Monitoring and genotyping of Norovirus in edible shellfish in Northern Italian seas (2018-2020)

Background Norovirus is an enteric virus with foodborne transmission. Bivalve shellfish are a main source of infections and outbreaks, which in the last years are showing an increasing trend. Therefore, in Italy a voluntary based monitoring plan to control the safety of bivalve shellfish was set up at provincial level. This study describes the occurrence and distribution of Norovirus in different areas of Northern Italy seas. Methods Between the years 2018-2020 622 samples (n = 173 oysters, n = 139 mussels, n = 300 clams, n = 10 other bivalve shellfish) were tested by OneStep RT Real Time PCR for Norovirus GI and GII and quantified according to the ISO 15216:2017. Positive samples were further analyzed to determine genotype by sequencing of the ORF1/ORF2 junction of the Norovirus genome. Results A total of 111 (18%) samples were positive for Norovirus (mussels 22%; oysters 18%; clams 16%, other molluscs 20%), positive samples were found mainly in the winter season. The mean contamination was 103 ge/µL and ranged 100-105 ge/µL. Thirty-two samples were typeable (GI n = 10; GII n = 22) with GI samples belonging to four genotypes and GII samples belonging to five genotypes. GII.4, particularly Sydney 2012 subtype (found in three oysters and three clams samples), was the most prevalent, followed by GII.3. Often more than one genotype was found in samples collected simultaneously in the same sampling site. The virus was found with relatively high occurrence both in samples collected close to the coast (38%) and in those harvested from farms located 3-6 nautical miles offshore (73%). Conclusions The overall prevalence of Norovirus in edible shellfish, particular those to be eaten raw, is moderately high (18%). No difference in contamination was highlighted among the different species or different sampling sites. One of the most prevalent variants found (i.e. GII.4 Sydney 2012) is a leading cause of infections worldwide. Key messages Given the moderately high proportion of positive samples (18%), oysters (which are often eaten raw), regardless of the origin, represent the main risk of infection. The presence of genotypes frequently involved in human infections strengthens the need for ongoing monitoring, which should be extended to all provinces involved in shellfish farming.

Purification, Toxicity and Functional Characterization of a New Proteinaceous Mussel Biotoxin from Bizerte Lagoon

The marine environment is known to be occupied by microorganisms. The potential toxicity of some of these marine microorganisms, that are capable of producing unknown biotoxins, has always been underestimated. Indeed, these biotoxins may be a threat to human health through the consumption of contaminated seafood and fish. For more than ten years, recurrent but atypical toxicity has been detected in mussels from Bizerte lagoon (North of Tunisia) during routine tests. In this study, we have isolated and characterized a new proteinaceous marine biotoxin, named Mussel Toxic Peptide (MTP). Using HPLC, electrophoresis and LC/MS studies, we showed that MTP has a protein characteristic UV-spectrum, can be visualized by protein specific reagents such as Coomassie, and has a molecular mass of 6.4 kDa. Patch-clamp experiments performed on cultured N18 neuroblastoma cells revealed that MTP (0.9–18 µM) markedly inhibited voltage-gated Na current, but was about 23 times less active in blocking voltage-gated K current at equimolar concentrations. To the best of our knowledge, this is the first time that a proteinaceous marine biotoxin with relatively high molecular mass is isolated and involved in the contamination of mussels harvested from shellfish farming areas.

Microbiological and Histological Analysis for the Evaluation of Farmed Mussels (Mytilus galloprovincialis) Health Status, in Coastal Areas of Italy

Shellfish farming is a relevant economic activity in Italy. The Gulf of La Spezia is one of the major production areas for mussels: the area is characterized by the presence of numerous human activities that could harm the quality of seawater. Additionally, the presence of potentially pathogenic microorganisms may influence the health status of animals, which must be constantly monitored. To have a clear view of the health conditions of the mussels (Mytilus galloprovincialis) farmed in this area, microbiological, parasitological, and histological analyses were performed. The study was conducted from November 2016 to October 2017. Overall, despite the presence of potentially pathogenic microorganisms for mussels, abnormal mortality rates were not reported during the monitoring period and the histological examination revealed no significant lesions. Our study confirms that studying different aspects together is a useful tool for assessing the health conditions of mussels and points out the importance of adverse environmental conditions for the expression of the pathogenicity of microorganisms.

Dinoflagellates in tropical estuarine waters from the Maraú River, Camamu Bay, northeastern Brazil

Dinoflagellates display great diversity in tropical regions and play an important role in the complex microbial food webs of marine and brackish environments. The goal of this study is to identify planktonic dinoflagellates and their distribution in the estuary of the Maraú River, Camamu Bay, state of Bahia, in a region with increasing use of shellfish farming. Samples were carried out monthly from August 2006 to July 2007 at four stations along the estuary. Plankton was sampled with a 20 μm mesh net. We identified 20 dinoflagellate species. The greatest species richness was observed in the genera Protoperidinium (five spp.), Tripos (four spp.), and Prorocentrum (three spp.). Based on literature, six species were classified as potentially harmful: Akashiwo sanguinea, Dinophysis caudata, Gonyaulax spinifera, Prorocentrum micans, Scrippsiella cf. acuminata, and Tripos furca. Protoperidinium venustum was recorded for the first time in coastal waters of Bahia.

Pseudo-nitzschia Blooms in a Coastal Upwelling System: Remote Sensing Detection, Toxicity and Environmental Variables

The NW coast of the Iberian Peninsula is dominated by extensive shellfish farming, which places this region as a world leader in mussel production. Harmful algal blooms in the area frequent lead to lengthy harvesting closures threatening food security. This study developed a framework for the detection of Pseudo-nitzschia blooms in the Galician rias from satellite data (MERIS full-resolution images) and identified key variables that affect their abundance and toxicity. Two events of toxin-containing Pseudo-nitzschia were detected (up to 2.5 μg L−1 pDA) in the area. This study suggests that even moderate densities of Pseudo-nitzschia in this area might indicate high toxin content. Empirical models for particulate domoic acid (pDA) were developed based on MERIS FR data. The resulting remote-sensing model, including MERIS bands centered around 510, 560, and 620 nm explain 73% of the pDA variance (R2 = 0.73, p < 0.001). The results show that higher salinity values and lower Si(OH)4/N ratios favour higher Pseudo-nitzschia spp. abundances. High pDA values seem to be associated with relatively high PO43, low NO3− concentrations, and low Si(OH)4/N. While MERIS FR data and regionally specific algorithms can be useful for detecting Pseudo-nitzschia blooms, nutrient relationships are crucial for predicting the toxicity of these blooms.