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Molecular identification of Trichinella species by multiplex PCR: new insight for Trichinella murrelli.

ABSTRACT: In order to identify Trichinella at the species level, the commonly used test is a multiplex PCR, allowing the discrimination of nine out of the twelve taxa described so far. This test is based on five primer pairs amplifying fragments of the large subunit rDNA. Each taxon produces one or two bands of different sizes, resulting in a specific band pattern. By multiplex PCR, Trichinella?murrelli shows two bands of 127?bp and 316?bp. However, a third band of 256?bp can occur. This band can lead to misidentification, since it is similar to the 253?bp band displayed by Trichinella?britovi. BLAST analysis confirmed that the 256?bp band is from T. murrelli. The aim of this short note is to inform analysts that T.?murrelli larvae may display either two- or three-band patterns.

SUBMITTER: Karadjian G

PROVIDER: S-EPMC5721686 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Molecular identification of Trichinella species by multiplex PCR: new insight for Trichinella murrelli.

Karadjian Grégory G Heckmann Aurélie A Rosa Giuseppe La G Pozio Edoardo E Boireau Pascal P Vallée Isabelle I

Parasite (Paris, France) 20171208

In order to identify Trichinella at the species level, the commonly used test is a multiplex PCR, allowing the discrimination of nine out of the twelve taxa described so far. This test is based on five primer pairs amplifying fragments of the large subunit rDNA. Each taxon produces one or two bands of different sizes, resulting in a specific band pattern. By multiplex PCR, Trichinella murrelli shows two bands of 127 bp and 316 bp. However, a third band of 256 bp can occur. This band can lead to ...[more]

PMID: 29219110

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Project description:BACKGROUND Human trichinellosis is a foodborne parasitic zoonotic disease caused by ingestion of raw or undercooked meat infected with nematode larvae of the genus Trichinella. In the USA, sporadic cases and outbreaks caused by the consumption of wild game meat infected with Trichinella have been reported. The current methods for diagnosis such as serology and microscopy are not specific, may result in false negative results, and cannot differentiate encapsulated Trichinella larvae to species level. The molecular protocols currently available for the differentiation of all encapsulate Trichinella species prevalent in North America have some limitations such as the inability to identify and resolve the presence of several Trichinella species in a single test. OBJECTIVES/METHODS In this study we developed and evaluated a multiplex TaqMan quantitative real-time polymerase chain reaction (qPCR) assay, which can simultaneously detect, identify and differentiate all species of encapsulated Trichinella occurring in North America i.e., T. nativa, T. spiralis, T. murrelli and Trichinella T6, even in cases of multiple infection in a single sample. We investigated two human biopsies and 35 wild animal meat samples considered as having a high likelihood of harboring Trichinella larvae obtained from the United States during 2009-2017. FINDINGS Using the multiplex assay describe here, 22 (59%) samples that tested positive contained Trichinella spp., were identified as: T. nativa (n = 7, including a human biopsy), T. spiralis (n = 9, including a human biopsy), T. murrelli (n = 3), Trichinella T6 (n = 1). Results also included two rare mixed infection cases in bears, a T. nativa/T. spiralis from Alaska and a T. spiralis/Trichinella T6 from California. The species identifications were confirmed using a conventional PCR targeting the rRNA ITS1-ITS2 region, followed by DNA sequencing analysis. The estimated limit of detection (LOD) was approximately seven larvae per gram of meat. MAIN CONCLUSIONS Differentiation of Trichinella spp. is needed to improve efforts on identification of case, optimize food safety control and better understand the geographic distribution of Trichinella species. The Trichinella qPCR multiplex proved to be a robust, easy to perform assay and is presented as an improved technique for identification of all known encapsulated species occurring in North America continent.

Dataset Information

Molecular identification of Trichinella species by multiplex PCR: new insight for Trichinella murrelli.

Publications

Molecular identification of Trichinella species by multiplex PCR: new insight for Trichinella murrelli.

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