Detection of Bacillus piliformis by specific amplification of ribosomal sequences.

In an effort to explore a sensitive species-specific detection system using the polymerase chain reaction (PCR) for B. piliformis, we sequenced 16S ribosomal DNA (rDNA) of the organism (MSK strain) isolated from the mouse and compared it with known rDNA sequences of the RJ strain isolated from the rat. Sequence homology between the MSK strain and the RJ strain was over 97%, but homology between the MSK strain and other bacterial species was less (70-83%). The results indicated that the sequences included B. piliformis species-specific regions. On the basis of the sequences, we designed a PCR primer set which amplifies B. piliformis rDNA specifically. The PCR with the primer set detected not only these two strains but also an HN strain of hamster origin, although it did not detect other organisms. Therefore, this primer set was considered to be specific for B. piliformis species. More than one organism (RJ strain) could be detected by the PCR method. Nine Jcl:Wistar rats were infected perorally with 2x10(4) RJ strain organisms, and three rats each were sacrificed on days 1, 3 and 5 postinoculation (p.i.) to investigate the presence of the organism in the liver, heart, cecum, spleen and mesenteric lymph nodes by PCR and the immunofluorescence test. On days 1 and 3 p.i., B. piliformis was not detected in any tissues of the six rats, but B. piliformis was detected in two of the three rats sacrificed on day 5 p.i. The presence of the pathogen was seen in both liver and heart (1/3), or in the cecum (1/3) by both methods.(ABSTRACT TRUNCATED AT 250 WORDS)