Identification of Abiotrophia adiacens and Abiotrophia defectiva by 16S rRNA gene PCR and restriction fragment length polymorphism analysis

Abstract

Abiotrophia adiacens and Abiotrophia defectiva, previously referred to as nutritionally variant streptococci, Streptococcus adjacens and Streptococcus defectivus, respectively, are causes of infective endocarditis. We describe a method of identifying these two species and also of distinguishing them from 15 other major etiological pathogens of infective endocarditis by means of 16S rRNA gene PCR amplification followed by restriction fragment length polymorphism analysis (PCR-RFLP). The 16S rRNA genes were successfully amplified with a set of universal primers from all 17 species of bacteria examined, including viridans group streptococci. The RFLP patterns of A. adiacens and A. defectiva obtained by HaeIII or MspI digestion were readily distinguished from each other and from those of other bacteria. When PCR analysis was performed with the supernatant of a suspension of a boiled colony, the 16S rRNA genes of 80 of 82 isolates (97%) of A. adiacens and all isolates (11 of 11) of A. defectiva were amplified. The HaeIII RFLP patterns of the isolates were the same as those of the corresponding type strains, although 28% of A. adiacens isolates revealed intraspecies polymorphism. The detection limit of this method was 0.1 pg of genomic DNA, as assessed by using the digoxigenin-labeling DNA detection system. Thus, the PCR-RFLP analysis that we developed is applicable for the routine detection of Abiotrophia from clinical specimens.