Histological and microbiological investigations in farmed mussels (Mytilus galloprovincialis) of La Spezia Gulf, Italy to evaluate the health status

Introduction: Mass mortality episodes in cultured mussels (Mytilus galloprovincialis) occurred in early February 2015 in the Gulf of La Spezia (Ligurian Sea, Italy). The mortality was suspected to be the consequence to dredging and excavation works of the fund to increase the depth of the seabed in La Spezia harbor, an important military, commercial and touristic port. Methodology: Histological and microbiological analysis were carried out in M. galloprovincialis of 4 different sites in the Gulf of La Spezia over a l-year period monthly from October 2015 to September 2016 excluding August. The histopathological examinations were performed on a total of 440 samples processed by standard histological techniques. The populations were “healthy” at the time of sampling. Histological aspects of mussels were evaluated considering 28 health parameters: pathogens, inflammatory and non-specific pathology. Bacteriological analysis were conducted for the isolation of Vibrio splendidus- related and V. aestuarianus strains from pieces of gills and/or mantle according to a method previously published by IFREMER. Colonies morphologically corresponding to Vibrio were selected and tested for DNA presence by a Real time PCR. To identify the pathogenicity of the isolated V. splendidus-related strains, PCR analysis was used to detect the DNA of virulence factors vsm and ompU. Results: Parasites observed were Steinhausia mytilovum 3.9%, Proctoeces maculatus 3.4%, Mytilicola intestinalis 0.9%, ciliated protozoa 2%, Eugymnanthea inquilina and Urastoma cyprinae 31%. Of significance, a previously undescribed haplosporidian was detected in a single mussel sample (0.2%) and was confirmed by in situ hybridization. Biomolecular investigation are currently in progress to confirm the genus of observed a protozoan looking like Perkinsus parasite (0.7%). Different pathological aspects regressive changes, host defense responses and progressive changes were detected in gills, digestive gland, gonads and mantle. It has been observed a single case of haemocytic neoplasia (0.2%). V. splendidus clade was detected in 72.7% (32/ 44) of shellfish samples while all samples were negative for V. aestuarianus. Among the positive samples 46.8% (15/ 32) contained virulence factors: in particular, 14 strains tested positive for vsm and only 1 to ompU, no samples showed positive for both.