Dataset Information

Rapid PCR-lateral flow assay for the onsite detection of Atlantic white shrimp.

ABSTRACT: The Atlantic white shrimp (Litopenaeus setiferus) is of great economic importance to the United States and risk being substituted with imported species due to a shortage in domestic production. To improve the current methods used for the identification of the Atlantic white shrimp species, we designed and validated a robust multiplex PCR-lateral flow assay for the onsite identification of L. setiferus. The standardized assay was validated using a miniaturized, low-cost PCR instrument with 68 shrimp, prawn, and fish samples, spread over fourteen seafood species. L. setiferus was simultaneously amplified by the multiplex assay to give three visual bands, which distinguished it from other species having either one or two bands on the dipstick. The standardized assay showed 100% inclusivity for target L. setiferus samples, 100% exclusivity for non-target samples and can be completed in less than two hours. The assay standardized in this study can be used for onsite testing of L. setiferus samples at processing facilities, restaurants, and wholesalers' facilities.

SUBMITTER: Kwawukume S

PROVIDER: S-EPMC9868375 | biostudies-literature | 2023 Jul

REPOSITORIES: biostudies-literature

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Rapid PCR-lateral flow assay for the onsite detection of Atlantic white shrimp.

Kwawukume Samuel S Velez Frank J FJ Williams David D Cui Leqi L Singh Prashant P

Food chemistry. Molecular sciences 20230116

The Atlantic white shrimp (<i>Litopenaeus setiferus</i>) is of great economic importance to the United States and risk being substituted with imported species due to a shortage in domestic production. To improve the current methods used for the identification of the Atlantic white shrimp species, we designed and validated a robust multiplex PCR-lateral flow assay for the onsite identification of <i>L. setiferus</i>. The standardized assay was validated using a miniaturized, low-cost PCR instrume ...[more]

PMID: 36698370

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Project description:Melioidosis is a severe infectious disease caused by gram-negative, facultative intracellular pathogen Burkholderia pseudomallei (B. pseudomallei). Although cases are increasing reported from other parts of the world, it is an illness of tropical and subtropical climates primarily found in southeast Asia and northern Australia. Because of a 40% mortality rate, this life-threatening disease poses a public health risk in endemic area. Early detection of B. pseudomallei infection is vital for prognosis of a melioidosis patient. In this study, a novel isothermal recombinase polymerase amplification combined with lateral flow dipstick (LF-RPA) assay was established for rapid detection of B. pseudomallei. A set of primer-probe targeting orf2 gene within the putative type III secretion system (T3SS) cluster genes was generated and parameters for the LF-RPA assay were optimized. Result can be easy visualized in 30 minutes with the limit of detection (LOD) as low as 20 femtogram (fg) (ca. 25.6 copies) of B. pseudomallei genomic DNA without a specific equipment. The assay is highly specific as no cross amplification was observed with Burkholderia mallei, members of the Burkholderia cepacia-complex and 35 non-B. pseudomallei bacteria species. Moreover, isolates from patients in Hainan (N = 19), Guangdong (N = 1), Guangxi (N = 3) province of China as well as in Australia (N = 3) and Thailand (N = 1) were retrospectively confirmed by the newly developed method. LODs for B. pseudomallei-spiked soil and blood samples were 2.1×103 CFU/g and 4.2×103 CFU/ml respectively. The sensitivity of the LF-RPA assay was comparable to TaqMan Real-Time PCR (TaqMan PCR). In addition, the LF-RPA assay exhibited a better tolerance to inhibitors in blood than TaqMan PCR. Our results showed that the LF-RPA assay is an alternative to existing PCR-based methods for detection of B. pseudomallei with a potentiality of early accurate diagnosis of melioidosis at point of care or in-field use.

Dataset Information

Rapid PCR-lateral flow assay for the onsite detection of Atlantic white shrimp.

Publications

Rapid PCR-lateral flow assay for the onsite detection of Atlantic white shrimp.

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