First description of Vibrio ostreicida as putative causal agent of disease in Mediterranean mussel (Mytilus galloprovincialis) and development of a PCR protocol for its rapid diagnosis

Abstract

In April 2022, mussels (Mytilus galloprovincialis) cultivated in the Ebro River delta (Spain) were affected by a mortality episode, involving 25 % of individuals, observed at a water temperature of 18 °C. Mussel samples from this site (3 M) were analysed for the presence of potentially pathogenic bacteria, in parallel with those obtained from an area where no mortality was reported (3 N). From all samples, a total of 113 bacterial isolates were obtained. When subjected to ERIC-PCR, the obtained band patterns showed a distinct profile in the 3 M samples, covering 13 % of the isolates. The genotypic and phenotypic characterization of three representative isolates of this profile (r167, r172, r190) allowed their identification as Vibrio ostreicida. In addition, the complete genome of isolate r172 was sequenced to identify functional features of interest. Histological examination revealed bacterial colonies and tissue damage compatible with bacterial toxicity in 3 M samples. From a preliminary assessment of the virulence of the r172 isolate by mussel challenge via injection (9 × 10⁶ cfu/mussel), a mean mortality of 50 % was observed in infected animals at 5 days. Moreover, a specific PCR protocol was designed for V. ostreicida using the gyrB gene as a target, able to detect the pathogen from both pure cultures and from tissues. To our knowledge, this is the first evidence of the association of V. ostreicida with a mortality episode in cultured M. galloprovincialis. The PCR protocol developed will be useful for screening and diagnosis of this vibrio species in different matrices and to clarify its importance as shellfish possible pathogen.