Abstract
BackgroundThe People’s Republic of China (P.R. China) is the presumptive home range of the rat lungworm Angiostrongylus cantonensis, a major aetiological agent of human eosinophilic meningitis. We present a study of the genetic variation of A. cantonensis in P.R. China. Our aim was to deepen the current knowledge pertaining to its origin and global spread from a molecular perspective.MethodsAdult A. cantonensis were collected in the frame of a national survey and identified based on morphological criteria. Polymerase chain reaction (PCR) was employed to amplify the target DNA sequences (cytochrome c oxidase subunit I (cox1), nicotinamide adenine dinucleotide dehydrogenase subunit 1 (nad1) and internal transcribed spacer (ITS)). The PCR product of cox1 was directly submitted to sequencing, while clone sequencing was used for nad1 and ITS. The identity of the samples was verified by comparing the sequences to those of accepted A. cantonensis specimens. The specific composition of substitutions in each gene was analysed, and the genotypes were compared based on the complete cox1, nad1 and ITS genes.ResultsWe characterised the complete mitochondrial genes cox1 and nad1 of 130 specimens and obtained 357 nuclear sequences containing two complete ITS (ITS1 and ITS2) and 5.8S rRNA of the same samples. All specimens were genetically confirmed as A. cantonensis. Two major groups (i.e. I and II) were identified according to the phylogeny of cox1 sequences. Group I could be further categorised into six distinct clades. Almost half of the specimens (47.7%) belong to the clade Ia and 22.3% to the group II. The former was widely distributed across the study region. A variable number of repeat units in three microsatellites was observed, resulting in considerable length variation in ITS. Intragenomic variation of ITS sequences was found in a large proportion of the samples. Genotyping showed a striking difference between mitochondrial DNA and ITS.ConclusionsOur results demonstrate that A. cantonensis is the only rat lungworm species in P.R. China and shows high genetic diversity. Results of diversity and genotyping of A. cantonensis can be impacted by the sequencing strategy and biomarker. Although ITS may be a valuable marker for interspecific identification, it is not suitable for studying the intraspecific variation of A. cantonensis due to its high intragenomic variation and current challenges for direct sequencing.
Highlights
Variation of mitochondrial genes The nad1 and cox1 genes of a total of 130 A. cantonensis specimens from 32 collection sites, representing the whole known endemic area of angiostrongyliasis in P.R
Among the 843 sites of the complete nad1 gene and 1577 sites of the complete cox1 gene, 171 variable sites were identified in each gene
We identified seven distinct clades based on the analysis of mitochondrial genes, which indicates a high diversity of A. cantonensis in its current presumptive home range
Summary
The People’s Republic of China (P.R. China) is the presumptive home range of the rat lungworm Angiostrongylus cantonensis, a major aetiological agent of human eosinophilic meningitis. The rat lungworm Angiostrongylus cantonensis is an important aetiological agent of human eosinophilic meningitis [1]. The life cycle of A. cantonensis involves rats as definitive host and mollusks as intermediate host. Humans acquire infections mainly through the consumption of undercooked snails that harbour infective larvae. Larvae mainly dwell in the vessels of the central nervous system (CNS) and only rarely migrate to the pulmonary arteries [2]. Inflammation in the CNS is the proximate cause of morbidity due to an infection with A. cantonensis. More than 3000 human cases have been reported [1]
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