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Bacteroides forsythus has been associated with destructive adult periodontitis. Up to now, detailed analysis by classical means was hampered by the fastidious nature of the organism. There is hope that the application of molecular detection methods such as indirect immunofluorescence or in situ hybridization (ISH) will allow for more rapid and accurate identification. Here we describe a B. forsythus-specific probe (BFV530), complementary to 16S rRNA, which correctly identified all B. forsythus isolates as confirmed by biochemical, protein, or fatty acid analysis. To assess whether this probe might be suitable for direct identification of B. forsythus in clinical specimens, a total of 92 subgingival plaque samples were analyzed. Fifty-five specimens were tested in parallel by culture, light microscopy, and filter hybridization. Unfortunately, the overall agreement between results of filter hybridization and conventional methods was 70.9% only. We therefore examined 37 new specimens by ISH and indirect immunofluorescence by using fluorescently labeled probe BFV530 or B. forsythus-specific monoclonal antibody 116BF1.2 (kindly provided by R. Gmür, Zurich, Switzerland), respectively. Agreement between these methods was 100%, indicating that ISH with probe BFV530 might be used to accurately identify B. forsythus directly in subgingival plaque samples.

Bacterial kidney disease in salmonid fish is caused by the slow-growing Gram-positive rod, Renibacterium salmoninarum. The partial sequence of 16S rRNA from R. salmoninarum was determined and compared with published bacterial 16S rRNA sequences. From this sequence information, a 30-bases-long oligonucleotide was designed and used as a specific probe for identification of R. salmoninarum in filter hybridization experiments. Strong specific hybridization signals were observed for all strains of R. salmoninarum tested. Furthermore, no cross-hybridization could be seen against 22 other bacterial species, among them other salmonid fish pathogens. The detection limit for the probe in direct filter hybridization by the dot-blot technique was 2.5 x 10(4) bacteria. It was also possible to detect R. salmoninarum in clinical samples by direct filter hybridization.

The partial sequences of 16S rRNA from Mycoplasma bovis and M. agalactiae were determined by dideoxynucleotide sequencing using reverse transcriptase. Two oligonucleotides complementary to different evolutionary variable regions of 16S rRNA from these two species were synthesized. The oligonucleotides were end-labelled with 32P and used as probes in filter hybridization experiments with different bovine, caprine and ovine mycoplasmas as samples. One of the probes, complementary to a sequence of the V8-region of both M. bovis and M. agalactiae, did not cross-hybridize to any bovine, caprine or ovine mycoplasmas except M. bovigenitalium and M. californicum. This probe is thus not useful for analysis of bovine samples, but can be used for detection of M. agalactiae in samples from goats and sheep, since M. bovigenitalium and M. californicum have never been isolated from these hosts and M. bovis only occasionally. The other probe, complementary to a sequence of the V6-region of M. bovis, gave some cross-hybridization with M. agalactiae but not with bovine mycoplasmas. M. agalactiae has never been isolated from cattle and this probe is therefore useful for rapid screening of bovine samples for M. bovis.