Assessment of Vibrio parahaemolyticus levels in oysters (Crassostrea virginica) and seawater in Delaware Bay in relation to environmental conditions and the prevalence of molecular markers to identify pathogenic Vibrio parahaemolyticus strains

Abstract

This study identifiedVibrio parahaemolyticusin oyster and seawater samples collected from Delaware Bay from June through October of 2016. Environmental parameters including water temperature, salinity, dissolved oxygen, pH, and chlorophyllawere measured per sampling event. Oysters homogenate and seawater samples were 10-fold serially diluted and directly plated on CHROMagarᵀᴹ Vibrio medium. PresumptiveV.parahaemolyticuscolonies were counted and at least 20% of these colonies were selected for molecular chracterization.V.parahaemolyticusisolates (n = 165) were screened for the presence of the species-specific thermolabile hemolysin (tlh) gene, the pathogenic thermostable direct hemolysin (tdh)/ thermostable related hemolysin (trh) genes, the regulatory transmembrane DNA-binding gene (toxR), andV.parahaemolyticusmetalloprotease (vpm) gene using a conventional PCR. The highest mean levels of the presumptiveV.parahaemolyticuswere 9.63×103CFU/g and 1.85×103CFU/mL in the oyster and seawater samples, respectively, during the month of July.V.parahaemolyticuslevels in oyster and seawater samples were significantly positively correlated with water temperature. Of the 165 isolates, 137 (83%), 110 (66.7%), and 108 (65%) weretlh+,vpm+, andtoxR+, respectively. Among theV.parahaemolyticus(tlh+) isolates, 7 (5.1%) and 15 (10.9%) weretdh+andtrh+, respectively, and 24 (17.5%), only oyster isolates, were positive for both genes. Potential pathogenic strains that possessedtdhand/ortrhwere notably higher in oyster (39%) than seawater (15.6%) isolates. The occurrence of totalV.parahaemolyticus(tlh+) was not necessarily proportional to the potential pathogenicV.parahaemolyticus. Co-occurrence of the five genetic markers were observed only among oyster isolates. The co-occurrence of the gene markers showed a relatedness potential oftdhoccurrence withvpm. We believe exploring the role ofV.parahaemolyticusmetalloprotease and whether it is involved in the toxic activity of the thermostable direct hemolysin (TDH) protein can be of significance. The outcomes of this study will provide some foundation for future studies regarding pathogenicVibriodynamics in relation to environmental quality.