Occurrence of Vibrio parahaemolyticus, Vibrio cholerae and Vibrio vulnificus in the clam Ruditapes philippinarum (Adams & Reeve, 1850) from Emilia Romagna and Sardinia, Italy

Abstract

Marine vibrios, <em>Vibrio parahaemolyticus</em>, <em>V. vulnificus</em> and <em>V. cholerae</em> are responsible of the majority of food-borne human infections by consumption of bivalve shellfish. The aim of the present study was to ascertain the occurrence of these bacteria, and their potential pathogenicity, in the Manila clam <em>R. philippinarum</em> from Emilia Romagna (ER) and Sardinia (SR) regions, Italy. Isolation was performed on CHROMagar<sup>TM</sup> vibrio with subculture on (thiosulfate-citrate-bile salts-sucrose) Agar and m-modified-cellobiose-polymyxin bcolistin (-CPC) Agar. Suspected strains were purified, biochemically characterized and genotyped by simplex polymerase chain reaction (PCR) for the specie-specific and pathogenic gene markers: <em>V. parahaemolyticus</em> (<em>tox</em>RP, <em>tdh</em> and <em>trh</em>); <em>V. vulnificus</em> (<em>vvhA</em>, <em>hsp</em>, <em>vcg</em>C, <em>vcg</em>E, CPS operon allele 1, CPS operon allele 2, 16s-rRNA operon allele A, 16s-rRNA operon allele B; <em>V. cholerae</em> (<em>toxR</em>C, <em>hlya</em>, <em>tcpI</em>, <em>tcpA</em>, <em>ctxA</em>, <em>ctxB</em>, <em>stn/sto</em>). Moreover a multiplex PCR was applied to the SR bivalve shellfish, for the simultaneous detection of the three targets directly on homogenate samples, targeting the species-specific gene for <em>V. cholerae</em> (<em>toxR</em>C), <em>V. parahaemolyticus</em> (<em>toxR</em>P) and <em>V. vulnificus</em> (<em>vvhA</em>). As a result of phenotyping and genotyping of isolates, bivalve shellfish from ER resulted positive for <em>V. parahaemolyticus</em> (27.8%) and <em>V. vulnificus</em> (10.1%), but negative for <em>V. cholerae</em>. Shellfish from SR resulted positive for <em>V. parahaemolyticus</em> (30.3%), <em>V. vulnificus</em> (6.1%) and <em>V. cholerae</em> (3%). No significant differences emerged between the two areas (P>0.05).