Characterization and CRISPR-based genotyping of clinical trh-positive Vibrio parahaemolyticus

Vibrio parahaemolyticus is a causative agent of gastroenteritis. Most of the clinical isolates carry either tdh and/or trh genes which are considered as the major virulence genes of this pathogen. In this study, the clinical isolates of V. parahaemolyticus carrying trh gene (n = 73) obtained from 1886 to 2012 from various countries were investigated for the urease production, haemolytic activity, and biofilm formation. In addition, the potential of clustered regularly interspaced short palindromic repeats (CRISPR)-based genotyping among these isolates was investigated.

In this study, no significant differences were observed in the urease production between tdh ⁺ trh1⁺ and tdh ⁺ trh2⁺ isolates (p = 0.063) and between the tdh ^- trh1⁺ and tdh ^- trh2⁺ isolates (p = 0.788). The isolates carrying only the trh gene showed variation in their haemolytic activity. The ratio of urease production and haemolytic activity between the trh1⁺ and trh2⁺ isolates and biofilm formation of trh ⁺ V. parahaemolyticus isolates were not significantly different. Sixteen of thirty-four tested isolates (47.0%) of trh ⁺ V. parahaemolyticus were positive for CRISPR detection. The discriminatory power index (DI) of CRISPR-virulence typing was higher than the DI obtained by CRISPR typing alone.

The tdh and trh genes were not involved in urease production in the trh ⁺ V. parahaemolyticus, and variation of haemolytic activity detected in V. parahaemolyticus carrying only the trh gene might be correlated to the sequence variation within trh1 and trh2 genes. Additionally, biofilm production of V. parahaemolyticus was not associated with harboring of virulence genes. For genotyping, CRISPR sequences combined with virulence genes can be used as genetic markers to differentiate trh ⁺ V. parahaemolyticus strains.