| Summary: | 碩士 === 國立成功大學 === 醫學檢驗生物技術學系 === 93 === Many species belonging to Enterococcus, Streptococcus, and nutritionally variant streptococci (NVS) including Abiotrophia and Granulicatella, are human pathogens. The feasibility of sequence analysis of the 16S-23S ribosomal DNA intergenic spacer (ITS) for identification of strains belonging to the four genera was evaluated. In this study, ITS sequences of 62 type strains (55 species) and 12 ITS sequences obtained from the GenBank database were used to construct an ITS database for identicication of strains belonging to Enterococcus, Streptococcus, and NVS. The ITS lengths of different species varied from 189 to 602 bp, with interspecies sequence similarities ranging from 0.31 to 0.996. ITS sequences were highly conserved among strains within the same species, with intraspecies similarity ranged from 0.98 to 1.0, except for strains of Granulicatella elegans. A total of 227 strains, including 89 reference strains and 138 clinical isolates, were identified by ITS sequencing. Cinical isolates producing discrepant species names by ITS sequencingwere reconfirmed by the Rapid ID 32 STREP system (bioMe’rieux Vitek, Marcy l’Etoile, France). A total of 27 strains (11.9%, 27/227), including reference strains and clinical isolates, were found to produce discrepant identification results between by ITS sequencing and. The sequences of 16S rRNA genes were further determined for species confirmation of strains producing discrepant identification. The sensitivity of ITS sequence analysis for species identification was 98.2% (226/230). Four strains were not identified by ITS sequencing because their corresponding ITS sequences were not available in the ITS sequence database. Based on ITS sequence, an oligonucleotide array was designed to identify these bacteria. A total of 391 strains, including 336 target strains (160 reference strains and 176 clinical isolates) and 55 nontarget strains were identified by the array. The sensitivity and specificity of the array were 100% (336/336) and 96.4% (53/55), respectively. In conclusion, both ITS sequence analysis and the array hybridization method could be used as accurate alternatives for species identification of Abiotrophia, Enterococcus, Granulicatella, and Streptococcus.
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