagattga 1440175660DNAStaphylococcus aureus 175 ttgaatttaa acaatgacca tggacctgat cccgaaaata ttttaccgat aaaagggaat 60 cggaatcttc aatttataaa acctactata acgaacgaaa acattttggt gggggaatat 120 tcttattatg atagtaagcg aggagaatcc tttgaagatc aagtcttata tcattatgaa 180 gtgattggag ataagttgat tataggaaga ttttgttcaa ttggtcccgg aacaacattt 240 attatgaatg gtgcaaacca tcggatggat ggatcaacat atccttttca tctattcagg 300 atgggttggg agaagtatat gccttcctta aaagatcttc ccttgaaagg ggacattgaa 360 attggaaatg atgtatggat aggtagagat gtaaccatta tgcctggggt gaaaattggg 420 gacggggcaa tcattgctgc agaagctgtt gtcacaaaga atgttgctcc ctattctatt 480 gtcggtggaa atcccttaaa atttataaga aaaaggtttt ctgatggagt tatcgaagaa 540 tggttagctt tacaatggtg gaatttagat atgaaaatta ttaatgaaaa tcttcccttc 600 ataataaatg gagatatcga aatgctgaag agaaaaagaa aacttctaga tgacacttga 6601761569DNAStaphylococcus aureus 176 atgaaaataa tgttagaggg acttaatata aaacattatg ttcaagatcg tttattgttg 60 aacataaatc gcctaaagat ttatcagaat gatcgtattg gtttaattgg taaaaatgga 120 agtggaaaaa caacgttact tcacatatta tataaaaaaa ttgtgcctga agaaggtatt 180 gtaaaacaat tttcacattg tgaacttatt cctcaattga agctcataga atcaactaaa 240 agtggtggtg aagtaacacg aaactatatt cggcaagcgc ttgataaaaa tccagaactg 300 ctattagcag atgaaccaac aactaactta gataataact atatagaaaa attagaacag 360 gatttaaaaa attggcatgg agcatttatt atagtttcac atgatcgcgc ttttttagat 420 aacttgtgta ctactatatg ggaaattgac gagggaagaa taactgaata taaggggaat 480 tatagtaact atgttgaaca aaaagaatta gaaagacatc gagaagaatt agaatatgaa 540 aaatatgaaa aagaaaagaa acgattggaa aaagctataa atataaaaga acagaaagct 600 caacgagcaa ctaaaaaacc gaaaaactta agtttatctg aaggcaaaat aaaaggagca 660 aagccatact ttgcaggtaa gcaaaagaag ttacgaaaaa ctgtaaaatc tctagaaacc 720 agactagaaa aacttgaaag cgtcgaaaag agaaacgaac ttcctccact taaaatggat 780 ttagtgaact tagaaagtgt aaaaaataga actataatac gtggtgaaga tgtctcgggt 840 acaattgaag gacgggtatt gtggaaagca aaaagtttta gtattcgcgg aggagacaag 900 atggcaatta tcggatctaa tggtacagga aagacaacgt ttattaaaaa aattgtgcat 960 gggaatcctg gtatttcatt atcgccatct gtcaaaatcg gttattttag ccaaaaaata 1020 gatacattag aattagataa gagcatttta gaaaatgttc aatcttcttc acaacaaaat 1080 gaaactctta ttcgaactat tctagctaga atgcattttt ttagagatga tgtttataaa 1140 ccaataagtg tcttaagtgg tggagagcga gttaaagtag cactaactaa agtattctta 1200 agtgaagtta atacgttggt actagatgaa ccaacaaact ttcttgatat ggaagctata 1260 gaggcgtttg aatctttgtt aaaggaatat aatggcagta taatctttgt atctcacgat 1320 cgtaaattta tcgaaaaagt agccactcga ataatgacaa ttgataataa agaaataaaa 1380 atatttgatg gcacatatga acaatttaaa caagctgaaa agccaacaag gaatattaaa 1440 gaagataaaa aacttttact tgagacaaaa attacagaag tactcagtcg attgagtatt 1500 gaaccttcgg aagaattaga acaagagttt caaaacttaa taaatgaaaa aagaaatttg 1560 gataaataa 15691771467DNAStaphylococcus epidermidis 177 atggaacaat atacaattaa atttaaccaa atcaatcata aattgacaga tttacgatca 60 cttaacatcg atcatcttta tgcttaccaa tttgaaaaaa tagcacttat tgggggtaat 120 ggtactggta aaaccacatt actaaatatg attgctcaaa aaacaaaacc agaatctgga 180 acagttgaaa cgaatggcga aattcaatat tttgaacagc ttaacatgga tgtggaaaat 240 gattttaaca cgttagacgg tagtttaatg agtgaactcc atatacctat gcatacaacc 300 gacagtatga gtggtggtga aaaagcaaaa tataaattac gtaatgtcat atcaaattat 360 agtccgatat tacttttaga tgaacctaca aatcacttgg ataaaattgg taaagattat 420 ctgaataata ttttaaaata ttactatggt actttaatta tagtaagtca cgatagagca 480 cttatagacc aaattgctga cacaatttgg gatatacaag aagatggcac aataagagtg 540 tttaaaggta attacacaca gtatcaaaat caatatgaac aagaacagtt agaacaacaa 600 cgtaaatatg aacagtatat aagtgaaaaa caaagattgt cccaagccag taaagctaaa 660 cgaaatcaag cgcaacaaat ggcacaagca tcatcaaaac aaaaaaataa aagtatagca 720 ccagatcgtt taagtgcatc aaaagaaaaa ggcacggttg agaaggctgc tcaaaaacaa 780 gctaagcata ttgaaaaaag aatggaacat ttggaagaag ttgaaaaacc acaaagttat 840 catgaattca attttccaca aaataaaatt tatgatatcc ataataatta tccaatcatt 900 gcacaaaatc taacattggt taaaggaagt caaaaactgc taacacaagt acgattccaa 960 ataccatatg gcaaaaatat agcgctcgta ggtgcaaatg gtgtaggtaa gacaacttta 1020 cttgaagcta tttaccacca aatagaggga attgattgtt ctcctaaagt gcaaatggca 1080 tactatcgtc aacttgctta tgaagacatg cgtgacgttt cattattgca atatttaatg 1140 gatgaaacgg attcatcaga atcattcagt agagctattt taaataactt gggtttaaat 1200 gaagcacttg agcgttcttg taatgttttg agtggtgggg aaagaacgaa attatcgtta 1260 gcagtattat tttcaacgaa agcgaatatg ttaattttgg atgaaccaac taatttttta 1320 gatattaaaa cattagaagc attagaaatg tttatgaata aatatcctgg aatcattttg 1380 tttacatcac atgatacaag gtttgttaaa catgtatcag ataaaaaatg ggaattaaca 1440 ggacaatcta ttcatgatat aacttaa 1467

Claims (131)

What is claimed is

1. A method using probes (fragments and/or oligonucleotides) and/or amplification primers which are specific, ubiquitous and sensitive for determining the presence and/or amount of nucleic acids from bacterial species selected from the group consisting ofEscherichia coli, Klebsiells pneumoniae, Pseudomonas aeruginosa, Proteus mirabilis, Streptococcus pneumoniae, Staphlyococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Staphlyococcus saprophyticus, Streptococcus pyogenes, Haemophilus influenzaeandMoraxells catarrhalisin a any sample suspected of containing said bacterial nucleic acid, wherein said bacterial nucleic acid or variant or part thereof comprises a selected target region hybridizable with said probes or primers; said method comprising the steps of contacting said sample with said probes or primers and detecting the presence and/or amount of hybridized probes and/or amplified products as an indication of the presence and/or amount of said bacterial species.

2. A method as defined inclaim 1 further using probes (fragments and/or oligonucleotides) and/or amplification primers which are universal and sensitive for determining the presence and/or amount of nucleic acids from any bacteria from any sample suspected of containing said bacterial nucleic acid, wherein said bacterial nucleic acid or variant or part thereof comprises a selected target region hybridizable with said probes or primers; said method comprising the steps of contacting said sample with said probes or primers and detecting the presence and/or amount of hybridized probes and/or amplified products as an indication of the presence and/or amount of said any bacteria.

3. A method as defined inclaim 1 further using probes (fragments and/or oligonucleotides) and/or amplification primers which are specific, ubiquitous and sensitive for determining the presence and/or amount of nucleic acids from an antibiotic resistance gene selected from the group consisting of bla_tem, Bla_rob, Bla_shv, aadB, aacC1, aacC2, aacC3, aacA4, mecA, vanA, vanH, vanX, satA, aacA-aphD, vat, vga, msrA, sul and int in any sample suspected of containing said bacterial nucleic acid, wherein said bacterial nucleic acid or variant or part thereof comprises a selected target region hybridizable with said probes or primers; said method comprising the steps of contacting said sample with said probes or primers and detecting the presence and/or amount of hybridized probes and/or amplified products as an indication of the presence and/or amount of said antibiotic resistance gene.

4. The method of any one of claims1,2 and3 which is performed directly on a sample obtained from human patients, animals, environment or food.

5. The method of any one of claims1,2 and3 which is performed directly on a sample consisting of one or more bacterial colonies.

6. The method of any one ofclaims 1 to5, wherein the bacterial nucleic acid is amplified by a method selected from the group consisting of:

a) polymerase chain reaction (PCR),

b) ligase chain reaction,

c) nucleic acid sequence-based amplification,

d) self-sustained sequence replication,

e) strand displacement amplification,

f) branched DNA signal amplification,

g) nested PCR, and

h) multiplex PCR.

7. The method ofclaim 6 wherein said bacterial nucleic acid is amplified by PCR.

8. The method ofclaim 7 wherein the PCR protocol is modified to determine within one hour the presence of said bacterial nucleic acids by performing for each amplification cycle an annealing step of only one second at 55° C. and a denaturation step of only one second at 95° C. without any elongation step.

9. A method for the detection, identification and/or quantification ofEscherichia colidirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO:3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofEscherichia coli,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofEscherichia coliin said test sample.

10. A method as defined inclaim 9, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-227 nucleotides in length which sequence is comprised in SEQ ID NO: 3 or a complementary sequence thereof,

2) an oligonucleotide of 12-278 nucleotides in length which sequence is comprised in SEQ ID NO: 4 or a complementary sequence thereof,

3) an oligonucleotide of 12-1596 nucleotides in length which sequence is comprised in SEQ ID NO: 5 or a complementary sequence thereof,

4) an oligonucleotide of 12-2703 nucleotides in length which sequence is comprised in SEQ ID NO: 6 or a complementary sequence thereof,

5) an oligonucleotide of 12-1391 nucleotides in length which sequence is comprised in SEQ ID NO: 7 or a complementary sequence thereof, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofEscherichia coli.

11. The method ofclaim 10, wherein the probe for detecting nucleic acid sequences fromEscherichia coliis selected from the group consisting of SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54 and a sequence complementary thereof.

12. A method for detecting the presence and/or amount ofEscherichia coliin a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofEscherichia coliDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6 and SEQ ID NO: 7;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofEscherichia coliin said test sample.

13. The method ofclaim 12, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 42 and SEQ ID NO: 43,

b) SEQ ID NO: 46 and SEQ ID NO: 47,

c) SEQ ID NO: 55 and SEQ ID NO: 56, and

d) SEQ ID NO: 131 and SEQ ID NO: 132.

14. A method for the detection, identification and/or quantification ofKlebsiella pneumoniaedirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO:8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofKlebsiella pneumoniae,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofKlebsiella pneumoniaein said test sample.

15. A method as defined inclaim 14, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-238 nucleotides in length which sequence is comprised in SEQ ID NO: 8 or a complementary sequence thereof,

2) an oligonucleotide of 12-385 nucleotides in length which sequence is comprised in SEQ ID NO: 9 or a complementary sequence thereof,

3) an oligonucleotide of 12-462 nucleotides in length which sequence is comprised in SEQ ID NO: 10 or a complementary sequence thereof,

4) an oligonucleotide of 12-730 nucleotides in length which sequence is comprised in SEQ ID NO: 11 or a complementary sequence thereof, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofKlebsiella pneumoniae.

16. The method ofclaim 15, wherein the probe for detecting nucleic acid sequences fromKlebsiella pneumoniaeis selected from the group consisting of SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 69 and a sequence complementary thereof.

17. A method for detecting the presence and/or amount ofKlebsiella pneumoniaein a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofKlebsiella pneumoniaeDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, and SEQ ID NO: 11;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofKlebsiella pneumoniaein said test sample.

18. The method ofclaim 17, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 61 and SEQ ID NO: 62,

b) SEQ ID NO: 67 and SEQ ID NO: 68,

c) SEQ ID NO: 135 and SEQ ID NO: 136, and

d) SEQ ID NO: 137 and SEQ ID NO: 138.

19. A method for the detection, identification and/or quantification ofProteus mirabilisdirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofProteus mirabilis,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofProteus mirabilisin said test sample.

20. A method as defined inclaim 19, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-225 nucleotides in length which sequence is comprised in SEQ ID NO: 12 or a complementary sequence thereof,

2) an oligonucleotide of 12-402 nucleotides in length which sequence is comprised in SEQ ID NO: 13 or a complementary sequence thereof,

3) an oligonucleotide of 12-157 nucleotides in length which sequence is comprised in SEQ ID NO: 14 or a complementary sequence thereof,

4) an oligonucleotide of 12-1348 nucleotides in length which sequence is comprised in SEQ ID NO: 15 or a complementary sequence thereof, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofProteus mirabilis.

21. The method ofclaim 20, wherein the probe for detecting nucleic acid sequences fromProteus mirabilisis selected from the group consisting of SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82 and a sequence complementary thereof.

22. A method for detecting the presence and/or amount ofProteus mirabilisin a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofProteus mirabilisDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, and SEQ ID NO: 15;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofProteus mirabilisin said test sample.

23. The method ofclaim 22, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 74 and SEQ ID NO: 75, and

b) SEQ ID NO: 133 and SEQ ID NO: 134.

24. A method for the detection, identification and/or quantification ofStaphylococcus saprophyticusdirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofStaphylococcus saprophyticus,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofStaphylococcus saprophyticusin said test sample.

25. A method as defined inclaim 24, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-172 nucleotides in length which sequence is comprised in SEQ ID NO: 21 or a complementary sequence thereof,

2) an oligonucleotide of 12-155 nucleotides in length which sequence is comprised in SEQ ID NO: 22 or a complementary sequence thereof,

3) an oligonucleotide of 12-145 nucleotides in length which sequence is comprised in SEQ ID NO: 23 or a complementary sequence thereof,

4) an oligonucleotide of 12-265 nucleotides in length which sequence is comprised in SEQ ID NO: 24 or a complementary sequence thereof, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofStaphylococcus saprophyticus.

26. The method ofclaim 25, wherein the probe for detecting nucleic acid sequences fromStaphylococcus saprophyticusis selected from the group consisting of SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 100, SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, SEQ ID NO: 104 and a sequence complementary thereof.

27. A method for detecting the presence and/or amount ofStaphylococcus saprophyticusin a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofStaphylococcus saprophyticusDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, and SEQ ID NO: 24;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofStaphylococcus saprophyticusin said test sample.

28. The method ofclaim 27, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 98 and SEQ ID NO: 99, and

b) SEQ ID NO: 139 and SEQ ID NO: 140.

29. A method for the detection, identification and/or quantification ofMoraxella catarrhalisdirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 28, SEQ ID NO: 29, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofMoraxella catarrhalis,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofMoraxella catarrhalisin said test sample.

30. A method as defined inclaim 29, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-526 nucleotides in length which sequence is comprised in SEQ ID NO: 28 or a complementary sequence thereof,

2) an oligonucleotide of 12-466 nucleotides in length which sequence is comprised in SEQ ID NO: 29 or a complementary sequence thereof, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofMoraxella catarrhalis.

31. The method ofclaim 30, wherein the probe for detecting nucleic acid sequences fromMoraxella catarrhalisis selected from the group consisting of SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117 and a sequence complementary thereof.

32. A method for detecting the presence and/or amount ofMoraxella catarrhalisin a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofMoraxella catarrhalisDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 28 and SEQ ID NO: 29;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofMoraxella catarrhalisin said test sample.

33. The method ofclaim 32, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 112 and SEQ ID NO: 113,

b) SEQ ID NO: 118 and SEQ ID NO: 119, and

c) SEQ ID NO: 160 and SEQ ID NO: 119.

34. A method for the detection, identification and/or quantification ofPseudomonas aeruginosadirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofPseudomonas aeruginosa,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofPseudomonas aeruginosain said test sample.

35. A method as defined inclaim 34, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-2167 nucleotides in length which sequence is comprised in SEQ ID NO: 16 or a complementary sequence thereof,

2) an oligonucleotide of 12-1872 nucleotides in length which sequence is comprised in SEQ ID NO: 17 or a complementary sequence thereof,

3) an oligonucleotide of 12-3451 nucleotides in length which sequence is comprised in SEQ ID NO: 18 or a complementary sequence thereof,

4) an oligonucleotide of 12-744 nucleotides in length which sequence is comprised in SEQ ID NO: 19 or a complementary sequence thereof,

5) an oligonucleotide of 12-2760 nucleotides in length which sequence is comprised in SEQ ID NO: 20 or a complementary sequence thereof, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofPseudomonas aeruginosa.

36. The method ofclaim 35, wherein the probe for detecting nucleic acid sequences fromPseudomonas aeruginosais selected from the group consisting of SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95 and a sequence complementary thereof.

37. A method for detecting the presence and/or amount ofPseudomonas aeruginosain a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofPseudomonas aeruginosaDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19 and SEQ ID NO: 20;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofPseudomonas aeruginosain said test sample.

38. The method ofclaim 37, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 83 and SEQ ID NO: 84, and

b) SEQ ID NO: 85 and SEQ ID NO: 86.

39. A method for the detection, identification and/or quantification ofStaphylococcus epidermidisdirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 36, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofStaphylococcus epidermidis,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofStaphylococcus epidermidisin said test sample.

40. A method as defined inclaim 39, wherein said probe is selected from the group consisting of an oligonucleotide of 12-705 nucleotides in length which sequence is comprised in SEQ ID NO: 36 and variants thereof which specifically and ubiquitously anneal with strains and representatives ofStaphylococcus epidermidis.

41. A method for detecting the presence and/or amount ofStaphylococcus epidermidisin a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofStaphylococcus epidermidisDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the following sequence: SEQ ID NO: 36;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofStaphylococcus epidermidisin said test sample.

42. The method ofclaim 41, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 145 and SEQ ID NO: 146, and

b) SEQ ID NO: 147 and SEQ ID NO: 148.

43. A method for the detection, identification and/or quantification ofStaphylococcus aureusdirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 37, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofStaphylococcus aureus, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofStaphylococcus aureusin said test sample.

44. A method as defined inclaim 43, wherein said probe is selected from the group consisting of an oligonucleotide of 12-442 nucleotides in length which sequence is comprised in SEQ ID NO: 37 and variants thereof which specifically and ubiquitously anneal with strains and representatives ofStaphylococcus aureus.

45. A method for detecting the presence and/or amount ofStaphylococcus aureusin a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofStaphylococcus aureusDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the following sequence: SEQ ID NO: 37;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofStaphylococcus aureusin said test sample.

46. The method ofclaim 45, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 149 and SEQ ID NO: 150,

b) SEQ ID NO: 149 and SEQ ID NO: 151, and

c) SEQ ID NO: 152 and SEQ ID NO: 153.

47. A method for the detection, identification and/or quantification ofHaemophilus influenzaedirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofHaemophilus influenzae,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofHaemophilus influenzaein said test sample.

48. A method as defined inclaim 47, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-845 nucleotides in length which sequence is comprised in SEQ ID NO: 25 or a complementary sequence thereof,

2) an oligonucleotide of 12-1598 nucleotides in length which sequence is comprised in SEQ ID NO: 26 or a complementary sequence thereof,

3) an oligonucleotide of 12-9100 nucleotides in length which sequence is comprised in SEQ ID NO: 27 or a complementary sequence thereof, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofHaemophilus influenzae.

49. The method ofclaim 48, wherein the probe for detecting nucleic acid sequences fromHaemophilus influenzaeis selected from the group consisting of SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: 107 and a sequence complementary thereof.

50. A method for detecting the presence and/or amount ofHaemophilus influenzaein a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofHaemophilus influenzaeDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 25, SEQ ID NO: 26 and SEQ ID NO: 27;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofHaemophilus influenzaein said test sample.

51. The method ofclaim 50, wherein said at least one pair of primers comprises the following pair: SEQ ID NO: 154 and SEQ ID NO: 155.

52. A method for the detection, identification and/or quantification ofStreptococcus pneumoniaedirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 34, SEQ ID NO: 35, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofStreptococcus pneumoniae,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofStreptococcus pneumoniaein said test sample.

53. A method as defined inclaim 52, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-631 nucleotides in length which sequence is comprised in SEQ ID NO: 30 or a complementary sequence thereof,

2) an oligonucleotide of 12-3754 nucleotides in length which sequence is comprised in SEQ ID NO: 31 or a complementary sequence thereof,

3) an oligonucleotide of 12-841 nucleotides in length which sequence is comprised in SEQ ID NO: 34 or a complementary sequence thereof,

4) an oligonucleotide of 12-4500 nucleotides in length which sequence is comprised in SEQ ID NO: 35 or a complementary sequence thereof, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofStreptococcus pneumoniae.

54. The method ofclaim 53, wherein the probe for detecting nucleic acid sequences fromStreptococcus pneumoniaeis selected from the group consisting of SEQ ID NO: 120, SEQ ID NO: 121 and a sequence complementary thereof.

55. A method for detecting the presence and/or amount ofStreptococcus pneumoniaein a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofStreptococcus pneumoniaeDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 34 and SEQ ID NO: 35;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofStreptococcus pneumoniaein said test sample.

56. The method ofclaim 55, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 78 and SEQ ID NO: 79,

b) SEQ ID NO: 156 and SEQ ID NO: 157, and

c) SEQ ID NO: 158 and SEQ ID NO: 159.

57. A method for the detection, identification and/or quantification ofStreptococcus pyogenesdirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 32, SEQ ID NO: 33, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofStreptococcus pyogenes,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofStreptococcus pyogenesin said test sample.

58. A method as defined inclaim 57, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-1337 nucleotides in length which sequence is comprised in SEQ ID NO: 32 or a complementary sequence thereof,

2) an oligonucleotide of 12-1837 nucleotides in length which sequence is comprised in SEQ ID NO: 33 or a complementary sequence thereof, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofStreptococcus pyogenes.

59. A method for detecting the presence and/or amount ofStreptococcus pyogenesin a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofStreptococcus pyogenesDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 32 and SEQ ID NO: 33;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofStreptococcus pyogenesin said test sample.

60. The method ofclaim 59, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 141 and SEQ ID NO: 142, and

b) SEQ ID NO: 143 and SEQ ID NO: 144.

61. A method for the detection, identification and/or quantification ofEnterococcus faecalisdirectly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, a sequence complementary thereof, a part thereof and a variant thereof, which specifically and ubiquitously anneals with strains or representatives ofEnterococcus faecalis,under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount ofEnterococcus faecalisin said test sample.

62. A method as defined inclaim 61, wherein said probe is selected from the group consisting of:

1) an oligonucleotide of 12-1817 nucleotides in length which sequence is comprised in SEQ ID NO: 1 or a complementary sequence thereof,

2) an oligonucleotide of 12-2275 nucleotides in length which sequence is comprised in SEQ ID NO: 2, and

variants thereof which specifically and ubiquitously anneal with strains and representatives ofEnterococcus faecalis.

63. A method for detecting the presence and/or amount ofEnterococcus faecalisin a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands ofEnterococcus faecalisDNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within one of the following sequences: SEQ ID NO: 1 and SEQ ID NO: 2;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount ofEnterococcus faecalisin said test sample.

64. The method ofclaim 63, wherein said at least one pair of primers is selected from the group consisting of:

a) SEQ ID NO: 38 and SEQ ID NO: 39, and

b) SEQ ID NO: 40 and SEQ ID NO: 41.

65. A method for the detection of the presence and/or amount of any bacterial species directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a universal probe which sequence is selected from the group consisting of SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130 and a sequence complementary thereof, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of the presence and/or amount of said any bacterial species in said test sample.

66. A method for detecting the presence and/or amount of any bacterial species in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing a pair of universal primers which sequence is defined in SEQ ID NO: 126 and SEQ ID NO: 127, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said any bacterial species DNA that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of the presence and/or amount of said any bacterial species in said test sample.

67. A method for evaluating a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene bla_tem(TEM-1) directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 161, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for a β-lactamase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene TEM-1.

68. A method as defined inclaim 67, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 161.

69. A method for evaluating a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene bla_tem(TEM-1) in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for a β-lactamase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 161;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene TEM-1.

70. A method for evaluating a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene bla_rob(ROB-1) directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 162, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for a β-lactamase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene ROB-1.

71. A method as defined inclaim 70, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 162.

72. A method for evaluating a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene bla_rob(ROB-1) in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for a β-lactamase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 162;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene ROB-1.

73. A method for evaluating a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene bla_shv(SHV-1) directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 163, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for a β-lactamase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene SHV-1.

74. A method as defined inclaim 73, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 163.

75. A method for evaluating a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene bla_shv(SHV-1) in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for a β-lactamase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 163;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene SHV-1.

76. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aadB directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 164, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for an aminoglycoside adenylyltransferase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aadB.

77. A method as defined inclaim 76, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 164.

78. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aadB in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for an aminoglycoside adenylyltransferase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 164;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aadB.

79. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC1 directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 165, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC1.

80. A method as defined inclaim 79, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 165.

81. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aaccl in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 165;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC1.

82. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC2 directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 166, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC2.

83. A method as defined inclaim 82, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 166.

84. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC2 in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 166;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC2.

85. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC3 directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 167, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC3.

86. A method as defined inclaim 85, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 167.

87. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC3 in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 167;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacC3.

88. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacA4 directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 168, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacA4.

89. A method as defined inclaim 88, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 168.

90. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacA4 in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 168;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacA4.

91. A method for evaluating a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene mecA directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 169, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for a penicillin-binding protein, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene mecA.

92. A method as defined inclaim 91, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 169.

93. A method for evaluating a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene mecA in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for a penicillin-binding protein that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 169;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to β-lactam antibiotics mediated by the bacterial antibiotic resistance gene mecA.

94. A method for evaluating a bacterial resistance to vancomycin mediated by the bacterial antibiotic resistance genes vanH, vanA and vanX directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 170, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance genes coding for vancomycin-resistance proteins, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to vancomycin mediated by the bacterial antibiotic resistance genes vanH, vanA and vanX.

95. A method as defined inclaim 94, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 170.

96. A method for evaluating a bacterial resistance to vancomycin mediated by the bacterial antibiotic resistance genes vanH, vanA and vanX in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance genes coding for vancomycin-resistance proteins that contain a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 170;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to vancomycin mediated by the bacterial antibiotic resistance genes vanH, vanA and vanX.

97. A method for evaluating a bacterial resistance to streptogramin A mediated by the bacterial antibiotic resistance gene satA directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 173, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for a streptogramin A acetyltransferase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to streptogramin A mediated by the bacterial antibiotic resistance gene satA.

98. A method as defined inclaim 97, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 173.

99. A method for evaluating a bacterial resistance to streptogramin A mediated by the bacterial antibiotic resistance gene satA in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for streptogramin A acetyltransferase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 173;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to streptogramin A mediated by the bacterial antibiotic resistance gene satA.

100. A method for evaluating a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacA-aphD directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 174, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase-phosphotransferase under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacA-aphD.

101. A method as defined inclaim 100, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 174.

102. A method for evaluating a bacterial resistance to aminoglycoside; antibiotics mediated by the bacterial antibiotic resistance gene aacA-aphD in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for an aminoglycoside acetyltransferase-phosphotransferase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 174;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to aminoglycoside antibiotics mediated by the bacterial antibiotic resistance gene aacA-aphD.

103. A method for evaluating a bacterial resistance to virginiamycin mediated by the bacterial antibiotic resistance gene vat directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 175, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for a virginiamycin acetyltransferase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to virginiamycin mediated by the bacterial antibiotic resistance gene vat.

104. A method as defined inclaim 103, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 175.

105. A method for evaluating a bacterial resistance to virginiamycin mediated by the bacterial antibiotic resistance gene vat in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for a virginiamycin acetyltransferase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 175;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to virginiamycin mediated by the bacterial antibiotic resistance gene vat.

106. A method for evaluating a bacterial resistance to virginiamycin mediated by the bacterial antibiotic resistance gene vga directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 176, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for an ATP-binding protein, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to virginiamycin mediated by the bacterial antibiotic resistance gene vga.

107. A method as defined inclaim 106, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 176.

108. A method for evaluating a bacterial resistance to virginiamycin mediated by the bacterial antibiotic resistance gene vga in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for an ATP-binding protein that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 176;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to virginiamycin mediated by the bacterial antibiotic resistance gene vga.

109. A method for evaluating a bacterial resistance to erythromycin mediated by the bacterial antibiotic resistance gene msrA directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 177, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for an erythromycin resistance protein under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of a bacterial resistance to erythromycin mediated by the bacterial antibiotic resistance gene msrA.

110. A method as defined inclaim 109, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 177.

111. A method for evaluating a bacterial resistance to erythromycin mediated by the bacterial antibiotic resistance gene msrA in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for an erythromycin resistance protein that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 177;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of a bacterial resistance to erythromycin mediated by the bacterial antibiotic resistance gene msrA.

112. A method for evaluating potential bacterial resistance to β-lactams, aminoglycosides, chloramphenicol and/or trimethoprim mediated by the bacterial antibiotic resistance gene int directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 171, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for an integrase, under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of potential bacterial resistance to β-lactams, aminoglycosides, chloramphenicol and/or trimethoprim mediated by the bacterial antibiotic resistance gene int.

113. A method as defined inclaim 112, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 171.

114. A method for evaluating potential bacterial resistance to β-lactams, aminoglycosides, chloramphenicol and/or trimethoprim mediated by the bacterial antibiotic resistance gene int in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for an integrase that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 171;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of potential bacterial resistance to β-lactams, aminoglycosides, chloramphenicol and/or trimethoprim mediated by the bacterial antibiotic resistance gene int.

115. A method for evaluating potential bacterial resistance to β-lactams, aminoglycosides, chloramphenicol and/or trimethoprim mediated by the bacterial antibiotic resistance gene sul directly from a test sample or from bacterial colonies, which comprises the following steps:

a) depositing and fixing on an inert support or leaving in solution the bacterial DNA of the sample or of a substantially homogenous population of bacteria isolated from this sample, or

inoculating said sample or said substantially homogenous population of bacteria isolated from this sample on an inert support, and lysing in situ said inoculated sample or isolated bacteria to release the bacterial DNA,

said bacterial DNA being in a substantially single stranded form;

b) contacting said single stranded DNA with a probe, said probe comprising at least one single stranded nucleic acid which nucleotidic sequence is selected from the group consisting of SEQ ID NO: 172, a sequence complementary thereof, a part thereof and a variant thereof, which specifically anneals with said bacterial antibiotic resistance gene coding for a sulfonamide resistance protein under conditions such that the nucleic acid of said probe can selectively hybridize with said bacterial DNA, whereby a hybridization complex is formed, said complex being detected by labelling means, the label being present on said probe or the label being present on a first reactive member of said labelling means, said first reactive member reacting with a second reactive member present on said probe; and

c) detecting the presence or the intensity of said label on said inert support or in said solution as an indication of potential bacterial resistance to β-lactams, aminoglycosides, chloramphenicol and/or trimethoprim mediated by the bacterial antibiotic resistance gene sul.

116. A method as defined inclaim 115, wherein said probe comprises an oligonucleotide of at least 12 nucleotides in length which hybridizes to SEQ ID NO: 172.

117. A method for evaluating potential bacterial resistance to β-lactams, aminoglycosides, chloramphenicol and/or trimethoprim mediated by the bacterial antibiotic resistance gene sul in a test sample which comprises the following steps:

a) treating said sample with an aqueous solution containing at least one pair of oligonucleotide primers having at least 12 nucleotides in length, one of said primers being capable of hybridizing selectively with one of the two complementary strands of said bacterial antibiotic resistance gene coding for a sulfonamide resistance protein that contains a target sequence, and the other of said primers being capable of hybridizing with the other of said strands so as to form an extension product which contains the target sequence as a template, said at least one pair of primers being chosen from within the sequence defined in SEQ ID NO: 172;

b) synthesizing an extension product of each of said primers which extension products contain the target sequence, and amplifying said target sequence, if any, to a detectable level; and

c) detecting the presence and/or amount of said amplified target sequence as an indication of potential bacterial resistance to β-lactams, aminoglycosides, chloramphenicol and/or trimethoprim mediated by the bacterial antibiotic resistance gene sul.

118. A nucleic acid having the nucleotide sequence of any one of SEQ ID NOs: 1 to 37, SEQ ID NOs: 161 to 177, a part thereof and variants thereof which, when in single stranded form, ubiquitously and specifically hybridize with a target bacterial DNA as a probe or as a primer.

119. An oligonucleotide having a nucleotidic sequence of any one of SEQ ID NOs: 38 to 160.

120. A recombinant plasmid comprising a nucleic acid as defined inclaim 118.

121. A recombinant host which has been transformed by a recombinant plasmid according toclaim 120.

122. A recombinant host according toclaim 121 wherein said host isEscherichia coli.

123. A diagnostic kit for the detection and/or quantification of the nucleic acids of any combination of the bacterial species defined in any one of claims9,14,19,24,29,34,39,43,47,52,57 and61, comprising any combination of probes defined therein.

124. A diagnostic kit for the detection and/or quantification of the nucleic acids of any combination of the bacterial species defined in any one of claims10,11,15,16,20,21,25,26,30,31,35,36,40,44,48,49,53,54,58,62 and65, comprising any combination of oligonucleotide probes defined therein.

125. A diagnostic kit for the detection and/or quantification of the nucleic acids of any combination of the bacterial species defined in any one of claims12,13,17,18,22,23,27,28,32,33,37,38,41,42,45,46,50,51,55,56,59,60,63,64 and66 comprising any combination of primers defined therein.

126. A diagnostic kit for the detection and/or quantification of the nucleic acids of any combination of the bacterial resistance genes defined in any one of claims67,70,73,76,79,82,85,88,91,94,97,100,103,106 and109 comprising any combination of probes defined therein.

127. A diagnostic kit for the detection and/or quantification of the nucleic acids of any combination of the bacterial resistance genes defined in any one of claims68,71,74,77,80,83,86,89,92,95,98,101,104,107 and110 comprising any combination of oligonucleotide probes defined therein.

128. A diagnostic kit for the detection and/or quantification of the nucleic acids of any combination of the bacterial resistance genes defined in any one of claims69,72,75,78,81,84,87,90,93,96,99,102,105,108 and111 comprising any combination of primers defined therein.

129. A diagnostic kit for the simultaneous detection and quantification of nucleic acids of any combination of the bacterial species defined inclaim 123, comprising any combination of the bacterial probes defined therein and any combination of the probes to the antibiotic resistance genes defined in any one of SEQ ID NOs: 161 to 177 in whole or in part.

130. A diagnostic kit for the simultaneous detection and quantification of nucleic acids of any combination of the bacterial species defined inclaim 124, comprising any combination of the bacterial oligonucleotide probes defined therein and any combination of oligonucleotide probes that hybridize to the antibiotic resistance genes defined in any one of SEQ ID NOs: 161 to 177.

131. A diagnostic kit for the simultaneous detection and quantification of nucleic acids of any combination of the bacterial species defined inclaim 125, comprising any combination of the primers defined therein and any combination of primers that anneal to the antibiotic resistance genes defined in any one of SEQ ID NOs: 161 to 177.