Newly developed SYBR Green-based quantitative real-time PCRs revealed coinfection evidence of Angiostrongylus cantonensis and A. malaysiensis in Achatina fulica existing in Bangkok Metropolitan, Thailand.

Autor:	Jakkul W; Department of Helminthology, Faculty of Tropical Medicine, Mahidol University, Thailand Mahidol University, Thailand., Chaisiri K; Department of Helminthology, Faculty of Tropical Medicine, Mahidol University, Thailand Mahidol University, Thailand., Saralamba N; Department of Molecular Tropical Medicine and Genetics, Faculty of Tropical Medicine, Mahidol University, Thailand., Limpanont Y; Department of Social and Environmental Medicine, Faculty of Tropical Medicine, Mahidol University, Thailand., Dusitsittipon S; Department of Parasitology and Entomology, Faculty of Public Health, Mahidol University, Thailand., Charoennitiwat V; Department of Helminthology, Faculty of Tropical Medicine, Mahidol University, Thailand Mahidol University, Thailand., Chan AHE; Department of Helminthology, Faculty of Tropical Medicine, Mahidol University, Thailand Mahidol University, Thailand., Thaenkham U; Department of Helminthology, Faculty of Tropical Medicine, Mahidol University, Thailand Mahidol University, Thailand.
Jazyk:	angličtina
Zdroj:	Food and waterborne parasitology [Food Waterborne Parasitol] 2021 Mar 15; Vol. 23, pp. e00119. Date of Electronic Publication: 2021 Mar 15 (Print Publication: 2021).
DOI:	10.1016/j.fawpar.2021.e00119
Abstrakt:	Angiostrongylus cantonensis is a well-known pathogen causing eosinophilic meningitis associated with angiostrongyliasis. Humans, as accidental hosts, are infected by consuming undercooked snails containing third-stage larvae. A. malaysiensis is closely related to A. cantonensis and has been described as a potential human pathogen. The two species distribution was recently reported to overlap in the same endemic area, particularly in the Indochina Peninsula. Similar morphological characteristics of the third-stage larva in the snail-intermediate host often lead to misidentification of the two species. Thus, we aimed to develop a sensitive and specific method to detect and discriminate Angiostrongylus third-stage larva by designing species-specific primers based on the mitochondrial cytochrome b gene. We developed the SYBR Green quantitative real-time PCR (qPCR) method for two species-specific detection assays, which could be conducted simultaneously. The method was subsequently employed to detect and identify third-stage larvae of Angiostrongylus isolated from infected Achatina fulica collected from six public parks in Bangkok Metropolitan, Thailand. The method was also a preliminary applied to detect parasite tissue debris in the patients' cerebrospinal fluid (CSF). SYBR Green qPCRs quantitatively detected approximately 10 -4  ng of genomic DNA from one larva, facilitating species-specific detection. Based on the pools of third-stage larvae isolated individually from the tissue of each infected A. fulica collected from the public parks, the qPCR results revealed that A. malaysiensis was the predominant species infecting 5.26% of the collected snails. In comparison, coinfection between A. malaysiensis and A. cantonensis was 5.97%, and no single infection of A. cantonensis was detected in A. fulica . Our SYBR Green qPCR method is a useful and inexpensive technique for A. cantonensis and A. malaysiensis discrimination, and the method has sufficient sensitivity to detect isolated larvae from a snail-intermediate host. The ratio of A. cantonensis and A. malaysiensis larvae infecting the snails can also be estimated simultaneously. Our qPCRs can be employed in a molecular survey of A. cantonensis and A. malaysiensis within intermediate hosts and for clinical diagnosis of angiostrongyliasis with CSF specimens in future studies. Competing Interests: None. (© 2021 The Authors.)
Databáze:	MEDLINE
Externí odkaz:	Zobrazit plný text záznamu