Elevated virulence in TCP-PAI carrying strains indicates the pathogenicity island's important role in Vibrio mediterranei pathogenicity to bivalves.

Several Vibrio spp. are important shellfish pathogens. Vibrio mediterranei, notorious for its pathogenicity to bivalve larvae, contains a pathogenicity island, TCP-PAI, homologous to the Vibrio cholerae TCP gene cluster. This study explored the role of TCP-PAI in V. mediterranei pathogenicity by comparing TCP-PAI (+) and TCP-PAI (-) strains isolated from larval and juvenile Chinese razor clams (Sinonovacula constricta) and blood clams (Tegillarca granosa). Three independent PCR assays were developed to detect TCP-PAI, tcpA, and tcpB, respectively. The assay amplified a 949 bp band for TCP-PAI (-) strains, a 13375 bp band for TCP-PAI (+) strains, a 415 bp band for tcpA, and a 400 bp band for tcpB, with detection limits of 1.98 × 103 copies/µL for TCP-PAI and 1.38 × 103 copies/µL for tcpA and tcpB. A total of 352 Vibrio spp. isolates were collected from larval clams, including 300 from healthy razor clams , 6 from healthy blood clams, and 46 from diseased blood clams. Three isolates (1% of total) from healthy razor clams were identified as V. mediterranei and all were determined to be TCP-PAI (-). Ten isolates (21.7% of total) from diseased blood clams were identified as V. mediterranei and all were TCP-PAI (+). No V. mediterranei was detected in healthy blood clams. Infection experiments showed TCP-PAI (+) strains were associated with up to 99% and 96.67% mortality in razor clams and blood clams, respectively, significantly higher than TCP-PAI (-) strains (71% and 48.89%). These findings highlight TCP-PAI as a key virulence factor in V. mediterranei pathogenicity. The developed PCR assays provide a tool for further studies on TCP-PAI's role and transmission, offering insights into V. mediterranei pathogenesis in bivalves.