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A Novel Sensitive Cell-Based Immunoenzymatic Assay for Palytoxin Quantitation in Mussels.

ABSTRACT: The marine algal toxin palytoxin (PLTX) and its analogues are some of the most toxic marine compounds. Their accumulation in edible marine organisms and entrance into the food chain represent their main concerns for human health. Indeed, several fatal human poisonings attributed to these compounds have been recorded in tropical and subtropical areas. Due to the increasing occurrence of PLTX in temperate areas such as the Mediterranean Sea, the European Food Safety Authority (EFSA) has suggested a maximum limit of 30 µg PLTX/kg in shellfish meat, and has recommended the development of rapid, specific, and sensitive methods for detection and quantitation of PLTX in seafood. Thus, a novel, sensitive cell-based ELISA was developed and characterized for PLTX quantitation in mussels. The estimated limits of detection (LOD) and quantitation (LOQ) were 1.2 × 10-11 M (32.2 pg/mL) and 2.8 × 10-11 M (75.0 pg/mL), respectively, with good accuracy (bias = 2.5%) and repeatability (15% and 9% interday and intraday relative standard deviation of repeatability (RSDr), respectively). Minimal interference of 80% aqueous methanol extract allows PLTX quantitation in mussels at concentrations lower than the maximum limit suggested by EFSA, with an LOQ of 9.1 µg PLTX equivalent/kg mussel meat. Given its high sensitivity and specificity, the cell-based ELISA should be considered a suitable method for PLTX quantitation.

SUBMITTER: Pelin M

PROVIDER: S-EPMC6116170 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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A Novel Sensitive Cell-Based Immunoenzymatic Assay for Palytoxin Quantitation in Mussels.

Pelin Marco M Sosa Silvio S Brovedani Valentina V Fusco Laura L Poli Mark M Tubaro Aurelia A

Toxins 20180814 8

The marine algal toxin palytoxin (PLTX) and its analogues are some of the most toxic marine compounds. Their accumulation in edible marine organisms and entrance into the food chain represent their main concerns for human health. Indeed, several fatal human poisonings attributed to these compounds have been recorded in tropical and subtropical areas. Due to the increasing occurrence of PLTX in temperate areas such as the Mediterranean Sea, the European Food Safety Authority (EFSA) has suggested ...[more]

PMID: 30110919

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Project description:CCR5 antagonists are a powerful new class of antiretroviral drugs that require a companion assay to evaluate the presence of CXCR4-tropic (non-R5) viruses prior to use in human immunodeficiency virus (HIV)-infected individuals. In this study, we have developed, characterized, verified, and prevalidated a novel phenotypic test to determine HIV-1 coreceptor tropism (VERITROP) based on a sensitive cell-to-cell fusion assay. A proprietary vector was constructed containing a near-full-length HIV-1 genome with the yeast uracil biosynthesis (URA3) gene replacing the HIV-1 env coding sequence. Patient-derived HIV-1 PCR products were introduced by homologous recombination using an innovative yeast-based cloning strategy. The env-expressing vectors were then used in a cell-to-cell fusion assay to determine the presence of R5 and/or non-R5 HIV-1 variants within the viral population. Results were compared with (i) the original version of Trofile (Monogram Biosciences, San Francisco, CA), (ii) population sequencing, and (iii) 454 pyrosequencing, with the genotypic data analyzed using several bioinformatics tools, i.e., the 11/24/25 rule, Geno2Pheno (2% to 5.75%, 3.5%, or 10% false-positive rate [FPR]), and webPSSM. VERITROP consistently detected minority non-R5 variants from clinical specimens, with an analytical sensitivity of 0.3%, with viral loads of ≥1,000 copies/ml, and from B and non-B subtypes. In a pilot study, a 73.7% (56/76) concordance was observed with the original Trofile assay, with 19 of the 20 discordant results corresponding to non-R5 variants detected using VERITROP and not by the original Trofile assay. The degree of concordance of VERITROP and Trofile with population and deep sequencing results depended on the algorithm used to determine HIV-1 coreceptor tropism. Overall, VERITROP showed better concordance with deep sequencing/Geno2Pheno at a 0.3% detection threshold (67%), whereas Trofile matched better with population sequencing (79%). However, 454 sequencing using Geno2Pheno at a 10% FPR and 0.3% threshold and VERITROP more accurately predicted the success of a maraviroc-based regimen. In conclusion, VERITROP may promote the development of new HIV coreceptor antagonists and aid in the treatment and management of HIV-infected individuals prior to and/or during treatment with this class of drugs.

Dataset Information

A Novel Sensitive Cell-Based Immunoenzymatic Assay for Palytoxin Quantitation in Mussels.

Publications

A Novel Sensitive Cell-Based Immunoenzymatic Assay for Palytoxin Quantitation in Mussels.

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