Dataset Information

Application of Six Detection Methods for Analysis of Paralytic Shellfish Toxins in Shellfish from Four Regions within Latin America.

ABSTRACT: With the move away from use of mouse bioassay (MBA) to test bivalve mollusc shellfish for paralytic shellfish poisoning (PSP) toxins, countries around the world are having to adopt non-animal-based alternatives that fulfil ethical and legal requirements. Various assays have been developed which have been subjected to single-laboratory and multi-laboratory validation studies, gaining acceptance as official methods of analysis and approval for use in some countries as official control testing methods. The majority of validation studies conducted to date do not, however, incorporate shellfish species sourced from Latin America. Consequently, this study sought to investigate the performance of five alternative PSP testing methods together with the MBA, comparing the PSP toxin data generated both qualitatively and quantitatively. The methods included a receptor binding assay (RBA), two liquid chromatography with fluorescence detection (LC-FLD) methods including both pre-column and post-column oxidation, liquid chromatography with tandem mass spectrometry (LC-MS/MS) and a commercial lateral flow assay (LFA) from Scotia. A total of three hundred and forty-nine shellfish samples from Argentina, Mexico, Chile and Uruguay were assessed. For the majority of samples, qualitative results compared well between methods. Good statistical correlations were demonstrated between the majority of quantitative results, with a notably excellent correlation between the current EU reference method using pre-column oxidation LC-FLD and LC-MS/MS. The LFA showed great potential for qualitative determination of PSP toxins, although the findings of high numbers of false-positive results and two false negatives highlighted that some caution is still needed when interpreting results. This study demonstrated that effective replacement methods are available for countries that no longer wish to use the MBA, but highlighted the importance of comparing toxin data from the replacement method using local shellfish species of concern before implementing new methods in official control testing programs.

SUBMITTER: Turner AD

PROVIDER: S-EPMC7761785 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Application of Six Detection Methods for Analysis of Paralytic Shellfish Toxins in Shellfish from Four Regions within Latin America.

Turner Andrew D AD Tarnovius Sophie S Hatfield Robert G RG Teixeira-Alves Mickael M Broadwater Maggie M Dolah Frances Van FV Garcia-Mendoza Ernesto E Medina Dinorah D Salhi Maria M Goya Alejandra B AB Barrera Fernanda F Carrasco Daniel D Rubilar Ignacio I Suarez-Isla Benjamin A BA

Marine drugs 20201203 12

With the move away from use of mouse bioassay (MBA) to test bivalve mollusc shellfish for paralytic shellfish poisoning (PSP) toxins, countries around the world are having to adopt non-animal-based alternatives that fulfil ethical and legal requirements. Various assays have been developed which have been subjected to single-laboratory and multi-laboratory validation studies, gaining acceptance as official methods of analysis and approval for use in some countries as official control testing meth ...[more]

PMID: 33287439

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Project description:Paralytic shellfish poisoning toxins (PSTs) are a family of more than 30 natural alkaloids synthesized by dinoflagellates and cyanobacteria whose toxicity in animals is mediated by voltage-gated Na(+) channel blocking. The export of PST analogues may be through SxtF and SxtM, two putative MATE (multidrug and toxic compound extrusion) family transporters encoded in PSTs biosynthetic gene cluster (sxt). sxtM is present in every sxt cluster analyzed; however, sxtF is only present in the Cylindrospermopsis-Raphidiopsis clade. These transporters are energetically coupled with an electrochemical gradient of proton (H(+)) or sodium (Na(+)) ions across membranes. Because the functional role of PSTs remains unknown and methods for genetic manipulation in PST-producing organisms have not yet been developed, protein structure analyses will allow us to understand their function. By analyzing the sxt cluster of eight PST-producing cyanobacteria, we found no correlation between the presence of sxtF or sxtM and a specific PSTs profile. Phylogenetic analyses of SxtF/M showed a high conservation of SxtF in the Cylindrospermopsis-Raphidiopsis clade, suggesting conserved substrate affinity. Two domains involved in Na(+) and drug recognition from NorM proteins (MATE family) of Vibrio parahaemolyticus and V. cholerae are present in SxtF/M. The Na(+) recognition domain was conserved in both SxtF/M, indicating that Na(+) can maintain the role as a cation anti-transporter. Consensus motifs for toxin binding differed between SxtF and SxtM implying differential substrate binding. Through protein modeling and docking analysis, we found that there is no marked affinity between the recognition domain and a specific PST analogue. This agrees with our previous results of PST export in R. brookii D9, where we observed that the response to Na(+) incubation was similar to different analogues. These results reassert the hypothesis regarding the involvement of Na(+) in toxin export, as well as the motifs L(398)XGLQD(403) (SxtM) and L(390)VGLRD(395) (SxtF) in toxin recognition.

Project description:In late February 2016, a harmful algal bloom (HAB) of Alexandrium catenella was detected in southern Chiloé, leading to the banning of shellfish harvesting in an extended geographical area (~500 km). On April 24, 2016, this bloom produced a massive beaching (an accumulation on the beach surface of dead or impaired organisms which were drifted ashore) of surf clams Mesodesma donacium in Cucao Bay, Chiloé. To determine the effect of paralytic shellfish poisoning (PSP) toxins in M. donacium, samples were taken from Cucao during the third massive beaching detected on May 3, 2016. Whole tissue toxicity evidence a high interindividual variability with values which ranged from 1008 to 8763 μg STX eq 100 g-1 and with a toxin profile dominated by GTX3, GTX1, GTX2, GTX4, and neoSTX. Individuals were dissected into digestive gland (DG), foot (FT), adductor muscle (MU), and other body fractions (OBF), and histopathological and toxin analyses were carried out on the obtained fractions. Some pathological conditions were observed in gill and digestive gland of 40⁻50% of the individuals that correspond to hemocyte aggregation and haemocytic infiltration, respectively. The most toxic tissue was DG (2221 μg STX eq 100 g-1), followed by OBF (710 μg STX eq 100 g-1), FT (297 μg STX eq 100 g-1), and MU (314 μg STX eq 100 g-1). The observed surf clam mortality seems to have been mainly due to the desiccation caused by the incapability of the clams to burrow. Considering the available information of the monitoring program and taking into account that this episode was the first detected along the open coast of the Pacific Ocean in southern Chiloé, it is very likely that the M. donacium population from Cucao Bay has not had a recurrent exposition to A. catenella and, consequently, that it has not been subjected to high selective pressure for PSP resistance. However, more research is needed to determine the effects of PSP toxins on behavioral and physiological responses, nerve sensitivity, and genetic/molecular basis for the resistance or sensitivity of M. donacium.

Dataset Information

Application of Six Detection Methods for Analysis of Paralytic Shellfish Toxins in Shellfish from Four Regions within Latin America.

Publications

Application of Six Detection Methods for Analysis of Paralytic Shellfish Toxins in Shellfish from Four Regions within Latin America.

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