Identification and characterization of Streptococcus agalactiae isolates using 16S rRNA sequencing and cellular fatty acid composition analysis

Abstract

This study was undertaken to apply 16S rRNA sequence and Cellular Fatty Acid (CFA) composition analysis techniques for the identification and characterization of seven individual bacteria isolates obtained from seven infected fish samples. All samples were cultured on brain heart infusion agar. All the seven isolates were Gram positive and were identified as Streptococcus sp. The 16S rRNA sequencing method yielded about 1500 bps for each strain where upon the sequence was compared to nucleotide data in Gene Bank using BLASTN 2.2.1 sequence alignment from NCBI for the nucleotide comparison. The 16S rRNA gene sequence for all the seven samples had no sequence variation between the isolates and gave a 100% similarity to plus, plus strand with Streptococcus agalactiae strain A909 Accession number NC_007432.1 and S. agalactiae strain H36B (Accession number AAJS01000007). Also the 16S rRNA sequence showed a high (92-93%) similarity between S. agalactiae and S. equi, S. suis and S. uberis. All strains appeared to be nearly identical to each other after CFA analysis using Library Generation System (LGS) software (MIDI) and were consistent to that of S. agalactiae ATCC 12386, the CFA analysis not only confirmed the results of 16S rRNA sequence but also indicated a possibility of single source of infection. Despite their accuracy to identify the poorly described, rarely isolated, or phenotypically aberrant strains, 16S rRNA gene sequence analysis and CFA analysis lacks widespread use beyond the large and reference laboratories because of technical and cost considerations.