MOLECULAR IDENTIFICATION Development of a PCR assay specific for

Peptostreptococcus anaerobius is a gram-positive anaerobic coccus that is widely distributed in the normal human flora. The organism has also been implicated as a causative agent of several systemic infections, including endocarditis and infections of the genitourinary and gastrointestinal tracts. Its role in oral disease is less well defined, although it has been implicated in periodontal disease, gingivitis and root canal infections. Identification of P. anaerobius in clinical samples is currently reliant upon traditional culture and biochemical methods. The aim of this study was to develop a novel PCR assay for the detection of P. anaerobius and to attempt detection of this organism in oral samples. PCR primers specific for P. anaerobius DNA were developed by alignment of bacterial 16S ribosomal RNA gene sequences and selection of sequences specific at their 39 ends for P. anaerobius. When used in a PCR assay, positivity for P. anaerobius DNA was indicated by the amplification of a 943-bp product. The primers were shown to be specific for P. anaerobius DNA, as no PCR products were obtained when genomic DNA from a wide range of other Peptostreptococcus species and other oral bacteria were used as templates. The PCR assay was then applied to the detection of P. anaerobius DNA in subgingival plaque samples from adult periodontitis patients and pus aspirates from subjects with acute dento-alveolar abscesses. All of 60 subgingival plaque samples from 16 patients were negative for P. anaerobius DNA. None of the 43 pus samples analysed contained P. anaerobius DNA. These results suggest that P. anaerobius is not a major pathogen in adult periodontitis and dento-alveolar abscesses. The PCR assay is a more rapid, sensitive and specific alternative to culture-based methods for identification of P. anaerobius in clinical samples