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PubMedDate not supplied

Vancouver workshop: overview and synthesis.

Investigators from the six PICES member nations conducted a May, 1999 interdisciplinary research assessment of Vancouver Harbour, BC, Canada. Sediment contaminants were analyzed at several sites along an assumed pollution gradient extending eastward into the harbour from the relatively less-impacted Howe Sound area (located outside and seaward, to the west). Subsets of a few representative benthic biota (English sole for general contaminants; specific mollusks for TBT) at the sampling sites were concurrently assessed for specific tissue contaminants. Measurements of biotic response to contaminants-across a wide spectrum of biological and ecosystem organization, and ranging from selected biomarkers and tissue responses, to observations at the organism and community level-were likewise conducted. Parameters examined included: P-450 expression, EROD induction and histopathologic assessment of toxicopathic liver lesions in English sole; prevalence and frequency of imposex in neogastropods; perturbations in benthic community responses to pollutants; tissue assays of various bivalve shellfish species for PSP. Results affirmed a consistent east-to-west trend in declining contaminant-associated effects, across a range of biological assessments, thus providing strong evidence that these effects are pollution-related. Specific findings revealed a significant distortion of benthic community structure along the pollution gradient, as well as biomarker evidence of histological and biochemical stress in English sole from central and inner harbour sites. Although the study reaffirms prior assumptions that various species of neogastropods have been extirpated from the harbour, most likely from the antifoulant uses of tributyltin, some of the adverse impacts from TBT may be declining. Also noted were strong biochemical inferences that sediments in the relatively "uncontaminated" Howe Sound, may well contain significant P450-inducing pollutants which were not included as analytes in the design of this limited and preliminary assessment. During the brief sampling period, PSP contamination did not appear to be a significant issue for the harbour.

Animals↗

PubMed2003

Implantable applications of chitin and chitosan.

Chitin, extracted primarily from shellfish sources, is a unique biopolymer based on the N-acetyl-glucosamine monomer. More than 40 years have lapsed since this biopolymer had aroused the interest of the scientific community around the world for its potential biomedical applications. Chitin, together with its variants, especially its deacetylated counterpart chitosan, has been shown to be useful as a wound dressing material, drug delivery vehicle and increasingly a candidate for tissue engineering. The promise for this biomaterial is vast and will continue to increase as the chemistry to extend its capabilities and new biomedical applications are investigated. It is interesting to note that a majority of this work has come from Asia. Japan has been the undisputed leader, but other Asian nations, namely Korea, Singapore, Taiwan and Thailand have also made notable contributions. More recently, China has joined the club to become an increasingly major research source for chitin and chitosan in Asia. This review surveys select works of key groups in Asia developing chitin and chitosan materials for implantable biomedical applications.

Animals↗

PubMed2002

Stress-induced immune changes in the oyster Crassostrea gigas.

Information concerning the effect of stress on invertebrate immune functions are scarce. The present study investigated the consequences of a 15-min mechanical disturbance on immune parameters in oysters Crassostrea gigas. As indicated by noradrenaline and dopamine measurements, the mechanical disturbance caused a transient state of stress in oysters. The number of circulating hemocytes, the migratory and phagocytic activities and reactive oxygen species production of hemocytes were measured before, during and after application of the stressor. Results show that all immune functions were significantly downregulated during stress and a transient period of immunostimulation was observed 30-240 min after the end of the disturbance. Taken together, these results suggest that stress can exert a profound influence on oyster immune functions and they may explain why stress and the outbreak of disease are often linked in shellfish culture. Furthermore, the present study strongly suggests that checking the stress status of animals may be necessary to avoid biases when studying oyster immune responses in vivo.

Animals↗

PubMed2003

First report and toxicological assessment of the cyanobacterium Cylindrospermopsis raciborskii from Portuguese freshwaters.

The freshwater cyanobacterium Cylindrospermopsis raciborskii has become increasingly prevalent in freshwaters worldwide. This species is a concern from a water quality perspective due to its known ability to produce a potent hepatotoxic alkaloid cylindrospermopsin, which has been implicated in outbreaks of human sickness and cattle mortality. C. raciborskii strains isolated from Brazil have also been found to produce the highly toxic paralytic shellfish poisons (PSPs). This article reports the toxicity of four strains of C. raciborskii taken from three reservoirs and one river in Portugal, as well as the occurrence of this species in other water bodies used for potable and recreational purposes. All four strains grown in pure culture in the laboratory were found to be toxic in the mouse bioassay at 8-24h after intraperitoneal administration of single doses ranging from 1337 to 1572 mgkg(-1) Histological examination indicated that liver damage was the primary lesion; in addition, there was inflammation in the intestine. HPLC/MS tests for the presence of cylindrospermopsin, microcystins, and PSP toxins were negative. The available evidence suggests that another toxin may be present. This constitutes the first report of toxic C. raciborskii in Europe and draws attention to the need for increased monitoring of this cyanobacterium in water bodies used for potable and recreational purposes.

Animals↗

PubMed1996

Okadaic acid treatment induces DNA adduct formation in BHK21 C13 fibroblasts and HESV keratinocytes.

Okadaic acid (OA), a toxin involved in diarrhetic shellfish poisoning (DSP), has been shown to be a potent tumor promoter in mouse skin and glandular stomach. However, more recent studies tended to show that OA can also act as a genotoxic. In this study, using the 32P-postlabelling method, DNA adduct formation was obtained in two cell lines (BHK21 C13 fibroblasts and HESV keratinocytes) after treatment by OA for 24 h with a dose range between 0.01 and 5 nM. Nineteen adducts were observed with BHK21 C13 cells and 15 with HESV ones. Low doses did not show adduct formation. Intermediate doses have given the most important number of adducts and with higher doses, the number of adducts decreased dose dependently. Ten adducts were similar in the two strains while 9 were specific of BHK21 C13 cell line and 5 of HESV one. The highest total DNA adduct level from origin parts was estimated at 95.6 adducts/10(9) nucleotides for BHK21 C13 fibroblasts (1 nM OA treatment) and 31.1 adducts/10(9) nucleotides for HESV keratinocytes (0.5 nM OA treatment). In this case, the major adduct (number 3) represented 20% for the fibroblastic cell line and 30% for the keratinocytic strain. The genotoxic effect of OA showed in this study should lead to a more careful survey of DSP outbreaks.

Animals↗

PubMed2002

Improved detection of human enteric viruses in foods by RT-PCR.

Human enteric viruses (including hepatitis A virus (HAV) and Norwalk-like viruses (NLVs)) are now recognized as common causes of foodborne disease. While methods to detect these agents in clinical specimens have improved significantly over the last 10 years, applications to food samples have progressed more slowly. In an effort to improve the sensitivity and speed of virus detection from non-shellfish food commodities by reverse transcription-polymerase chain reaction (RT-PCR), we (i) evaluated multiple RNA extraction methods; (ii) compared alternative NLV primer sets; and (iii) developed a one-step RT-PCR method. Hamburger and lettuce samples, processed for virus concentration using a previously reported filtration-extraction-precipitation procedure, were inoculated with HAV or NV. Several RNA extraction methods (guanidinium isothiocyanate, microspin column, QIAshredder Homogenizer, and TRIzol) and primer pairs were compared for overall RNA yield (microg/ml), purity (A(260)/A(280)), and RT-PCR limits of detection. The use of TRIzol with the QIAshredder Homogenizer (TRIzol/Shred) yielded the best RT-PCR detection limits (<1 RT-PCR amplifiable units/reaction for NV), and the NVp110/NVp36 primer set was the most efficient for detecting NV from seeded food samples. A one-step RT-PCR protocol using the TRIzol/Shred extraction method and the NVp110/NVp36 or HAV3/HAV5 primer sets demonstrated improved sensitivity (>10-fold) over the routinely used two-step method. HAV RNA was detected by RT-PCR at initial inoculum levels corresponding to <10 and <100 PFU per 300 microl sample concentrate (corresponding to 6 g food sample) for hamburger and lettuce, respectively. NV RNA was detected by RT-PCR at initial inoculum levels <5 and <50 RT-PCR amplifiable units per 300 microl concentrate (corresponding to 6 g food sample) for hamburger and lettuce, respectively. Residual RT-PCR inhibitors were effectively removed as evidenced by the ability to detect viral RNA in food concentrates without prior dilution. The methods reported here show promise for rapid, sensitive detection of human enteric viruses in foods.

DNA Primers↗

PubMed2003

Accumulation and transformation of DSP toxins in mussels Mytilus galloprovincialis during a toxic episode caused by Dinophysis acuminata.

The time course of several outbreaks of the diarrhetic shellfish poisoning (DSP) producer Dinophysis acuminata and the consequent kinetic of accumulation and loss of toxins in mussels Mytilus galloprovincialis feeding on them was studied. Samples of mussels and seawater were frequently (2-3 times a week) collected from a raft in the Ri;a de Vigo. DSP toxins content of mussels and water was analyzed by HPLC-FD and phytoplankton was quantified in an inverted light microscope. Only okadaic acid (OA) and some of its conjugated forms (OA CF), estimated by enzymatic hydrolysis, were found in the plankton samples obtained, comprised mainly of D. acuminata cells. The main accumulated form in mussels was OA reaching a maximum of 10.1 microg OA g(-1) in the digestive gland (d.g.) in 16 days, falling below the quarantine level (ca. 2 microg OA g(-1) d.g.) by 45 days. The low polarity conjugated forms (LPCF), estimated by hexane extraction, accounted for 6.2% of the total toxin burden of the mussels. To quantify the rates of the processes involved in the accumulation, transformation and loss of the toxins, two dynamic models, a one-compartment and a two-compartment, including OA and its conjugated forms as variables were designed and implemented. The one-compartment model provided a good fit to the OA and LPCF actual data (r(2)=0.92 and r(2)=0.94, respectively). The two-compartment model did not fit the data markedly better than its one-compartment counterpart (r(2)=0.93 and r(2)=0.95, for OA and LPCF, respectively). High hydrolysis rates were estimated for most of the OA CF, which means that these forms came largely from the ingested plankton. The low estimated acylation rates support the previous point and suggest that the formation of LPCF by direct acylation of the OA is of little importance in M. galloprovincialis. Only in cases where the intoxication period is very long, can the formed acyl-derivatives be important, because they seem to accumulate for a long time in the mussels, as suggested by the low hydrolysis and depuration rates estimated from model fitting.

Animals↗

PubMed2002

Combined use of LC/MS and a biological test for rapid identification of marine mycotoxins produced by Trichoderma koningii.

Trichoderma koningii Oudemans, a strain isolated from a shellfish farming area, was selected for its high frequency in samples and its ability to produce metabolites when cultured in natural seawater. Combined use of LC/MS and a biological test on blowfly larvae allowed the characterization of four compounds after purification in only two steps (VLC and HPLC). ESI/MS, a powerful tool for rapid identification and sequence determination of peptides, confirmed that these compounds were peptide, alpha-aminoisobutyric acid and amino alcohol (peptaibols), the usual metabolites of Trichoderma.

Chromatography, Liquid↗

PubMed2001

Occurrence of Listeria monocytogenes in food in Chile.

Out of 2145 food samples analysed 77 were found contaminated with Listeria monocytogenes in Santiago, Chile. Samples were: 603 ice-cream (3.5% contaminated), 256 soft cheese (0.8%), 155 hard cheese (0%), 229 baby milk bottles (0%), 634 processed meat products (3.6%) and 268 crustaceous shellfish (11.6%). Three different isolation media were used: for 318 samples, Modified McBride Agar (MMA), Lithium chloride Phenylethanol Moxalactam agar, and Polymyxin Acriflavine Lithium chloride Ceftazidime Aesculin Mannitol agar; for 1827 samples MMA was replaced by Listeria Selective Agar Oxford Formulation. Isolates were classified as follow: serovar 1/2a (25 isolates), serovar 4b (20), serovar 1/2b (19), serovar 3b (7), serovar 1/2c (2), untypable (4). A high variety of phagovars was detected although 52% of strains was untypable.

Bacteriophage Typing↗

PubMed2004

Foodborne viruses: an emerging problem.

Several groups of viruses may infect persons after ingestion and then are shed via stool. Of these, the norovirus (NoV) and hepatitis A virus (HAV) are currently recognised as the most important human foodborne pathogens with regard to the number of outbreaks and people affected in the Western world. NoV and HAV are highly infectious and may lead to widespread outbreaks. The clinical manifestation of NoV infection, however, is relatively mild. Asymptomatic infections are common and may contribute to the spread of the infection. Introduction of NoV in a community or population (a seeding event) may be followed by additional spread because of the highly infectious nature of NoV, resulting in a great number of secondary infections (50% of contacts). Hepatitis A is an increasing problem because of the decrease in immunity of populations in countries with high standards of hygiene. Molecular-based methods can detect viruses in shellfish but are not yet available for other foods. The applicability of the methods currently available for monitoring foods for viral contamination is unknown. No consistent correlation has been found between the presence of indicator microorganisms (i.e. bacteriophages, E. coli) and viruses. NoV and HAV are highly infectious and exhibit variable levels of resistance to heat and disinfection agents. However, they are both inactivated at 100 degrees C. No validated model virus or model system is available for studies of inactivation of NoV, although investigations could make use of structurally similar viruses (i.e. canine and feline caliciviruses). In the absence of a model virus or model system, food safety guidelines need to be based on studies that have been performed with the most resistant enteric RNA viruses (i.e. HAV, for which a model system does exist) and also with bacteriophages (for water). Most documented foodborne viral outbreaks can be traced to food that has been manually handled by an infected foodhandler, rather than to industrially processed foods. The viral contamination of food can occur anywhere in the process from farm to fork, but most foodborne viral infections can be traced back to infected persons who handle food that is not heated or otherwise treated afterwards. Therefore, emphasis should be on stringent personal hygiene during preparation. If viruses are present in food preprocessing, residual viral infectivity may be present after some industrial processes. Therefore, it is key that sufficient attention be given to good agriculture practice (GAP) and good manufacturing practice (GMP) to avoid introduction of viruses onto the raw material and into the food-manufacturing environment, and to HACCP to assure adequate management of (control over) viruses present during the manufacturing process. If viruses are present in foods after processing, they remain infectious in most circumstances and in most foods for several days or weeks, especially if kept cooled (at 4 degrees C). Therefore, emphasis should be on stringent personal hygiene during preparation. For the control of foodborne viral infections, it is necessary to: Heighten awareness about the presence and spread of these viruses by foodhandlers; Optimise and standardise methods for the detection of foodborne viruses; Develop laboratory-based surveillance to detect large, common-source outbreaks at an early stage; and Emphasise consideration of viruses in setting up food safety quality control and management systems (GHP, GMP, HACCP).

Caliciviridae Infections↗

PubMed2004

Prevalence of Clostridium botulinum in food raw materials used in REPFEDs manufactured in France.

Food raw materials used in refrigerated processed foods of extended durability (REPFEDs) manufactured in France were surveyed for Clostridium botulinum types A, B and E using PCR-Enzyme-linked Immunosorbent assay (PCR-ELISA) and mouse bioassay for detection respectively of cells and toxins in enrichment broth. Portions of 25 to 50 g of food were analysed. A total of 8 out of the 102 samples of fish and shellfish, 12 out of the 143 samples of meat and poultry, 1 out of the 62 samples of aroma, sauce and gravy, 4 out of the 25 samples of thickening agents, 3 out of the 26 samples of dehydrated dairy ingredients, and none of the 65 samples of spices, herbs and dehydrated mushroom were positive for C. botulinum in PCR-ELISA, i.e., 6.6% of all the samples tested. The 28 positive samples comprised 10 type A, 10 type B, 4 with both types A and B, and 4 undetermined by PCR typing. No sample positive for type E was detected. Of the 28 samples positive in PCR-ELISA, 15 were also positive in the mouse bioassay. The MPN count was between 1 and 3 C. botulinum/kg of food, which is similar to or in the lower range of values reported in the literature.

Clostridium botulinum↗

PubMed1997

Genotypic diversity of Campylobacter lari isolated from mussels and oysters in The Netherlands.

In order to gain insight into the epidemiology of Campylobacter lari infection in The Netherlands due to the consumption of raw mussels and oysters, batches of these shellfish were screened for the presence of Campylobacter spp. during a 6 month period in 1993-1994. Apparently, 41 out of 59 batches of mussels and 11 out of 41 batches of oysters were colonized with Campylobacter spp. A subset of the isolates was further characterized by additional phenotypic tests, numerical analysis of electrophoretic protein patterns, and genotyping by random amplification of polymorphic DNA (RAPD). Protein electrophoretic analysis of 39 Campylobacter spp. cultured from 24 batches of mussels and oysters, revealed that all isolates, except two, were C. lari. Two strains with an aberrant protein pattern were identified as C. coli and C. hyointestinalis, respectively. Nalidixic acid susceptible campylobacters (NASC) and urease-positive thermophylic campylobacters (UPTC) did not form separate clusters and should be considered biovars only. Several strains were both urease positive and nalidixic acid susceptible, which represents a new biovar within C. lari. The results of RAPD demonstrated the presence of 3 distinct genetic variants, implying that even within a single batch of shell fish, relatively extensive DNA polymorphisms can be found. It is therefore apparent that this complex group of C. lari is characterized by a high degree of genetic diversity, implying the presence of a heterogenous population of C. lari in crustacean organisms living in marine waters in a restricted area in The Netherlands.

Animals↗

PubMed1999

Occurrence of Vibrio and other pathogenic bacteria in Mytilus galloprovincialis (mussels) harvested from Adriatic Sea, Italy.

Sixty-two samples of Mytilus galloprovincialis (mussels) harvested from approved shellfish waters in the Adriatic Sea were examined for the presence of Vibrio, Salmonella, Campylobacter, and verocytotoxin producing Escherichia coli. Vibrio spp. were isolated from 48.4% of samples; the species most frequently found were V. alginolyticus (32.2%) and V. vulnificus (17.7%), followed by V. cincinnatiensis (3.2%), V. parahaemolyticus (1.6%), V. fluvialis (1.6%) and V. cholerae non-O1 (1.6%). V. parahaemolyticus resulted negative to Kanagawa-phenomenon and to PCR amplification of tdh gene. V. cholerae resulted negative to PCR amplification of sto gene. No Salmonella, Campylobacter, or E. coli verocytotoxin-producing strains were isolated. The results of this study suggest the potential risk of ingesting raw or undercooked mussels due to the frequent presence of potentially pathogenic Vibrio species.

Animals↗

PubMed2001

Neutralisation and binding of VHS virus by monovalent antibody fragments.

We have previously reported the cloning and characterisation of the heavy and light chain variable domain genes encoding three monoclonal antibodies (Mabs) that bind viral haemorrhagic septicaemia virus (VHSV). Two of these antibodies, 3F1H10 and 3F1A2 both neutralised the virus though 3F1A2 appeared to recognise a broader range of virus isolates. The variable domains of these two antibodies differ by only four residues (Lorenzen et al., 2000a. Fish Shellfish Immunol. 10, 129-142). To further study the mechanism of neutralisation, Fab fragments as well as a series of recombinant bacterial single chain antibody (scAb) fragments were generated from the three anti-VHSV Mabs and their variable domain genes, respectively. Fabs and scAbs derived from the neutralising Mabs were both able to neutralise the VHSV type 1 isolate DK-F1. In addition, a series of scAb fragments were produced using the 3F1H10 variable heavy (VH) chain and variable light (Vkappa) chain domains but containing, either alone or in dual combination, each of the four different residues present in 3F1A2. The dissociation constants of Mabs 3F1H10 and 3F1A2 and their respective Fab and scAb fragments were measured by BIAcore analysis and found to correlate with the capacity of each molecule to neutralise DK-F1. These investigations, together with computer assisted molecular analysis of the theoretical influence of each mutation on antigen binding, led to the identification of a single mutation at position 35a in the VH domain as having the most marked impact on viral neutralisation.

Animals↗

PubMed2000

Application of DNA vaccine technology to aquaculture.

The aquaculture industry needs to augment its global production and efficiency to meet the increasing consumer needs for fish and shellfish products. Unfortunately, infectious diseases have been a major impediment to the development and profitability of fish farms. While vaccines offer the most efficient way to control infectious pathogens, current products have only been successful against some diseases. These are mostly bacterial, and there are still several important diseases, mainly of viral and parasitic origin, for which no prophylactic treatment exists. DNA vaccines, compared to traditional antigen vaccines, have several practical and immunological advantages that make them very attractive for the aquaculture industry. The early success of DNA vaccines in animal models was very encouraging, but fish are unique in many aspects, and findings with other classes of vertebrate, namely mammals and birds, do not necessarily apply to aquatic animals. However, more recent studies with reporter genes showed that fish cells efficiently express foreign proteins encoded by eukaryotic expression vectors. A piscine-specific backbone vector might eventually improve immune responses to DNA vaccines, but there is already strong direct evidence for the induction of protective immunity with currently available plasmids. Immune responses to plasmid DNA injected intramuscularly (IM) into fish are characterized by the production of antibodies, which have been shown to be neutralizing in two different viral disease models. There is also indirect evidence suggesting the induction of cell-mediated immunity. Despite this evidence, immune responses to DNA vaccines have only been poorly characterized in fish because of the limited knowledge of the piscine immune system, and the small number of studies on the subject. Apart from optimizing the efficiency of DNA vaccines, other important issues, such as safety and production cost will be determinants for the potential application of this technology in commercial fish farms. Alternative methods of administration will also have to be developed for small fish and low-valued species, for which IM injection is not practical and/or cost effective.

Administration, Oral↗

PubMed2001

Aquaculture pharmaceuticals and biologicals: current perspectives and future possibilities.

With the continued expansion of cultured fin and shellfish species, aquaculture has become a key component of the animal health industry. The ever-increasing demand for quality pharmaceutical and biological products, to combat bacterial and viral infections, calls for the development of modern formulations and novel drug delivery systems. The purpose of this paper is to provide a general overview of important fish pathogens, manifested fish diseases, and control methods. Currently available pharmaceutical and biological products are tabulated. Emphasis is placed on the extension of formulation and process technologies, commonly encountered in human pharmaceuticals, to the aquaculture arena. New research efforts in the area of novel vaccine delivery systems, i.e. enteric-coated beads and biodegradable microspheres, are also summarized.

Anesthetics↗

PubMed1989

Occupational dermatitis in food handlers and bakers.

Of 1346 patients with suspected contact dermatitis examined in Toronto, 10 bakers and 10 food handlers were found to have occupational contact dermatitis. Although not dissimilar in age, years in the trade, or length of illness, men were significantly overrepresented in the population of bakers (100% vs 40%, p less than 0.05). One baker had occupational asthma related to rye flour. Compared with the bakers, significantly more of the food handlers had allergic contact dermatitis (70% vs 20%, p less than 0.05). In addition to food components, cinnamates and rubber were found to be causes of allergic response. One food handler had contact urticaria caused by shellfish. The standard screening tests, combined with the use of contactants specific to occupational history, yielded a definitive determination of the existence of a contact allergy in our subjects.

Adult↗

PubMed1997

Neurotoxin domoic acid produces cytotoxicity via kainate- and AMPA-sensitive receptors in cultured cortical neurones.

Domoic acid, a naturally occurring kainoid, has been responsible for several outbreaks of fatal poisoning after shellfish ingestion, and we examined its neurotoxic mechanism in cultured murine cortical neurones. Using observations of neuronal viability and morphology, exposure to domoic acid for 24 h was found to induce substantial concentration-dependent neuronal cell death. Domoic acid-mediated neuronal death was attenuated by the non-N-methyl-D-aspartate receptor antagonist 6-cyano-7-nitro-quinoxaline-2,3-dione and the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor-selective antagonist LY293558 ((3S,4aR,6R,8aR)-6-[2-(1H-tetrazol-5-yl)-ethyl]-1,2,3, 4,4a,5,6,7,8,8a-decahydroisoquinoline-3-carboxylic acid), but unaffected by NS-102 (5-nitro-6,7,8,9-tetrahydrobenzo[g]indole-2, 3-dione-3-oxime)--a low-affinity kainate receptor antagonist. Domoic acid was equipotent with (S)-AMPA (EC50 values 3.8 and 3.4 microM respectively); however, (S)-AMPA induced only 50% cell death compared to > 80% cell death induced by domoic acid. Kainate also killed > 80% of cortical neurones; however, domoic acid was about 19 times more potent than kainate (EC50 75 microM). We show the potent neurotoxicity of domoic acid for the first time in a pure neuronal model and indicate that domoic acid acts via high-affinity AMPA- and kainate-sensitive glutamate receptors to produce excitotoxic cell death.

Animals↗