PCR-based molecular characterization and in-silico analysis of food-borne trematode parasites Paragonimus westermani, Fasciolopsis buski and Fasciola gigantica from Northeast India using ITS2 rDNA

Abstract

Food-borne fluke infections/trematodiases are emerging as a major public health problem worldwide with over 40 million people affected and over 10% of world population at risk of infection. The major concentration of these infections is in Southeast Asian and Western Pacific Regions, where the epidemiological factors (including the prevalent socio-cultural food habits) are conducive for transmission of these infections. The preponderance of these infections is usually in food deficit poor communities that lack access to proper sanitary infrastructure. While targeting health for all, especially the poor rural tribal communities, it is imperative to take these infections into account. Bayesian analysis phylogeny of food-borne trematode parasites under study showed that they are closely related phylogenetic groups. To focus the control strategies at the target populations, the aim of the present study was to establish molecular methods for accurate discrimination between common food-borne trematodes parasites Paragonimus (lung fluke), Fasciolopsis (giant intestinal fluke) and Fasciola (liver fluke), the infections of which commonly prevail in NE India. In the first step, we amplified and sequenced the second internal transcribed spacer (ITS2) region of ribosomal DNA, utilizing nucleotide differences between the multiple sequence alignments of the parasites under study. Based upon the differences in nucleotide sequences of conserved regions, we designed species-specific primers that can unequivocally discriminate one species from another. ITS2 sequence motifs allowed an accurate in-silico distinction of the trematodes. The data indicate that ITS2 motifs (≤ 50 bp in size) can be considered promising tool for trematode species identification. Using molecular morphometrics that is based on ITS2 secondary structure homologies, phylogenetic relationships with various isolates of several trematode species have been discussed. The present results suggest that the ITS2 specific primers can be used for epidemiological investigations of the prevalence of trematodiasis.

Molecular confirmation of trematodes in the snail intermediate hosts from Ratchaburi Province, Thailand

“…Molecular biological methods have been acknowledged as the most efficient and accurate tools that can be used for the identification of several parasitic organisms including trematodes [1,14,[19][20][21]. The internal transcribed spacer 2 (ITS2) of the 18S rDNA gene was selected and used in this study.…”

“…Conventional PCR methods have been widely applied for the purposes of identification because the DNA method is very sensitive, highly accurate and can be rapidly applied. Therefore, the sequence data have proven to be helpful in the study of species identification and geographical distribution for numerous species such as Paragonimus westermani, Fasciolopsis buski and Fasciola gigantica [20], Explanatum explanatum, Paramphistomum epiclitum and Calicophoron calicophorum [25].…”

Molecular characterization of the Indian poultry nodular tapeworm, Raillietina echinobothrida (Cestoda: Cyclophyllidea: Davaineidae) based on rDNA internal transcribed spacer 2 region

“…In recent years, several molecular tools including sequence data related to conserved domains have unveiled their utility in molecular identification, taxonomy, systematics and phylogeny of floral, faunal and microbial species, too (Huang et al 2006;Prasad et al 2009a, b;Wiemers et al 2009;Yao et al 2010;Sharma et al 2012). Since Woese and Fox (1977), the nuclear ribosomal repeat cistrons have also been widely implied for parasites (Littlewood and Olson 2001;Waeschenbach et al 2007;Jyrwa et al 2009;Goswami et al 2009;Tandon et al 2010;Prasad et al 2011;Ghatani et al 2012). Molecular phylogeny has advanced our understanding of comparative studies in basic and applied biology and more generally of their position within the phylum Platyhelminthes (Brooks and Hoberg 2001;Tandon et al 2010).…”

Molecular characterization of three species belongs to the Allocreadioidea, Hemiuroidea and Plagiorchioidea (Platyhelminthes: Trematoda) infecting freshwater fishes in India

“…In India, for platyhelminthes taxonomy morphological differences have been widely used to discriminate between species. Though, traditional diagnostic tools are now complemented by molecular techniques for resolving the taxonomic confusions coupled with species descriptions (Prasad et al, 2011;Chaudhary et al, 2015;Chaudhary et al, 2016). In India, there is a scarcity of molecular tools in studies of trematode parasites.…”