Abstract:
Objective To establish a multilocus sequence typing （MLST） assay for Vibrio alginolyticus for the clonal classification of V. alginolyticus and the study of the geographical distribution and source of V. alginolyticus.
Methods Eight housekeeping genes of V. alginolyticus （atpA, dnaE, ftsZ, gyrB, mdh, purM, pyrH, tnaA） were screened and specific primers were designed. The housekeeping gene sequences of laboratory-isolated strains were sequenced by polymerase chain reaction （PCR） amplification, and compared with genomes in the database by BLASTn. The resolution of the 8 genes was evaluated, and the allele profiles and sequence types （STs） were defined based on sequence differences of the 8 housekeeping genes. ST clustering analysis was performed and minimum spanning tree and concatenated sequences based maximum-likelihood tree were constructed.
Results The evaluation of housekeeping gene polymorphism suggested that the selected housekeeping genes had abundant mutation loci and resolution, with different rates of genetic evolution in the evolutionary course, which could meet the requirements of MLST analysis. A total of 184 STs were defined in the allele profile, i.e. ST001-ST184, and 24 clonal groups were identified, i.e. CC1-CC24. Most strains showed a fragmented distribution, and no related phylogenetic characteristics among the strains were found in terms of isolation region, isolation year and host source.
Conclusion The MLST assay for the detection of V. alginolyticus was established, and the V. alginolyticus isolates analyzed in this study showed a high genetic diversity.
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