Bivalve Molluscs Research Articles

Exposure to microplastics renders immunity of the thick-shell mussel more vulnerable to diarrhetic shellfish toxin-producing harmful algae

Despite both microplastics (MPs) and harmful algae blooms (HABs) may pose a severe threat to the immunity of marine bivalves, the toxification mechanism underlying is far from being fully understood. In addition, owing to the prevalence and sudden occurrence characteristics of MPs and HABs, respectively, bivalves with MP-exposure experience may face acute challenge of harmful algae under realistic scenarios. However, little is known about the impacts and underlying mechanisms of MP-exposure experience on the susceptibility of immunity to HABs in bivalve mollusks. Taking polystyrene MPs and diarrhetic shellfish toxin-producing Prorocentrum lima as representatives, the impacts of MP-exposure on immunity vulnerability to HABs were investigated in the thick-shell mussel, Mytilus coruscus. Our results revealed evident immunotoxicity of MPs and P. lima to the mussel, as evidenced by significantly impaired total count, phagocytic activity, and cell viability of haemocytes, which may result from the induction of oxidative stress, aggravation of haemocyte apoptosis, and shortage in cellular energy supply. Moreover, marked disruptions of immunity, antioxidant system, apoptosis regulation, and metabolism upon MPs and P. lima exposure were illustrated by gene expression and comparative metabolomic analyses. Furthermore, the mussels that experienced MP-exposure were shown to be more vulnerable to P. lima, indicated by greater degree of deleterious effects on abovementioned parameters detected. In general, our findings emphasize the threat of MPs and HABs to bivalve species, which deserves close attention and more investigation.

Traditional knowledge and socioeconomic aspects of small-scale bivalve fishing on the Amazon coast: A case study of Iphigenia brasiliensis on the Island of Maranhão

The bivalve mollusk Iphigenia brasiliensis, locally known as tarioba, is an important fishing resource exploited along the coast of Maranhão. Despite its socioeconomic relevance to numerous shellfish communities on the Amazon coast, little information is available on this species. This study aimed to evaluate the socioeconomic profiles and traditional local knowledge of I. brasiliensis fishermen working on Maranhão Island. For this purpose, 106 guided interviews were conducted with workers selected using the snowball method in the four municipalities of the region: São Luís, Raposa, Paço do Lumiar, and São José de Ribamar. The results showed that shellfish gatherers are predominantly young men with reasonable education and low income who start practicing the activity early, are influenced by family and friends, and have traditional knowledge verticalized between generations. In the region, three methods of capturing mollusks were identified: diving, using hands and feet, and with the support of a stick, with subsistence being the main demand for the activity. Dozens of mollusks are marketed in natura, with prices varying between municipalities, and processing is conducted only by the final consumer.

Procjena utjecaja matriksa na kvantifikaciju lipofilnih toksina u plodovima mora

Phycotoxin accumulation in seafood can cause human intoxication and significant economic losses in seafood-producing areas. To protect consumer safety, maximum permitted levels in bivalve molluscs have been set, with appropriate analytical methods for their determination and quantification. The reference method for lipophilic phycotoxins, commonly referred to as lipophilic toxins, is liquid chromatography tandem mass spectrometry (LCMS/MS method), where coeluting components from the matrix can affect the efficiency of ionisation, resulting in erroneous quantification. In this study, the matrix effect was evaluated using the slope ratio analysis method, which involves comparing the slopes of the calibration curves in the matrix with those in pure solvent. The extent of the matrix effect was investigated in mussels, oysters, queen scallops and ascidians. The study covered all phycotoxins under Regulation EU 853/2004 for which certified standards are commercially available, including unregulated pectenotoxin 2. The results indicated that all lipophilic toxins were susceptible to this effect when LC-MS/MS was used for their determination. Significant ion suppression was evident for most analytes in all matrices, except for okadaic acid and dinophysistoxin 2 in bivalves, where significant ion enhancement was demonstrated, and dinophysistoxin 1 in oyster and scallop extract where no significant effect on ionisation was observed. Further analysis revealed no significant differences between the slope of mussel matrix-matched calibration and that of other bivalve matrices. Given this minor difference, the mussel matrix-matched calibration curve could be applied to minimise the matrix effect and to quantify phycotoxins in bivalve matrices analysed here, with the exception of the okadaic acid group in ascidians, which requires matrix-matched calibration prepared with the blank extract of these mentioned species. Given the risks phycotoxins pose to human health, ongoing analytical method development is necessary in this field to properly control food safety and ensure consumer health.

Identification and expression analysis of two steamer-like retrotransposons in the Chilean blue mussel (Mytilus chilensis)

BackgroundDisseminated neoplasia (DN) is a proliferative cell disorder of the circulatory system of bivalve mollusks. The disease is transmitted between individuals and can also be induced by external chemical agents such as bromodeoxyuridine. In Mya arenaria, we have cloned and characterized an LTR-retrotransposon named Steamer. Steamer mRNA levels and gene copy number correlates with DN and can be used as a marker of the disease. So far, the only mollusk where a retrotransposon expression relates to DN is Mya arenaria. On the other hand, it has been reported that the Chilean blue mussel Mytilus chilensis can also suffers DN. Our aim was to identify retrotransposons in Mytilus chilensis and to study their expression levels in the context of disseminated neoplasia.ResultsHere we show that 7.1% of individuals collected in August 2018, from two farming areas, presents morphological characteristics described in DN. Using Steamer sequence to interrogate the transcriptome of M. chilensis we found two putative retrotransposons, named Steamer-like elements (MchSLEs). MchSLEs are present in the genome of M. chilensis and MchSLE1 is indeed an LTR-retrotransposon. Neither expression, nor copy number of the reported MchSLEs correlate with DN status but both are expressed at different levels among individual animals. We also report that in cultured M. chilensis haemocytes MchSLEs1 expression can be induced by bromodeoxyuridine.ConclusionsWe conclude that SLEs present in Mytilus chilensis are differentially expressed among individuals and do not correlate with disseminated neoplasia. Treatment of haemocytes with a stressor like bromodeoxyuridine induces expression of MchSLE1 suggesting that in Mytilus chilensis environmental stressors can induce activation of LTR-retrotransposon.

Contaminants of Emerging Concern: Antibiotics Research in Mussels from the Coasts of the Tyrrhenian Sea (Sardinia, Italy).

Contaminants of emerging concern (CECs) are compounds found in several environmental compartments whose ubiquitous presence can cause toxicity for the entire ecosystem. Several personal care products, including antibiotics, have entered this group of compounds, constituting a major global threat. It is essential to develop simple and reliable methods by which to quantify these contaminants in several matrices. In this work, mussels were chosen as sentinel organisms to assess environmental pollution and the safety of bivalve mollusk consumption according to the "One Health perspective". A liquid chromatographic tandem mass spectrometry method (LC-MS/MS) was developed for the quantification of two macrolides, erythromycin (ERY) and azithromycin (AZI), in mussels. This new method was validated according to international guidelines, showing high selectivity, good recoveries (>60% for both of them), sensitivity, and precision. The method was successfully applied for ERY and AZI research in mussels farmed along the Sardinian coasts (Italy), demonstrating itself to be useful for routine analysis by competent authorities. The tested macrolides were not determined in the analyzed sites at concentrations above the limits of detection (LODs). These results demonstrate the food safety of mussels (as concerns the studied antibiotics) and a negligible amount of pollution derived from these drugs in the studied area.

Ponto-Caspian amphipod co-location with zebra mussel beds (Dreissena polymorpha) is influenced by substrate size and population source

AbstractThe global spread of non-native species is leading to an increasing frequency of multiple co-occurring non-native species. We examined the co-occurrence of the bivalve mollusc Dreissena polymorpha (zebra mussel) with three Ponto-Caspian amphipods (Dikerogammarus villosus, Dikerogammarus haemobaphes, and Chelicorophium curvispinum) across England and Wales in association with in-situ substrate size. For all three amphipod species, substrate grain size where amphipods co-occurred with D. polymorpha was significantly finer than when recorded in isolation. Subsequently, we confirmed this via aquarium experiments. We examined the occurrence of D. villosus with D. polymorpha when present with cobbles, gravel, or sand from three population sources (co-location with abundant D. polymorpha populations, co-location with low populations, and naïve). Experiments demonstrated that D. villosus actively sought shelter on or near D. polymorpha, with their co-location being significantly more prevalent in finer grained substrates (sand > gravel > cobble). The strength of this co-location differed by population source, with those co-located with high D. polymorpha densities demonstrating a greater association. Our analyses and experiments indicate that D. polymorpha may facilitate Ponto-Caspian amphipod establishment in otherwise suboptimal locations, whereby mussel shells provide favourable structural habitat for the amphipods, analogous to the presence of coarse-grained benthic sediment.

Analysis of the original notifications in the Tuscany region “Rapid Alert System for Food and Feed” in the seven-year period 2015-2021

The original notifications (n=1355) managed by the Rapid Alert System for Food and Feed of the Tuscany region in the seven-year period 2015-2021 were analyzed. 68.9% of them were classified as alert notifications, and they mostly originated (56.3%) from official controls on the market, followed by the company's own checks (29.1%). Italy was the most represented notifying country (73.3%) and the most represented country of origin of the notified products (64.5%). Out of the 28 considered food categories, “fish and products thereof” (F), “cereals and bakery products” (C), “bivalve mollusks and products thereof” (B), and “meat and meat products (other than poultry)” (M) were the most notified. F were especially notified for the presence of heavy metals (45.7%), C for undeclared allergens (32.1%), B for microbial contaminants (Escherichia coli, 49.6%), and M for pathogenic microorganisms (55.9%), mostly Salmonella spp. and Listeria monocytogenes. This study provides an updated framework for the main food safety issues at the regional level. The Rapid Alert System for Food and Feed should be periodically analyzed to evaluate historical trends and emerging or poorly known hazards. This is especially important for the planning of official control activities and the prevention of risks through the food supply chain.

First Report of Glochidia (Mollusca: Lamellibranchiata) Infestation in Aquarium Fish (Flowerhorn) from Iran

Glochidia are the larva stage of bivalve mollusks (Lamellibranchiata). These temporary obligate parasites are apparent on the fins and gills and rarely on the surface of fish in spring, summer, and autumn. In spring 2018, a flowerhorn (cichlid) with spots appearing body and gills and white small mobile organisms on the floor of an aquarium were referred to the parasitology laboratory in the School of Veterinary Medicine, Science and Research Unit, Tehran Islamic Azad University. The organisms were sampled and clear in Potas 10% fixed by Glycerin gelatin and examined with a light microscope; the samples were diagnosed as Glochidia.

Molecular Identification and Subtype Analysis of Blastocystis sp. Isolates from Wild Mussels (Mytilus edulis) in Northern France.

Blastocystis sp. is the most common single-celled eukaryote colonizing the human gastrointestinal tract worldwide. Because of the proven zoonotic potential of this protozoan, sustained research is therefore focused on identifying various reservoirs of transmission to humans, and in particular animal sources. Numerous groups of animals are considered to be such reservoirs due to their handling or consumption. However, some of them, including mollusks, remain underexplored. Therefore, a molecular epidemiological survey conducted in wild mussels was carried out in Northern France (Hauts-de-France region) to evaluate the frequency and subtypes (STs) distribution of Blastocystis sp. in these bivalve mollusks. For this purpose, 100 mussels (Mytilus edulis) were randomly collected in two sampling sites (Wimereux and Dannes) located in the vicinity of Boulogne-sur-Mer. The gills and gastrointestinal tract of each mussel were screened for the presence of Blastocystis sp. by real-time polymerase chain reaction (qPCR) assay followed by direct sequencing of positive PCR products and subtyping through phylogenetic analysis. In parallel, sequences of potential representative Blastocystis sp. isolates that were previously obtained from temporal surveys of seawater samples at marine stations offshore of Wimereux were integrated in the present analysis. By taking into account the qPCR results from all mussels, the overall prevalence of the parasite was shown to reach 62.0%. In total, more than 55% of the positive samples presented mixed infections. In the remaining mussel samples with a single sequence, various STs including ST3, ST7, ST14, ST23, ST26 and ST44 were reported with varying frequencies. Such distribution of STs coupled with the absence of a predominant ST specific to these bivalves strongly suggested that mussels might not be natural hosts of Blastocystis sp. and might rather be carriers of parasite isolates from both human and animal (bovid and birds) waste. These data from mussels together with the molecular identification of isolates from marine stations were subsequently discussed along with the local geographical context in order to clarify the circulation of this protozoan in this area. The identification of human and animal STs of Blastocystis sp. in mussels emphasized the active circulation of this protozoan in mollusks and suggested a significant environmental contamination of fecal origin. This study has provided new insights into the host/carrier range and transmission of Blastocystis sp. and emphasized its potential as an effective sentinel species for water quality and environmental contamination.

A Comparative Study of Mercury Bioaccumulation in Bivalve Molluscs from a Shallow Estuarine Embayment.

In estuarine food webs, bivalve molluscs transfer nutrients and pollutants to higher trophic levels. Mercury (Hg) pollution is ubiquitous, but it is especially elevated in estuaries historically impacted by industrial activities, such as those in the U.S. Northeast. Monomethylmercury (MeHg), the organic form of Hg, is highly bioaccumulative and transferable in the food web resulting in the highest concentrations in the largest and oldest marine predators. Patterns of Hg concentrations in marine bivalve molluscs, however, are poorly understood. In this study, inorganic Hg (iHg), MeHg, and the total Hg (THg) in soft tissues of the northern quahogs (Mercenaria mercenaria), eastern oysters (Crassostrea virginica), and ribbed mussels (Geukensia demissa) from eastern Long Island sound, a temperate estuary of the western North Atlantic Ocean was investigated. In all three species, concentrations of THg remained similar between the four sampling months (May, June, July, and September), and were mostly independent of animal size. In quahogs, MeHg and iHg displayed significant (p < 0.05) positive (iHg in May and June) and negative (MeHg in July and September) changes with shell height. Variability in concentrations of THg, MeHg, and iHg, both inter- and intra-specifically was high and greater in quahogs and oysters (THg: 37, 39%, MeHg: 28, 39%, respectively) than in mussels (THg: 13%, MeHg: 20%). The percentage of THg that was MeHg (%MeHg) was also highly variable in the three species (range: 10-80%), highlighting the importance of measuring MeHg and not only THg in molluscs.

Neuroendocrine functions of monoamines in invertebrates: Focus on bivalve molluscs

Monoamines (MA) such as serotonin, catecholamines (dopamine, norepinephrine, epinephrine), and trace amines (octopamine, tyramine), are neurotransmitters and neuroendocrine modulators in vertebrates, that contribute to adaptation to the environment. Although MA are conserved in evolution, information is still fragmentary in invertebrates, given the diversity of phyla and species. However, MA are crucial in homeostatic processes in these organisms, where the absence of canonical endocrine glands in many groups implies that the modulation of physiological functions is essentially neuroendocrine.In this review, we summarize available information on MA systems in invertebrates, with focus on bivalve molluscs, that are widespread in different aquatic environments, where they are subjected to a variety of environmental stimuli. Available data are reviewed on the presence of the different MA in bivalve tissues, their metabolism, target cells, signaling pathways, and the physiological functions modulated in larval and adult stages. Research gaps and perspectives are highlighted, in order to enrich the framework of knowledge on MA neuroendocrine functions, and on their role in adaptation to ongoing and future environmental changes.

Determination of hydrophilic marine biotoxins in bivalve molluscs by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS)

This work describes a method for the determination of paralytic shellfish toxins in mussels (M. galloprovincialis), oysters (Crassostrea gigas), and clams (Ruditapes decussatus) samples. The acetic acid extraction method is used for simultaneous analysis of Paralytic Shellfish Poisoning (PSP) and the separation of these compounds was achieved on ethylene bridged hybrid (BEH) Amide column with gradient mobile phase composed of acetonitrile (MeCN) and water (as eluent A and eluent B) both containing 2 mM ammonium formate and 50 mM formic acid. The method specificity, linearity, sensitivity, precision, and accuracy were successfully evaluated towards the detection of 13 hydrophilic marine biotoxins analyzed together (single run) in different sample matrixes such as mussels. oysters. and clams. Calibration curves were found to be linear (R2 ≥ 0.98). average Recovery% of 98.5–99.9%. limits of detection (LODs): 0.16–1.92 µg·kg−1; limits of quantitation (LOQs): 0.54–6.41 µg·kg−1) and the relative standard deviation (RSD%) of the precision was ≤ 2.8%. The method was applied for the determination of PSP in shellfish collected from several Tunisian locations. All LC-MS/MS results of analyzed molluscs collected from different Tunisia locations, reveal no PSP toxin group contamination and that the amount found does not exceeding the threshold of 800 µg STXequ·kg−1.

Monitoring of bivalve mollusk harvesting areas: the relevance of &lt;i&gt;Salmonella&lt;/i&gt; spp.

The microbiological monitoring of bivalve mollusk harvesting areas in the Marche region is based on the parameters of Escherichia coli and Salmonella spp. However, Regulation EU/2019/627 stated criteria based on E. coli only to determine the health status of these areas. Therefore, the reason for Salmonella spp. monitoring, as provided in the Marche region, could be aimed at reducing the risk of placing on the market contaminated bivalve mollusks. This study, using the results of microbiological monitoring carried out in the Marche region from 2015 to 2022 and the methods based on Bayes' theorem and Poisson’s distribution, evaluated the effectiveness and efficiency of Salmonella spp. monitoring in reducing the risk to the consumer. The results show that i) the use of a single sample unit significantly reduced the possibility of detecting non-compliance with the microbiological safety criterion; ii) the time taken to report positive results (average of approximately 10 days) did not allow the timely implementation of control measures; iii) the prevalence of positive outcomes was quite sporadic: a random trend of positivity is recognizable on a geographical and monthly basis for mussels and a geographical basis for striped clams; iv) considering the predictive value of E. coli against Salmonella spp., the specificity is very high and the negative predictive value versus Salmonella spp. would be &gt;80%. In conclusion, the study shows that the monitoring of Salmonella spp. has a limited effect on reducing the risk to the consumer; however, in the cost/benefit assessment, other aspects not covered by this study should be considered.

Leveraging Plasma-Activated Seawater for the Control of Human Norovirus and Bacterial Pathogens in Shellfish Depuration.

Cold plasma is a promising alternative for water treatment owing to pathogen control and a plethora of issues in the agriculture and food sectors. Shellfish pose a serious risk to public health and are linked to large viral and bacterial outbreaks. Hence, current European regulations mandate a depuration step for shellfish on the basis of their geographical growth area. This study investigated the inactivation of relevant viral and bacterial pathogens of three plasma-activated seawaters (PASWs), and their reactive oxygen and nitrogen species (RONS) composition, as being primarily responsible for microbial inactivation. Specifically, F-specific (MS2) and somatic (φ174) bacteriophage, cultivable surrogate (murine norovirus, MNV, and Tulane virus, TV), and human norovirus (HuNoV GII.4) inactivation was determined using plaque counts and infectivity assays, including the novel human intestinal enteroid (HIE) model for HuNoV. Moreover, the kinetic decay of Escherichia coli, Salmonella spp., and Vibrio parahaemolyticus was characterized. The results showed the complete inactivation of phages (6-8 log), surrogates (5-6 log), HuNoV (6 log), and bacterial (6-7 log) pathogens within 24 h while preventing cytotoxicity effects and preserving mussel viability. Nitrites (NO2-) were found to be mostly correlated with microbial decay. This research shows that PASWs are a suitable option to depurate bivalve mollusks and control the biohazard risk linked to their microbiological contamination, either viral or bacterial.

PMAxx-RT-qPCR to Determine Human Norovirus Inactivation Following High-Pressure Processing of Oysters.

Norovirus is the leading cause of viral gastroenteritis globally. While person-to-person transmission is most commonly reported route of infection, human norovirus is frequently associated with foodborne transmission, including through consumption of contaminated bivalve molluscan shellfish. Reverse transcription (RT)-qPCR is most commonly used method for detecting human norovirus detection in foods, but does not inform on its infectivity, posing challenges for assessing intervention strategies aimed at risk elimination. In this study, RT-qPCR was used in conjunction with a derivative of the photoreactive DNA binding dye propidium monoazide (PMAxx™) (PMAxx-RT-qPCR) to evaluate the viral capsid integrity ofnorovirus genogroup I and II (GI and GII) in shellfish following high pressure processing (HPP). Norovirus GI.3 and GII.4 bioaccumulated oysters were subjected to HPP at pressures of 300 and 450MPa at 15°C, and 300, 450 and 600MPa at 20°C. Samples were analysed using both RT-qPCR and PMAxx-RT-qPCR. For each sample, norovirus concentration (genome copies/g digestive tissue) determined by RT-qPCR was divided by the PMAxx-RT-qPCR concentration, giving the relative non-intact (RNI) ratio. The RNI ratio values relate to the amount of non-intact (non-infectious) viruses compared to fully intact (possible infectious) viruses. Our findings revealed an increasing RNI ratio value, indicating decreasing virus integrity, with increasing pressure and decreasing pressure. At 300MPa, for norovirus GI, the median [95% confidence interval, CI] RNI ratio values were 2.6 [1.9, 3.0] at 15°C compared to 1.1 [0.9, 1.8] at 20°C. At 450MPa, the RNI ratio values were 5.5 [2.9, 7.0] at 15°C compared to 1.3 [1.0, 1.6] at 20°C. At 600MPa, the RNI ratio value was 5.1 [2.9, 13.4] at 20°C. For norovirus GII, RT-qPCR and PMAxx-RT-qPCR detections were significantly reduced at 450 and 600MPa at both 15°C and 20°C, with the median [95% CI] RNI ratio value at 300MPa being 1.1 [0.8, 1.6]. Following HPP treatment, the use of PMAxx-RT-qPCR enables the selective detection of intact and potential infectious norovirus, enhancing our understanding of the inactivation profiles and supporting the development of more effective risk assessment strategies.

Reductive stress and cytotoxicity in the swollen river mussel (Unio tumidus) exposed to microplastics and salinomycin

Multistress effects lead to unpredicted consequences in aquatic ecotoxicology and are extremely concerning. The goal of this study was to trace how specific effects of the antibiotic salinomycin (Sal) and microplastics (MP) on the bivalve molluscs are manifested in the combined environmentally relevant exposures. Unio tumidus specimens were treated with Sal (0.6 μg L−1), MP (1 mg L−1, 2 μm size), and both at 18 °C (Mix) and 25 °C (MixT) for 14 days. The redox stress and apoptotic enzyme responses and the balance of Zn/Cu in the digestive gland were analyzed. The shared signs of stress included a decrease in NAD+/NADH and Zn/Cu ratios and lysosomal integrity and an increase in Zn-metallothioneins and cholinesterase levels. MP caused a decrease in the glutathione (GSH) concentration and redox state, total antioxidant capacity, and Zn levels. MP and Mix induced coordinated apoptotic/autophagy activities, increasing caspase-3 and cathepsin D (CtD) total and extralysosomal levels. Sal activated caspase-3 only and increased by five times Cu level in the tissue. Due to the discriminant analysis, the cumulative effect was evident in the combined exposure at 18 °C. However, under heating, the levels of NAD+, NADH, GSH, GSH/GSSG and metallothionein-related thiols were decreased, and coordination of the cytosolic and lysosomal death stimuli was distorted, confirming that heating and pollution could exert unexpected synergistic effects on aquatic life.

Effect of Dietary Supplementation with Lactiplantibacillus plantarum I On Queen Scallop Aequipecten opercularis Under Simulated Climate Change Conditions

Abstract This study examined the effects of dietary supplementation of queen scallop Aequipecten opercularis with an indigenous strain of lactic acid bacteria (LAB), Lactiplantibacillus plantarum I, previously isolated from its digestive tract, on gut microbial populations and growth rates during cultivation under simulated climate change conditions (pH 7.8, T = 16 ± 2 °C). After one month of feeding, the results showed a noticeable reduction in aquaculture diseases causing pathogens while maintaining sufficient viable Lpb. plantarum I cells. A higher pH and temperature resulted in higher growth rates, measured by the weight and length of scallops, compared to the control group. The results obtained shed light on the influence of the addition of lactic acid bacteria on the growth of bivalve mollusks under normal and climate change conditions, and provide control of pathogenic microorganisms. In the context of climate change, host-pathogen interactions need to be recognized and put under control by applying natural solutions to minimize the environmental footprint.

Erratum to: Effect of Hypoxia on Amino Acid Content in Hemolymph and Protein Hydrolysate of the Bivalve Mollusk Anadara kagoshimensis

Erratum to: Effect of Hypoxia on Amino Acid Content in Hemolymph and Protein Hydrolysate of the Bivalve Mollusk Anadara kagoshimensis

Prevalence of bacterial pathogens in bivalve shellfish harvested from the brackish waters of Niger Delta, Nigeria

The presence of bacterial pathogens in bivalve shellfish harvested from the marine ecosystem globally has continued to raise serious food safety concerns, thereby undermining the nutritional and health benefits derived from their consumption. Four species of bivalve shellfish: Bloody cockle (Anadara senilis), Donax clam (Donax rugosus), Knife clam (Tagelus adansonaii) and Mangrove oyster (Crassosstra gasar) harvested from the brackish waters of the Niger Delta were assessed for their bacterial pathogens. Standard methods of analysis were employed to assess microbiological hazards accumulated by shellfish species. The results of the microbiological analysis revealed that the total viable count (TVC) was lower than the recommended limit (5.7 log cfu/g) for fresh bivalve shellfish while Vibrio cholerae, Vibrio parahaemolyticus, Salmonellaspp, Shigellaspp, Listeria monocytogenes and E. coli were far above the stipulated FDA standard for molluscian shellfish. The loads of microbiological hazards in bivalve species indicated that in all the locations, Bloody cockle and Donax clam accumulated more bacterial pathogens while the samples harvested from Ibeno and Iko Town brackish waters were lower in pathogenic loads when compared to shellfish harvested from Andoni and Bonny location. The result obtained in this study is of public health importance since, it will serve as a guide towards improved processing techniques and stimulate actions towards the development of novel surveillance as well as, prevention and control strategies, that will help to reduce foodborne disease outbreaks associated with shellfish.

Detection of pathogenic Vibrio spp. in foods: polymerase chain reaction-based screening strategy to rapidly detect pathogenic Vibrio parahaemolyticus, Vibrio cholerae, and Vibrio vulnificus in bivalve mollusks and preliminary results.

The majority of human diseases attributed to seafood are caused by Vibrio spp., and the most commonly reported species are Vibrio parahaemolyticus, Vibrio vulnificus, and Vibrio cholerae. The conventional methods for the detection of Vibrio species involve the use of selective media, which are inexpensive and simple but time-consuming. The present work aimed to develop a rapid method based on the use of multiplex real-time polymerase chain reaction (PCR) to detect V. parahaemolyticus, V. vulnificus, and V. cholerae in bivalve mollusks. 30 aliquots of bivalve mollusks (Mytilus galloprovincialis) were experimentally inoculated with two levels of V. parahaemolyticus, V. vulnificus, and V. cholerae. ISO 21872-1:2017 was used in parallel for qualitative analysis. The limit of detection of 50% was 7.67 CFU/g for V. cholerae, 0.024 CFU/g for V. vulnificus, and 1.36 CFU/g for V. parahaemolyticus. For V. vulnificus and V. cholerae, the real-time PCR protocol was demonstrated to amplify the pathogens in samples seeded with the lowest and highest levels. The molecular method evaluated showed a concordance rate of 100% with the reference microbiological method. V. parahaemolyticus was never detected in samples contaminated with the lowest level, and it was detected in 14 samples (93.33%) seeded with the highest concentration. In conclusion, the developed multiplex real-time PCR proved to be reliable for V. vulnificus and V. cholerae. Results for V. parahaemolyticus are promising, but further analysis is needed. The proposed method could represent a quick monitoring tool and, if used, would allow the implementation of food safety.