NUCLEIC ACID PROBES FOR THE DETECTION OF CHLAMYDIA TRACHOMATIS

Abstract: Nucleic acid probes capable of hydridizing to rRNA of Chlamydia trachomatis and not to rRNA of non-Chlamydia are described along with methods utilizing such probes for the detection of Chlamydia trachomatis in clinical samples.

Description: NUCLEIC ACID PROBES FOR THE DETECTION OF CHLAMYDIA TRACHOMATIS Field of the Invention This invention relates to detecting bacteria belonging to the species Chlamydia trachomatis and more specifically provides nucleic acid probes and compositions along with methods for their use for the specific detection of Chlamydia trachomatis. Background of the Invention The term "Chlamydia trachomatis" as used herein, refers to bacteria classified as such in Bergey's Manual of Systematic Bacteriology (P. H. A. Sneath, ed., 1986, PP. 729-736, Williams & Wilkins). Chlamydia trachomatis is a member of the genus Chlamydia, which has only one other member, Chlamydia psittaci. Chlamydia are a unique group of intracellular parasites. Chlamydia has a complex life cycle that is characterized by tvo forms, the elementary body (EB) which like a virus is infectious but not metabolically active, and the reticulate body (RB) which is metabolically active, not infectious and divides by binary fission like bacteria. The RB eventually matures into the EB which is released by the infected cell, a process which take about 2 days. Because of their role in ocular, respiratory and sexually transmitted diseases, these organisms are a major health risk through the world (Nichols, R. L. and Manire, G. P.: Chlamydiae, in Microbiology 3rd Ed, Davis, B. D., Dulbecco, R., Eisen, H. N. and Ginsburg, H. S. {ed) . 1980, 776-784, Harper & Row). C. trachomatis is responsible for some 350 million cases of trachoma in the developing world (Dawson, C. R., Proceedings of the Fifth International Symposium on Human Chlamydial Infections. Lund, Sweden, June 15-19, 1982, Mardh, P. A., Holmes, K. K. , Oril, J. D., Piot, P., Schachter, J., [edj, 1982, 71-81, Elsevier Biomedical Press, Amsterdam, New York, Oxford). This is particularly tragic since trachoma is also the leading cause of preventable blindness (Jones, B. R. , 1975, Trans. Opthalmol. Soc. UK, 95: 16-33). Infection by this organism is now recognized as the most common sexually transmitted disease in the industrialized world (Centers for Disease Control, Division of sexually transmitted disease. Chlamydia trachomatis infections. Policy guidelines for prevention and control. MMWR 1985; 34: 53S-74S), accounting for more than 4 million cases in the United States alone (Judson FN. 1985, J.Reprod. Med. 30: 269-272), Direct and indirect costs run into billions of dollars. Among the more serious complications of infection are ecotropic pregnancy and tubal infertility (Washington et al-, 1987, JAMA 257: 2070-2072). These two manifestations of C. trachomatis infection reflect two further groupings: the trachoma biovar which causes ocular end genital infections and the lymphogranuloma veretrum (LGV) biovar. These tvo biovars are further subdivided in 15 serotypes: A, B, Ba, C (usually associated with follicular conjunctivitis), D through K, and LI through L3. Diagnosis of the disease is often lengthy and labor intensive. Cell culture is considered the "gold standard" for chlamydial diagnostics. Because of the time-consuming and tedious nature of cell culture, and the difficulty of growing many clinical isolates in tissue culture, a number of rapid laboratory methods have recently become available (see reviev by Barnes RC, 1989, Clin. Microbiol. Rev. 2: 119-136). The vast majority are antibody-based tests. While in certain instances these rapid assays have proven to be more sensitive or specific than cell culture, their utility lies in their rapidity. It is an aspect of the present invention to provide nucleic acid probes which are specific for Chlamydia, which do not react with other bacteria or fungi which may be present in sampled materials, and which nay be used in a variety of assay systems. It is another aspect of the present invention to provide probes which can hybridize to target regions which can be rendered accessible to probes under normal assay conditions. It is yet another aspect of the present invention to provide assays which avoid many of the disadvantages associated with traditional, multi-day culturing techniques. While Kohne et al. (Biophysical Journal 8:1104-1118, 1968) discuss one method for preparing probes to rRNA sequences, they do not provide the teaching necessary to make Chlamydia trachomatis specific probes. Pace and Campbell (Journal of Bacteriology 107:543-547, 1971) discuss the homology of .ribosomal ribonucleic acids from diverse bacterial species and a hybridization method for quantitating such homology levels. Similarly, Sogin, Sogin and Woese (Journal of Molecular Evolution 1:173-184, 1972) discuss the theoretical and practical aspects of using primary structural characterization of different ribosomal RNA molecules for evaluating phylogenetic relationships. Fox, Pechman and Woese (International Journal of Systematic Bacteriology 27:44-57, 1977) discuss the comparative cataloging of 16S ribosomal RNAs as an approach to prokaryotic systematics. These references, however, fail to relieve the deficiency of Kohne's teaching with respect to Chlamydia trachomatis and in particular, do not provide Chlamydia trachomatis specific probes useful in assays for detecting Chlamydia trachomatis in clinical and other samples. Ribosomes are of profound importance to all organisms because they serve as the only known means of translating genetic information into cellular proteins, the main structural and catalytic elements of life. A clear manifestation of this importance is the observation that all cells have ribosomes. Riboso es contain three distinct RNA molecules which, at least in Escherichia coli, are referred to as 5S, 16S and 23S rRNAs. These names historically are related to the size of the RNA molecules, as determined by their sedimentation rate. In actuality, however, ribosomal RNA molecules vary substantially in size between organisms. Nonetheless, 5S, 16S, and 23S rRNA are commonly used as generic names for the homologous RNA molecules in any bacteria, and this convention vill be continued herein. As used herein, probe(s) refer to synthetic or biologically produced nucleic acids (DNA or RNA) which, by design or selection, contain specific nucleotide sequences that allow them to hybridize under defined predetermined stringencies, specifically (i.e., preferentially, see below - Hybridization) to target nucleic acid sequences. In addition to their hybridization properties, probes also may contain certain constituents that pertain to their proper or optimal functioning under particular assay conditions. For example, probes may be nodified to carry detection ligands (e.g. fluorescien, 32-P, biotin, etc.), or to facilitate their capture onto a solid support (e. g., poly-deoxyadenosine "tails"). Such modifications are elaborations on the basic probe function which is its ability to usefully discriminate between target and non-target organisms in a hybridization assay. Hybridization traditionally is understood as the process by which, under predetermined reaction conditions, two partially or completely complementary strands of nucleic acid are allowed to come together in - an antiparallel fashion to form a double-stranded nucleic acid with specific and stable hydrogen bonds. The stringency of a particular set of hybridization conditions is defined by the length and base composition of the probe/target duplex, as veil as by the level and geometry of mispairing between the two nucleic acids. Stringency may also be governed by such reaction parameters as the concentration and type of ionic species present in the hybridization solution, the types and concentrations of denaturing agents present, and/or the temperature of hybridization. Generally, as hybridization conditions become more stringent, longer probes are preferred if stable hybrids are to be formed. As a corollary, the stringency of the conditions under which a hybridization is to take place (e. g. , based on the type of assay to be performed) will dictate certain characteristics of the preferred probes to be employed. Such relationships are well understood and can be readily manipulated by those skilled in the art. As a general matter, dependent upon probe length, such persons understand stringent conditions to mean approximately 35°C-65°C in a salt solution of approximately 0.9 molar. Summary of the Invention In accordance with the various principles and aspects of the present invention, there are provided nucleic acid probes and probe sets comprising DNA or RNA sequences which hybridize, under specific conditions, to the ribosomal RNA molecules (rRNA) or rRNA genes (rDNA) of Chlamydia trachomatis but which do not hybridize, under the same conditions, to the rRNA or rDNA of other related bacteria which may be present in test samples. Therefore the probe(s) of the present invention provide the basis for development of a valuable nucleic acid hybridization assay for the specific detection of Chlamydia trachomatis in clinical samples. In our experience such nucleic acid hybridization based assays have been discovered to impart enhanced performance capabilities with respect to most currently available, microbiological methods for detection of bacteria in test samples, generally including: a) increased sensitivity; i. e. f the ability to detect fewer said bacteria in a given sample; b) potentially significant reductions in assay cost due to the use of inexpensive reagents and reduced labor; c) accurate identification of even biochemically unusual strains of the target bacteria; d) faster results because such tests do not require the isolation of the target bacterium from the sample prior to testing. It has been discovered that other advantages incurred by directing the probes of the present invention against rRNA include the fact that the rRNAs detected constitute a significant component of cellular mass. Although estimates of cellular ribosome content vary, actively growing E. coli nay contain upwards of 50,000 ribosomes per cell, and therefore 50,000 copies of each of the rRNAs (present in a 1:1:1 stoichiometry in ribosomes). Chlamydia trachomatis elementary bodies are known to contain far fever ribosomes (hundreds). Nevertheless, this number is still larger than most other potential cellular target molecules such as genes or RNA transcripts thereof, which are less ideal since they are present in much lower abundance. A further unexpected advantage is that the rRNAs (and the genes specifying them) appear not to be subject to lateral transfer between contemporary organisms. Thus, the rRNA primary structure provides an organism-specific molecular target, rather than a gene-specific target as would likely be the case, for example of a plasmid-borne gene or product thereof vhich may be subject to lateral transmission between contemporary organisms. Additionally, the present invention provides probes to Chlamydia trachomatis rRNA target sequences which are sufficiently similar in all Chlamydia trachomatis strains tested that they can hybridize to the target region in all such Chlamydia trachomatis. Advantageously, these same rRNA target sequences are sufficiently different in most non- Chla ydia trachomatis rRNAs that, under conditions where probes 781, 860, 861, 879, 1153, 1203, 1220, 1318, 1319, 1320, 1321, 1322, 1323, 1325 and 1479 hybridize to Chlamydia trachomatis rRNAs and they do not hybridize to most non-Chlamydia trachomatis rRNAs. These probe characteristics are defined as inclusivity and exclusivity, respectively. The other probes of the present invention, probes 783, 882, and 1324 are fully as inclusive for Chlamydia trachomatis strains as the probes mentioned above and, in addition, these probes also hybridize to a few other bacteria. Probe 782 is the only probe which hybridizes to Chlamydia psittaci and not to Chlamydia trachomatis. The discovery that probes could be generated with the extraordinary inclusivity and exclusivity characteristics of those of the present invention with respect to Chlamydia trachomatis was unpredictable and unexpected. Brief Description of the Tables Further understanding of the principles and aspects of the present invention may be made by reference to the tables wherein: Table 1 - Shows the nucleotide sequences of the preferred 16S rRNA- targeted probes of the present invention aligned upon their Chlamydial target nucleotide sequences. The corresponding portions of the 16S rRNA from E. coli is shown for comparison. RNA (target) sequences are written 5' to 3', probe sequences are DNA and written 3' to 5'. Probes are shown along with the "core" region of variation upon which their inclusivity and exclusivity behavior are based. Table 2 - Shows the nucleotide sequences of the preferred 23S rRNA- targeted probes of the present invention aligned upon their Chlamydial target nucleotide sequences. The corresponding portions of the 23S rRNA from E. coli is shown for comparison. RNA (target) sequences are written 5' to 3', probe sequences are DNA and written 3' to 5'. Probes are shown along with the "core" region of variation upon which their inclusivity and exclusivity behavior are based. Table 3 - Exemplifies the inclusivity behavior of the preferred 16S and 23S rRNA probes toward a representative. sampling of Chlamydia trachomatis serovars in a liquid-hybridization assay. Table 4 - Exemplifies the exclusivity behavior of the preferred 16S and 23S rRNA probes toward a representative sampling of non-Chlamydia trachomatis bacteria in a dot blot hybridization assay. Detailed Description of the Invention and Best Mode Probe Development Strategy. 'The first step taken in the development of the probes of the present invention involved identification of regions of 16S and 23S rRNA which potentially could serve as target sites for Chlamydia trachomatis specific nucleic acid probes. As a practical matter, it is difficult to predict, a priori, which non-Chlamydia trachomatis organisms might be present in any test sample. Because of the large number of such potential non-Chlamydia trachomatis bacteria, demonstrating exclusivity for any given probe sequence is not only unpredictable but also extremely difficult and laborious. A more rigorous criterion was adopted to obviate the need to know what non- Chlamydia trachomatis bacteria might be present in all test samples that ultimately will be screened using the probes. This entailed knowledge of the phylogenetic relationships among Chlamydia trachomatis and between Chlamydia trachomatis and other groups of bacteria. Specifically, an operating but previously unproven hypothesis was adopted that the exclusivity criterion could be satisfied by determining that if a particular target region in Chlamydia trachomatis rRNA could be identified which was sufficiently different from the homologous region in the rRNA of representative yet close evolutionary relatives of Chlamydia trachomatis, then a probe to such a sequence could be used to distinguish between Chlamydia trachomatis and the relatives by hybridization assay. Based on phylogenetic observations, it then was extrapolated that rRNA sequences of more distantly related organisms, even though their actual identity may not necessarily be known, should be predictably different in a particular region of sequence than the aforementioned close evolutionary relative of Chlamydia trachomatis. However, it cannot be predicted, a priori, whether such regions exist or if they do, vhere within the rRNA such regions will be located. As the first step in identifying regions of Chlamydia trachomatis rRNA which could potentially serve as useful target sites for nucleic acid hybridization probes, complete nucleotide sequences of the 16S and 23S rRNAs from Chlamydia trachomatis was determined. The nucleotide sequences were determined by standard laboratory protocols either by cloning (Maniatis et al., 1982, Molecular Cloning; A Laboratory Manual, Cold Spring Harbor Laboratory, New York, pp 545) and sequencing (Maxam and Gilbert, 1977, Proceedings of the National Academy of Science, USA 74:560-564: Sanger et al. , 1977, Proceedings of the National Academy of Science, USA 74:5463-5467) the genes which specify the rRNAs, and/or by direct sequencing of the rRNAs themselves using reverse transcriptase (Lane et al. , 1985, Proceedings of the National Academy of Science, USA 82:6955-6959). The determined Chlamydia trachomatis rRNA nucleotide sequences were compared to other available rRNA nucleotide sequences, in particular to Chlamydia psittaci, which also was determined as part of this work. Comparison of the sequences of Chlamydia trachomatis and its close relative Chlamydia psittaci proved especially valuable. Several regions of sequence were identified which appeared to be different in the two species of Chlamydia and between Chlamydia trachomatis and non- Chlamydia bacteria. The locations of these regions within the 16S and 23S rRNA sequences are shown in Tables 1 and 2, respectively. Oligonucleotide probes, 28 - 36 nucleotides in length, were designed which would hybridize preferentially to Chlamydia trachomatis, Chlamydia psittaci or both. These were designed : 1) to maximally utilize the nucleotide sequence differences useful for distinguishing Chlamydia trachomatis from Chlamydia psittaci or Chlamydia from other bacteria (indicated as upper case letters in the region of Core Variation, Tables 1 and 2) and, 2) to minimize the effect of self complementarity both locally within the target rRNA and between probe molecules. Optimizing these parameters as well as others discussed above (Background) results in probes of preferred specificity and hybridization efficiency. Table 3 exemplifies the inclusivity behavior of the preferred probes toward a representative sampling of Chlamydia trachomatis and a few non-Chlamydia trachomatis bacteria in a liquid hybridization assay. Table 4 exemplifies the exclusivity behavior of the preferred probes toward a representative sampling of non-Chlamydia trachomatis bacteria in a dot-blot hybridization assay. Physical Description of the Probes. The foregoing probe selection strategy yielded a number of probes useful for identifying Chlamydia trachomatis bacteria in samples. Probes 781, 782, 783, 860, 861, 879, 882, 1153 and 1203 are designed from 16S rRNA sequences and probes 1318 through 1325 and probe 1479 are designed from 23S rRNA sequences. The following preferred oligonucleotide probes are disclosed herein: 16S rRNA-Targeted Probes: Probe 781: 5'-CTTTAACGTTACTCGGATGCCCAAATATCGCCACAT-3' Probe 782: 5'-CTTTAATATGTTTTAGATGCCTAAACATACCACAT-3' Probe 783: 5'-CGGAAAACGACATTTCTGCCGCGGTCAAATACATG-3' Probe 860: 5- * -CGCTCAAATCCAGCGGGTATTAACCGCCTTCTC-3' Probe 861: 5'-GCCGACTCGGGGTTGAGCCCCGATCTTTGACAA-3' Probe 879: 5'-CGGATGGGGTTGAGACCATCCACATCAA-3' Probe 882: 5'-TGTGTATATGTCCTTGCGGAAAACGACATTTCTGC-3' Probe 1153: 5'-CCACTAAACAATCGTCGAAACAATTGCTCCGTTCG-3' Probe 1203: 5'-CCACTAAACAATTGCCGAAACAATTGCTCCGTTCG-3' 23S rRNA-Targeted Probes: Probe 1220: 5' -CCGGGGCTCCTATCGTTCCATAGTCACCCTAAAAG-3' Probe 1318: 5'-TACCTCCGGGTCTTTGCTTATCACCAGCTCGCC-3' Probe 1319: 5'-GTATTCAGCATGCAATGGTAGTCTATTACTCTA-3' Probe 1320: 5'-TCGGCAGGTGTCGCTTTGCATACCTATGTATTC-3' Probe 1321: 5'-CGAGCCTTATCAGCTCGGTTTAGGCTATTCCCC-3' Probe 1322: 5'-AACTAGGAGTCCTGATCCTTTATCCTCAATCCT-3' Probe 1323: 5'-TCAGGTGTTGAGGTCGGTCTTTCTCTCCTTTCG-3' Probe 1324: 5'-AGATTCCCCTTGATCGCGACCTGATCTTATGTT-3' Probe 1325: 5'-AACCGTTCTCATCGCTCTACGGACTCTTCCAAT-3' Probe 1479: 5'-CTCCTACCGCGTGTCTTATCGACACACCCGCGA-3' The probes and their target sequences in the 16S and 23S rRNAs of Chlamydia trachomatis and Chlamydia psittaci are shown in Tables 1 and '2. The corresponding nucleotide positions of the E. coli 16S and 23S rRNAs also are shown. Since the E. coli sequences were among the first full 16S and 23S sequences obtained, the assigned position numbers are a convenient standard for explicitly identifying the homologous regions in the Chlamydia rRNAs under consideration. More than one probe has been designed to a number of the target regions shown in Tables 1 and 2 corresponding variously to complements of the sequences of Chlamydia trachomatis serovars LGV or K, or Chlamydia ps ttaci, or all three. The particular sequence upon which each probe is based (i.e., is complementary to) is provided in Tables 1 and 2 by inspection of the aligned probe and target sequences. Thus, for example, it can be seen in Table 1 that probe 1203 is complementary to the Chlamydia trachomatis serovar LGV sequence through this target region and the related probe 1153 is complementary to the C. trachomatis serovar K sequence. However, it is expected (and desirable) that some cross-hybridization between serovars by one or both probes will take place. Likewise, probes 781 and 782 are based on Chlamydia trachomatis (LGV is identical to K in this region) and Chlamydia psittaci 16S rRNA sequences, respectively. The specific hybridization behaviors of the probes described above are dependent to a significant extent on the assay format, in which they are employed. Conversely, the assay format vill dictate certain of the optimal design features of particular probes. The "essence" of the probes of the invention is not to be construed as restricted to the specific string of nucleσtides in the probes named above. For example, the length of these particular oligonucleotides vas optimized for use in the dot blot assay (and certain other anticipated assays) described below. It is well known to one skilled in the art that optimal probe length will be a function of the stringency of the hybridization conditions chosen and hence the length of the instant probes may be altered in accordance therewith. Also, in considering sets comprised of more than one probe, it is desirable that all probes behave in a compatible manner in any particular format in which they are both employed. Thus, the exact length of a particular probe will to a certain extent reflect its specific intended use. The "essence" of the probes described herein resides in the discovery and utilization of the Chlamydia trachomatis specific sequences described above and given in Tables 1 and 2 (Core Variation). Hybridization Analysis of Probe Behavior. The sequence data in Tables 1 and 2 suggest that the probes of the present invention should exhibit a variety of useful hybridization properties with respect to the specific detection of Chlamydia trachomatis, Chlamydia psittaci or both to the exclusion of other bacteria. However, relatively few Chlamydia trachomatis and Chlamydia psittaci sequences vere inspected. It is possible that sequence variation might exist in other Chlamydia trachomatis serotypes not inspected by sequence analysis. Such variation might reduce or eliminate hybridization by the prospective probes to some or many untested Chlamydia trachomatis serotypes. Equally as important as the inclusivity behavior of the probes, is their exclusivity behavior, i.e., their reactivity toward non-Chlamydia bacteria. The number and types of non-Chlamydia strains which might be encountered in a potentially Chlamydia containing test sample are extremely large. Therefore, the behavior of the probes toward representative Chlamydia trachomatis and non-Chlamydia trachomatis bacteria was determined by hybridization analysis using a liquid hybridization and a dot blot procedures. While hybridization data for each of the individual probes of the present invention are provided, it should be noted that useful combinations (sets) of probes which exhibit hybridization behaviors that are the sum of the individual probes also is explicitly predicted by the data. Example 1: Inclusivity Analysis of probe hybridization behaviour in liquid-hybridization assay. The probes of the present invention or derivatives thereof could potentially be of significant value in a variety of hybridization formats. One such format, a dual probe, sandwich-type hybridization assay formats (e.g. the homopolymer capture, dual probe, liquid hybridization format described in USSN 277, 579; USSN 169,646, or USSN 233,683), is used in this example. In general in such an application, an oligonucleotide probe is modified at its 3' terminus to contain a tract of deoxyadenosine (dA) residues ca. 20 - 200 residues long. This would be used to "capture" the target rRNA (following liquid hybridization) from the test sample onto a solid support (e.g., beads, plastic surface, filter, etc.) which had been suitably derivatized with poly-deoxythymidine (dT) for this purpose. A second probe is used as a "detection" probe and would be derivatized by some detectable ligand (e.g. 32-P, fluorescien, biotin, etc.). In principle, the detection probe could be an oligonucleotide or a longer DNA or RNA probe. Detection of the presence of the target nucleic acid in a test sample then is indicated by the capture of the detection ligand onto the solid surface through the series of hybridization interactions: SOLID I \ (TARGET NUCLEIC ACID) / JTTTTTTTTn \ / SUPPORT I 111111 1111111111111 111111111111 I 11 nAAAAAAAAA(Capture Probe) (Detection Probe) i I Ligand Ligand This could occur only if the target nucleic acid is present in the test sample. In this example, each oligonucleotide individually is used as a capture probe (for this purpose ca. 200-500 dA residues were appended to their 3' termini using terminal deoxynucleotidyl transferase). P-32 labeled "riboprobes," specific for either 16S or 23S rRNA, were prepared as described below and used as "generic" detection probes. Both riboprobes were prepared by transcription from plasmid vectors containing portions of the E. coli (16S rRNA) or C. psittaci (23S rRNA) genes. The 16S riboprobe is an antisense transcript, ca. 567 nucleotides long, generated from a Hindlll subfragment of the E. coli 16S rRNA gene (E. coli positions 1 to 567). The 23S riboprobe is an antisense transcript, ca. 855 nucleotides long, generated from a Pstl- EcoRI portion of the C. psittaci 23S rRNA gene (E. coli positions 1 to 855). Radioactive phosphorous 32 is incorporated into the riboprobes during the transcription reactions by using P-32 substituted nucleotide triphospbate precursors according to standard protocols. Detection of the presence of the target nucleic acid in the test sample then is indicated by the capture of the detection ligand (P-32) onto the solid surface through series of hybridization interactions described above. The inclusivity behaviors of the oligonucleotide probes were tested in a version of the liquid hybridization format discussed above. The 16S and 23S riboprobes are expected to hybridize to all Chlamydia and non-Chlamydia 16S and 23S rRNAs, respectively. Therefore, a positive hybridization signal in this example assay in indicative of the hybridization behavior of the oligonucleotide probe employed in each experiment. Alternatively, these oligonucleotide probes could have been used to capture and detect P-32-labeled Chlamydia 16S and 23S rRNAs prepared, for example, by growth of the Chlamydia strains in radioactive phosphorous. Fifteen serotypes of Chlamydia trachomatis have been tested. Serovar K was obtained from the American Type Culture Collection (ATCC). The rest (serovars A-LGV), were obtained from the Washington Research Foundation. Briefly, formalin-fixed Elementary Bodies (EBs) of the indicated serovars of Chlamydia trachomatis were lysed in a solution of 1.0 mg/ml proteinase-K (Boehranger Mannheim Corp.) and 1.6% Sarkosine (Sigma) at 65 β C for 15 minutes (final volume of 0.07ml). The samples were removed from the incubator and an equal volume of 5.0 M GuSCN (in 0.1H Tris-HCl pH 7.0, 10% dextran sulphate) containing poly-dA tailed capture probe and P-32 labeled detector probe (either 16S or 23S riboprobes appropriate to each oligonucleotide probe being tested) at a concentration of 80 ng/ml and 30 ng/ml (specific activity 1 x 10^ cpm/ug) was added. Hybridization of the probes to the target nucleic acids was allowed to proceed at 37°C for 30 minutes after which time the target complex (dA capture probe : target : detector probe ) was captured onto oligo-dT derivatized magnetic beads. This capture was accomplished by adding one tenth of a milliliter (0.1 ml) of a mixture containing 0.0625% (w/v) magnetic beads, 4% BSA, 10 mM EDTA, 0.2% Sarkosine, 0.1M Tris-HCl pH 8.0, and 0.05% Bronopol to the hybridization reaction and incubating the mixture for 5 minutes at 37 β C. Following the capture of the probe/target complexes onto the beads, 0.2 ml of a solution containing 1.0M GuSCN, lOmM EDTA, 0.1M Tris pH 7.0, 0.5% Sarkosine, 0.2% BSA, and 0.1% antifoam was added to the reaction tube, and the tube placed in a magnetic field. The beads (with the attached probe/target complexes) were drawn to the sides of the tube by the magnetic field and the liquid phase (containing the non-hybridized target and detector molecules) was removed from the tube by aspiration. The captured target complex remaining in the tube was washed in this manner 2 more times. The separated beads finally were suspended in 0.1 ml of a solution containing 3.25H GuSCN, 65mM EDTA, 0.1M Tris-HCl, pH 7.0, 0.5% Sarkosine and 0.5% BSA and incubated at 37 e C for 5 minutes. This treatment releases the target complex from the bead via the "melting" of the dT:dA bond between the bead and the dA-tailed capture probe. The solution was removed and mixed with a fresh aliquot of beads (0.1 ml) and the target complex was "recaptured" at 37°C for 5 minutes as described above. The wash procedure described above also was repeated. After the third wash and separation, the beads were resuspended in 0.1 ml of the wash buffer and the beads were filtered onto a nitrocellulose membrane and exposed to x-ray film. The extent of hybridization of each probe was determined by visual inspection of the exposed x-ray film. The results are shown in Table 3. ++++ indicates a hybridization signal equivalent to that of the "control" Chlamydia trachomatis serovar for which a perfect match between probe and target sequences has been explicitly determined by sequence analysis. +++, ++, + and - represent, respectively, very strong, strong, weak and undetectable hybridization signals. Hybridization of the probes to approximately 1 x 10-* of the non- Chlamydia (negative control) bacteria shown in Table 3 was scored in the same fashion. Example 2: Dot blot analysis of probe hybridization behavior. Dot blot analysis, in accordance with well known procedures, involves immobilizing a nucleic acid or a population of nucleic acids on a filter such as nitrocellulose, nylon, or other derivatized membranes which readily can be obtained commercially, specifically for this purpose. Either DNA or RNA can be easily immobilized on such a filter and subsequently can be probed or tested for hybridization under any of a variety of conditions (i.e., stringencies) with nucleotide sequences or probes of interest. Under stringent conditions, probes whose nucleotide sequences have greater complementarity to the target sequence will exhibit a higher level of hybridization than probes containing less complementarity. For the experiment shown in Table 4, one tenth of a microgratn of purified RNA (Lane e_t al. , 1985, Proceedings of the National Academy of Science, USA 82:6955-6959) from each of the indicated organisms was spotted on nitrocellulose filters. The oligo- nucleotide probes were end-labeled with radioactive phosphorous 32, using standard procedures. For the oligonucleotide probes described herein, hybridization to rRNA targets at 60°C for 14-16 hours (in a hybridization solution containing 0.9 M NaCl, 0.12 M Tris-HCl, pH 7.8, 6 mM EDTA, 0.1 M KP04, 0.1% SDS, 0.1% pyrophosphate, 0. 002% ficoll, 0.02% BSA, and 0.002% polyvinylpyrrolidine), followed by three 15 minute post-hybridization washes at 60°C (in 0.03 M NaCl, 0.004 M Tris-HCl, pH 7.8, 0.2 mM EDTA, and 0.1% SDS) to remove unbound probes, would be sufficiently stringent to produce the levels of specificity demonstrated in Table 4. Following hybridization and washing as described above, the hybridization filters were exposed to X-ray film and the intensity of the signal "scored" visually with respect to control spots of known amount of target material (RNA) as described above (Example 1). While the description of the invention has been made with reference to detecting rRNA, it will be readily understood that the probes described herein and probes complementary to those described herein also will be useful for the detection of the genes (DNA) which specify the rRNA and, accordingly, such probes are to be deemed equivalents to the described probes and encompassed vithin the spirit and scope to the present invention and the appended claims. TABLE It CHLAMYDIA 16S rRNA PROBES AND TARGET SEQUENCES Pos. * (E. coli) 63 105 I i Escherichia coli GUCGAACGGUAACAGGAAGAAGCUUGCUUCUUUGCUGACGAGUGGCG Chlamydia psittaci GUCGAACGGAAUAAUGACUU CGGUUGUUAUUUAGUGGCG C. trachomatis LGV GUCGAACGGAGCAAUUGUUU CGGCAAUUGUUUAGUGGCG C. trachomatis K GUCGAACGGAGCAAUUGUUU CGACGAUUGUUUAGUGGCG Core Variation GUuu cgRCRAuuG Probe 1203 GCTTGCCTCGTTAACAAA GCCGTTAACAAATCACC-5' Probe 1153 GCITGCCTCGTTAACAAA GCTGCTAACAAATCACC-5' Pos. # (Ξ. coli) 179 198 I I Escherichia coli GCAUAACGD-CGC AAGACC AAAGAG Chlamydia psittaci GAAUGUGGU-AUGUUUAGGCAUCUAAAACAUAUUAAAGAA C. trachomatis LGV GAAUGUGGCGAUAOUUGGGCAUCCGAGUAACGUUAAAGAA C. trachomatis K GAAUGUGGCGAUAUUUGGGCAUCCGAGUAACGUUAAAGAA Core Variation A-auGuuuAggcaucUAaAACaUA Probe 781 TACACCGCTATAAACCCGTAGGCTCATTGCAAI TC-5' Pos. # (E. coli) 453 485 I Escherichia coli AAGGGAGUAAAGUUAAUACCUUUGCUCAUUGACGUUA Chlamydia psittaci AAGAGAGAUUGGCUAAUAUCCAAUCGAUUUGAGCGUA C. trachomatis LGV AAGAGAAGGCGGUUAAUACCCGCUGGAUUUGAGCGUA Core Variation AGGCggUuaauaCccGCuG Probe 860 CTCTTCCGCCAATTATGGGCGACCTAAACTCGC-5 ' TABLE 1 (cont'd) j CHLAMYDIA 16S rRNA PROBES AND TARGET SEQUENCES Pos. # (E. coli) 602 634 I I Escherichia coli UCAGAUGUGAAAUCCCCGGGCUCAACCUGGGAACUGCAU Chlamydia psittaci UUAGAUGUUAAAUCUUGGGGCUCAACCCCAAGCCAGCAU C. trachomatis LGV UUAGUUGUCAAAGAUCGGGGCUCAACCCCGAGUCGGCAU C. trachomatis K UUAGUUGUCAAAGAUCGGGGCUCAACCCCGAGUCGGCAU Core Variation CaaaGAuCggrggcucaaccccGagUcG Probe 861 AACAGTTTCTAGCCCCGAGTTGGGGCTCAGCCG-5' Pos. # (E. coli) 827 853 I I Escherichia coli ACUUGGAGGUUGUGCCCU-UGAGGCGUGGCUU i Chlamydia psittaci ACUUGAUGUGGAUAGUCUCAACCCUAUCCGUG C. trachomatis LGV ACUUGAUGUGGAUGGUCUCAACCCCAUCCGUG C. trachomatis K ACUUNAUGUGGAUGGUCUCAACCCNAUCNGUG Core Variation GgucucaacccC Probe 879 AACTACACCTACCAGAGTTGGGGTAGGC-5' Pos. # (E. coli) 995 1046 I I Escherichia coli GACAUCCACGGAA-GUUUUCAGAGAUGAGAAUGUGCCUUCGGGAACCGUGAGACAGG Chlamydia psittaci GACAUGUAUUUGACCGCGGCAGAAAUGUCGUUUUC-CGCAAGGACAGAUA-CACAGG C. trachomatis LGV GACAUGUAUAUGACCGCGGCAGAAAUGUCGUUUUC-CGCAAGGACAUAUA-CACAGG Core Variation GUaUWUGaCCGCGGcagaAaugUcGUuUuC-cGCAAggaCAKAuA-C Probe 783 GTACATAAACTGGCGCCGTCTTTACAGCAAAAG-GC-5' Probe 882 CGTCTTTACAGCAAAAG-GCGTTCCTGTATAT-GTGT-5' TABLE 2: CHLAMYDIA 23S rRNA PROBES AND TARGET SEQUENCES Pos. * (E. coli) 80 113 I I Escherichia coli UCGGUAAGGUGAUAUGAACCGUUAUAACCGGCGAUUUC C. trachomatis LGV CCGGCGAGCUGGUGAUJMGCAAAG-ACCCGGAGGUAUC Chlamydia psittaci UCGGCGAGCUGGUAUAAAGCUAUG-ACCCGGAGGUCUC Core Variation GAUaagcAaAg-acccggagguA Probe 1318 CCGCUCGACCACUAUUCGUUUC-UGGGCCUCCAU-5' Pos. fr (E. coll) 135 159 164 188 I I I I Escherichia coli UGUGUUUCGACACACUAUCAUUA---ACUGAAUCCAUAGGUUAAUGAGGCGA-ACCGGGGGA C. trachomatis LGV GGUAGAGUAAUAGACUACCAUUGCAUGCUGAAUACAUAGGUAUGCAAAGCGACACCUGCCGA Chlamydia psittaci GGUAGAUUAAUCAUCUACCAUUAUACGUUGAAUACAUAGG-CGUAUAAGGCACNCCUGUUGA I Core Variation GuaauAGAcuaggauuGCaUgC Probe 1319 AUCUCAUUAUCUGAUGGUAACGUACGACUUAUG-5' Core Variation UAUGCAaagCGacaccugCC Probe 1320 CUUAUGUAUCCAUACGUUUCGCUGUGGACGG-5' Pas. # (E. coli) 258 269 269 297 Escherichia coli G IGGGAGCAGCCC I I AGAGCCUGAAUCAGUGUGUGUGUUAGUGIGA C. trachomatis LGV GGGGAAUAGCCUAAACCGAGCUGAUAAGGCUCGGGGUUGUAGGAUUGAGGAUAAAGGAUCAGGACUCCUAGUUGA Chlamydia psittaci GGGGAAAAGCCUAAACC-ACAUUUUUAAUGU-GGGGUUGUAGGGUCGAUAACAUGGGAUCUUAAGUUUUAGUUGA Core Variation UagccuaaaccGaGCuGAuAaGGCuC Probe 1321 CCCCUUAUCGGAUUUGGCUCGACUAUUCCGAGC-5* Core Variation AuUgaGGaUaAAggaucAGGaCuCC Probe 1322 UCCUAACUCCUAUUUCCUAGUCCUGAGGAUCAA-5' TABLE 2 (cont'd): CHLAMYDIA 23S rRNA PROBES AND TARGET SEQUENCES Pos. # (E. coli) 343 371 I I Escherichia coli CACAAAAAUGCACAUGCUGUGAGCUCGAU GAGU C. trachomatis LGV GACGAAAGGAGAGAAAGACCGACCUCAACACCUGAGU Chlamydia psittaci GACGAAAAAAC-AAGAGACUCUAUUCGAUACCUGAGU Core Variation GGAGAGaAagacCGACCucAaC Probe 1323 GCUUUCCUCUCUUUCUGGCUGGAGUUGUGGACU-5' Pos. # (E. coli) 1158 1193 I I Escherichia coli AGCUGCGGCAGCGACGCUUAUGCGUUGUUGGGUAGGGGAG C. trachomatis LGV CAUCGCGGGUGUGUCGAUAAGA CACGCGGUAGGAGAG Chlamydia psittaci AAUCGCGGGUGUAUAUUUAUAU AUCGCGGUAGGAGAG Core Variation GuCGAuaAGA CA Probe 1479 AGCGCCCACACAGCTATTCT GTGCGCCATCCTC-5' Pos. # (E. coli) 1457 1489 I I Escherichia coli GGUUGUCCCGGUUUAAGCGUGUAGGCUGGUUUUCC—AGG C. trachomatis LGV CGAUUGGAAGAGUCCGUAGAGCGAUGAGAACGGUUAGUAGG Chlamydia psittaci CGAUUGGAAAUGUCCGUAUCACAAUGAGACCGGUUAGUAGG Core Variation GAguccguaGAGcGaugagaA Probe ll325 UAACCUUCUCAGGCAUCUCGCUACUCUUGCCAA-5' Pos. # (E. coli) 1503 1531 I I Escherichia coli CAAAUCCGGAAAAUCAAGGCUGAGGCGUGAUGACGAGG CAC C. trachomatis LGV CAAAUCCGCUAACAUAAGAUCAGGUCGCGAUCAAGGGGAAUCUUC Chlamydia psittaci CAAAUCCGCUAACAUAAGGUUAGGUUGUGGUUAAGGGAAAUCUUC Core Variation AuCagguCgCgAuCaagggG Probe 1324 UUGUAUUCUAGUCCAGCGCUAGUUCCCCUUAGA-5' Pos. # (E. coli) 1708 1746 I I Escherichia coli CGCUGAUAUGUAGGUGAGGUCCCUCGCGGAUGGAGCUGAAAUC C. trachomatis LGV AGCCUU—UUAGGGUGACUAUGGA-ACGAUAGGAGCCCCGGGG Chlamydia psittaci AGCCUC—UUAGGGUGAUUgcCUUUACGGCAUGAGCUCCGGGG Core Variation U—uuagggugaCuAUGGA-acgAUaGgagcC Probe 1220 GGAA—AAϋCCCACUGAUACCU-UGCUAUCCUCGGGGCC-5* TABLE 3a: LIQUID HYBRIDIZATION INCLUSIVITY DATA FOR 16S rRNA-TARGETED PROBES PROBE HYBRIDIZATION Genus species Strain 1153 1203 781 860 861 879 Chlamydia trachomatis LGV-1 ++++ ++++ +-H-+ +++ ++++ Chlamydia trachomatis LGV-2 ++++ ++++ ++++ +++ ++++ Chlamydia trachomatis LGV-3 ++++ ++++ ++++ +-H-+ ++++ Chlamydia trachomatis A ++++ Chlamydia trachomatis B ++++ + ++++ +++ I t-o Chlamydia trachomatis Ba + ++++ + +++ ++++ •fc- l Chlamydia trachomatis C + ++++ +++ Chlamydia trachomatis D + ++++ + ++++ ++++ Chlamydia trachomatis E ++++ ++++ + ++++ ++++ Chlamydia trachomatis F ++ ++++ + ++ Chlamydia trachomatis G + ++++ + ++++ ++++ Chlamydia trachomatis H + ++++ + ++ ++ Chlamydia trachomatis I + ++++ + +++ ++++ Chlamydia trachomatis J + ++++ + +++ ++++ Chlamydia trachomatis K ++++ ++++ ++++ ++++ ++++ ++++ Bacillus cereus GT0008 Camphylobacter jejunii GT0022 Candida albicans GT0128 Coryπebacter genitalium GT0045 Escherichia coli GT1720 Neisseria gonorrea GT0315 "+++++" « positive control level of hybribiza ion, "+" = barely detectable, "-" = zero. TABLE 3B: LIQUID HYBRIDIZATION INCLUSIVITY DATA FOR 23S rRNA-TARGETED PROBES PROBE HYBRIDIZATION Genus species strain 1318 1319 1320 1321 1322 1323 1324 1325 1220 Chlamydia trachomatis LGV-1 ++++ ++++ ++++ ++++ ++++ +++ ++++ ++++ ++++ Chlamydia trachomatis LGV-2 ++++ +++ ++++ ++++ +++ +++ ++ +++ ++++ Chlamydia trachomatis LGV-3 ++++ ++++ ++++ ++++ +++ ++++ +++ +++ ++++ Chlamydia trachomatis A ++++ ++++ ++++ ++++ ++ ++++ ++ +++ ++ Chlamydia trachomatis B ++++ ++++ ++++ ++++ +++ ++++ ++ +++ ++ I to Chlamydia trachomatis Ba ++++ ++++ ++++ ++++ +++ ++++ ++++ ++++ ++++ \J\ I Chlamydia trachomatis C ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ Chlamydia trachomatis D ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ Chlamydia trachomatis E ++++ ++++ ++++ ++++ ++++ ++ ++++ ++++ ++++ Chlamydia trachomatis F ++++ +++ ++++ ++++ ++++ + +++ +++ ++ Chlamydia trachomatis G ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ Chlamydia trachomatis H ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++ Chlamydia trachomatis I ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ Chlamydia trachomatis J ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ Chlamydia trachomatis K ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ Bacillus cere ε GT0008 - - - - - - - - - Camphylobacter jejunli GT0022 - - - - - - - - - Candida albicans GT0128 - - - - - - - - - Corynebacter genitaliu GT0045 - - ■- Escherichia coli GT1720 - - - - - - - - - Neisseria gonorrea GT0515 - - - - - - - - - "++++" = positive level of hybridization, "+" = barely detectable, "-" = zero. TABLE 4A: CHLAMYDIA 16S EXCLUSIVITY DOT-BLOT DATA PROBE HYBRIDIZATION Genus species strain 1153 1203 781 860 861 879 783 882 Chlamydia trachomatis (EB) LGV + +++ ++++ ++++ ++++ ++++ ++++ +++ Chlamydia trachomatis (RB) LGV - +++ ++++ ++++ ++++ ++++ +++ ++++ Chlamydia psittaci (EB +RB) LGV - - - - - - +++ +++ Acinetobaterr calcoaceticus GJC0002 - - - - - - - - Acinetobacter lwoffii GT0004 - - - - - - - - Aeromonas sobria GT0007 - - - - - - - - Bacillus cereus GT000B - - - - - - - - Bacteroides fragilis ATCC29771 - - - - - • - Bacteroides mellanioogenicus GT0011 - - - - - - - - I Bacteroides thetaiotamicron GT0527 - - - - - - - - t Bifidobacterium dentium GT0012 - - - - - - - - I Camphylobacter jejunii GT0022 - - - - - - - - Candida albicans 223-87 - - - - - - - - Candida albicans 819-88 - - - - - - - - Candida tropicalis 750 - - - - . - Candida tropicalis 224-87 - - - - - - - - Candida diversus GT0030 - - - - - - - - Citrobacter freundii GT0687 - - - - - - - - Cloεtridium perfringens ATCC13124 - - - - - - - - Clostridium gentalium GT0045 - - - - - - - - Enterobacter agglomeranε GT06B3 - - - - - - - - Enterobacter cloacae GT0686 - - - - - - - - Enterobacter sakazakii GT0062 - - - - - - - - Escherichia coli 1665 - - - - - - - - Escherichia hermanii GT0232 - - - - - - - - Escherichia vulneris GT0233 - - - - - - - - Fusobacterium necrophorum GT0238 - - - - - - - - TABLE 4A (cont'd): CHLAMYDIA 16S EXCLUSIVITY DOT-BLOT DATA PROBE HYBRIDIZATION Genus species strain 1153 1203 781 860 861 879 783 882 Fusobacterium prauεnitzii ATCC27768 - - - - - - - - Flavobacterium meningosepticum GT0037 - - - - - - - - Giardia * CDC ( human ) - - + _ + _ _ _ Hafnia alvei GT0241 - - - - - - - - Haemophilus influenzae GT0244 - - - - - - - - Kingella dentrifleans GT0245 - - - - - - - - Kingella indologenes GT0246 - . - Kingella kingae GT0247 - - - - - ■ - I klebsiella pneumonia 1500 - - - - - - - - (S3 • ^1 Lactobacillus acidophilus GT0256 - - - - - - - - I Kingella kingae GT0247 - - - - - - - - Lactobacillus minitus GT0257 - - - - - - - - Lactobacillus casei GT0805 - - - - - - - - Lactobacillus plantaru GT0258 - - - - - Listeria monocytogenes IG3299 - - - - - Morganella morganii GT0303 - - - - - - - - Moraxella osloensis GT0301 - - - + + Neisεeria cinerea GT0307 - - - - - - - - Neisseria flaveεcens GT0310 - - - - - - - - Neisserla gonorrhoea ATCC19424 - - - - - - - - Neisseria meningitidis GT0349 - - - - - - - - Neiεεeria mucoεa . GT0353 - - - - - - - - Peptostreptococcus anaerobius GT0359 - - - - - - - - Plesiomoπas shigelliodes ATCC14029 - - - - - - - - Proteus mirabilis 1496 - - - - - - - - Proteus vulgariε GT0368 - - - - - - - - Providencia alcalificiens GT0371 - - - - - - - - TABLE 4A (cont'd): CHLAMYDIA 16S EXCLUSIVITY DOT-BLOT DATA PROBE HYBRIDIZATION Genus species strain 1153 1203 781 860 861 879 783 BB2 Providencla rettgeri GT0373 - - - - - - - - Providencia stuartii GT0375 - - - - - - - - Pseudo onas acidovorans GT0376 - - - - - - - - Pseudomonas aeruginoεa 1908 - - - - - - - - Salmonella arizona GT0799 - - - - - - - - Salmonella typhimuriu GT0389 - - - - - - - - Serratia arcescens GT0392 - - - - - • - Shigella boydii C-13 RF 974 - I Shigella dysenteriae RF 970 - - - - - - - - 00 I Shigella flexnerii GT0798 - - - - - - - - Shigella sαnnei RF 968 - - - - - - - - Staphylococcus aureus GT0399 - - - - - - - - Staphylococcus epidermitis GT0401 - - - - - - - - Streptococcus sanguis GT0411 - - - - - - - - Streptococcus agalactiae GT0405 - - - - - - - - Streptococcus faecalis GT0406 - - - - - - - - Streptococcuε faecium GT0407 - - - - - - - - Streptococcus mutans GT0412 - - - - - - - - Streptococcus salivarius GT0410 - - - - - - - - Torulopεis globrata 2001 - - - - - - - - Vibrio parahemolyticus GT0568 - - - - - - - - Xanthomonas maltophilia GT0417 - - - - - - - - Yerslnia enterocolitica GT0419 - - - - - - - - L-cell RNA - _ _ + - - _ - Human RNA f om Whole Blood - - - - - - - - Human stool - - - - - - - - * Inclusivity and Exclusivity data was collected front overnight exposures. AA Each organism is represented by lOOng of CsTFA purified RNA. TABLE 4B (cont'd): CHLAMYDIA 23S EXCLUSIVITY DOT-BLOT DATA PROBE HYBRIDIZATION Genus species strain 1318 1319 1320 1321 1322 1323 1324 1325 1479 1220 Pseudomonas aeruginosa 1908 - - - - - - - - - - Salmonella arizona GT0799 - - - - - - ++ - - - Salmonella typhimurium GT0389 - - - - - - + - - - Serratia marcescens GT0392 - - - - - - - - - - Shigella boydii C-13 RF 974 - - - _ - - + - - Shigella dysenteriae RF 970 - - - - - - ++ - - - Shigella flexnerii GT0798 - - - - - - ++ - - - Shigella sonnei RF 968 - - - - - .- + - - - Staphylococcus aureus GT0399 - - - - - - - - - - I Staphylococcus epidermitis GT0401 - - - - - - - - - - * O I Streptococcus sanguis GT0411 - - - - - - - - - - Streptococcus agalactiae GT0405 - - - - - - - - - - Streptococcus faecalis GT0406 - - - - - - - - - - Streptococcus faecium GT0407 - - - - - - - - - - Streptococcus mutans GT0412 - - - - - - - - - - Streptococcus salivarius GT0410 ~ ~ . ~ ~ - - - - - - Torulopεis globrata 2001 - - ' - - - - - - - Vibrio parahe olyticuε GT056B - - - - - - - - - - Xanthomonas maltophilia GT0417 - - - - - - - - - - Yersinia eπterocolitica GT0419 - - - - - - - - - - McCoy cell RNA - - - - - - - - - ND Human RNA f om Whole Blood - - _ _ _ _ _ _ _ _ A Inclusivity and Exclusivity data was collected from overnight exposures. AA Each organism is represented by lOOng of CsTFA purified RNA. TABLE 4B (cont'd): CHLAMYDIA 23S EXCLUSIVITY DOT-BLOT DATA PROBE HYBRIDIZATION Genus ppecies strain 1318 1319 1320 1321 1322 1323 1324 1325 1479 1220 Giardla : CDC ( human ) Hafnla alvei GT0241. - - - - - - ++ - - - Hae ophilus iπfluenzae GT0244 - - - - - - - - - - Kingella dentrifleans GT0245 - - - - - - - - - - Kingella indologenes GT0246 - - - - - - - - - - Kingella kingae GT0247 - - - - - - + - - - klebsiella pneumonia 1500 - - - - - - - - - - Lactobacillus acidophilus GT0256 - - - - - -- - - - - Lactobacillus minitus GT0257 - - - - - - - - - - I Lactobacillus casei GT0B05 - - - - - - - - - - O Lactobacillus plantarum GT0258 - - - - - - - - - - I Moraxella osloensis GT0301 - - - - - - - + - - Neisseria cinerea GT0307 - - - - - - - - - - Neisseria flavescens GT0310 - - - - - - - - - - Neisseria gonorrhoea ATCC19424 - - - - - - - - - - Neisseria meningitidis GT0349 - - - - - - - - - - Neisseria mucoεa GT0353 - - - - - - - - - - Peptostreptococcus anaerobius GT0359 - - - - - - - - - - Plesiomonas shigelliodes ATCC14029 - - - - - - - - - - Proteus mirabilis 1496 - - - - - - - - - - Proteus vulgaris GT0368 - - - - - - - - - - Providencia alcaliflciens GT0371 - - - - - - - - - - Providencla rettgeri GT0373 - - - - - - - - - - Providencia stuartil GT0375 - - - - - - - - - - Pseudomonas acidovorans GT0376 - - - - - - - - - - TABLE 4B: CHLAMYDIA 23S EXCLUSIVITY DOT-BLOT DATA PROBE HYBRIDIZATION Genus species strain 1318 1319 1320 1321 1322 1323 1324 1325 1479 1220 Chlamydia trachomatis (LGV) ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ ++++ Acinetobaterr calcoaceticus GT0002 - - - - - - - - - - Acinetobacter lwoffii GT0004 - - - - - - - - - - Aeromonas sobria GT0007 - - - - - - - - - - Bacillus cereuβ GT0008 - - - - - - - - - - Bacteroides fragilis ATCC29771 - - - - - - - - - - Bacteroides mellanioogenicus GT0011 - - - - - - - - - - Bacteroides thetaiotamicron GT0527 - - - - - .- Bifidobacterium dentium GT0012 - - - - - - - - - - c Io Candida albicans ATCC60193 - - - - - - - - - - * Candida albicans ATCC36232 - - - - - - - - - - Candida tropicalis 750 - - - - - - - - - - Candida diversus GT0030 - - - - - - ++ - - - Citrobacter freundii GT0687 - - - - - - ++ - - - Cloεtridium perfringenε ATCC13124 - - - - - - - - - - Clostridium gentalium GT0045 - - - - - - - - - - Enterobacter agglomeranε GT0683 - - - - - - ++ - - - Enterobacter cloacae GT0686 - - - - - - + - - Enterobacter εakazakii GT0062 - - - - - - + - - - Escherichia coli 1665 - - - - - - + - - - Escherichia hermanii GT0232 - - - - - - +++ - Escherichia vulneris GT0233 - - - - - - +++ - Fuεobacterium necrophorum GT0238 - - - - - - - - - - Fusobacterium prausnitzii ATCC27768 - - - - - - - - - - Flavobacterium meningoεepticum GT0037 - - - - - - - - - -

Claims: Vhat is claimed is: 1. A nucleic acid fragment capable of hybridizing, under predetermined stringency conditions, to rRNA or rDNA of Chlamydia trachomatis and not to rRNA or rDNA of non-Chlamydia bacteria. 2. The nucleic acid fragment of claim 1 vherein said fragment is not capable of hybridizing, under said conditions to rRNA or rDNA of the non-Chlamydia organisms listed in Tables 3 and 4. 3. The nucleic acid fragment of claim 2, comprising a probe sequence selected from the group consisting of probes 781, 782, 783, 860, 861, 879, 882, 1153, 1203, 1220, 1318, 1319, 1320, 1321, 1322, 1323, 1324, 1325, 1479 and their complementary sequences. •4. The nucleic acid fragment of claim 1, vherein said fragment is complementary to at least 90% of a sequence comprising any ten consecutive nucleotides of probes 781, 782, 783, 860, 861, 879, 882, 1153, 1203, 1220, 1318, 1319, 1320, 1321, 1322, 1323, 1324, 1325, 1479. 5. The nucleic acid fragment of claim 1, vherein said fragment is homologous to at least 90S. of a sequence comprising any ten consecutive nucleotides of probes 781, 782, 783, 860, 861, 879, 882, 1153, 1203, 1220, 1318, 1319, 1320, 1321, 1322, 1323, 1324, 1325, 1479. 6. A set of probes comprising at least tvo nucleic acids fragments, at least one of which is selected from the group consisting of probes 781, 782, 783, 860, 861, 879, 882, 1153, 1203, 1220, 1318, 1319, 1320, 1321, 1322, 1323, 1324, 1325, 1479 and their complementary sequences. 7. A method of detecting the presence of Chlamydia trachomatis in a sample comprising: a) contacting said sample vith at least one nucleic acid fragment of claim 1, under conditions that allow said fragment to hybridize to rRNA or rDNA of Chlamydia trachomatis, if present in said sample, to form hybrid nucleic acid complexes, vherein said nucleic acid fragment is capable of hybridizing, under predetermined stringency conditions, to rRNA or rDNA of Chlamydia trachomatis and not to rRNA or rDNA of non-Chlamydia trachomatis; and b) detecting said hybrid nucleic acid complexes as an indication of the presence of said Chlamydia trachomatis in said sample. 8. The method of claim 7 vherein said nucleic acid fragment comprises a probe sequence selected from the group consisting of probes 781, 782, 783, 860, 861, 879, 882, 1153, 1203, 1220, 1318, 1319, 1320, 1321, 1322, 1323, 1324, 1325, 1479 and their complementary sequences. J

Inventors: BUHARIN AMELIA, KING WALTER, LANE DAVID J, MAHAN DONALD, SHAH JYOTSNA, WILLIAMS CHARLOTTE, BUHARIN, AMELIA, KING, WALTER, LANE, DAVID, J., MAHAN, DONALD, SHAH, JYOTSNA, WILLIAMS, CHARLOTTE
Applicants: GENE TRAK SYSTEMS, GENE-TRAK SYSTEMS
Application year: 1990
Main classification: C12Q1
IPC codes: C12N15, C12Q1