Epidemiology of cercarial stage of trematodes in freshwater snails from Chiang Mai province,Thailand

Objective:To investigate the epidemiological situation of cercarial trematodes infection in freshwater snails from different water resources in Chiang Mai province,Thailand.Methods:The snail specimens were collected from 13 districts of Chiang Mai province during April 2008 to February 2012.The prevalence of cercarial infection in snails was investigated using the crushing method.The drawing was done with the help of a camera lucida for the morphological study.Results:A total of 2479 snail individuals were collected and classified into 7 families,11 genera,and 14 species,Among them,8 snails species were found to be infected with an overall prevalence of 17.27%(428/2479),which infected with nine groups of cercariae;gymnocephalous cercaria,strigea cercaria,megalurous cercaria,monostome cercaria,parapleurolophocercous cercaria(Haplorchis cercaria),pleurolophocercous cercaria,furcocercous cercaria(Transversotrema cercaria),xiphidiocercaria,and virgulate cercaria.The parapleurolophocercous cercaria was found to be the dominant type among the cercarial infection in the snails(64.25%).Conclusions:The various species of snails found in the research location act as the intermediate hosts for the high prevalence of parasitic infection of many species of mammals.This work will provide new information on both the distribution and first intermediate host of trematodes.

Epidemiological situation and molecular identification of cercarial stage in freshwater snails in Chao-Phraya Basin, Central Thailand

Objective: To investigate the prevalence of cercarial trematode infection in snails and to examine the reconstruction of the phylogenetic relationship to explain the molecular system of cercarial stage trematodes to estimate the infection rate of in the definite host from the Chao-Phraya Basin.Methods: The snails were collected from 10 provinces of the Chao-Phraya Basin,Thailand by stratified sampling method. The snails were examined for cercarial infection by the crushing method. All DNA specimens were amplified with internal transcribed spacer 3(ITS3) and ITS4 primer based on PCR technique. The sequence data were aligned and used to reconstruct the phylogenetic tree by unweighted pair-group method with arithmetic means with 10 000 bootstraps.Results: The overall rate of cercarial infection was found to be 5.90%(122/2 067). Snails in the family Thiaridae were found to be in the highest prevalence followed by Lymnaeidae, Bithyniidae, Planorbidae, Viviparidae, and Ampullariidae, respectively, while the Buccinidae family(Clea helena) did not reveal any infections. The frequently found species of cercariae were parapleurolophocercous cercariae, cercariae and megarulous cercariae. The monophyletic tree separated the snails into five groups comprised of Heterophyidae, Strigeidae, Lecithodendriidae, Philophthalmidae and Echinostomatidae using the sequence of Angiostrongylus cantonensis as an out-group.Conclusions: This study was the first to report on cercarial infection in the Chao-Phraya Basin, Thailand. This revealed that a high variety of freshwater snails were infected by cercariae stage trematodes with a high prevalence. The sequence data of ITS2 can be used to investigate the phylogenetic relationships of trematodes at the family level and in each clade of different families separated by the definitive hosts.

Epidemiology and molecular genotyping of echinostome metacercariae in Filopaludina snails in Lamphun Province,Thailand

Objective:To analyze the prevalence of echinostome metacercariae in Filopaludina dorliaris(F.dorliaris)and Filopaludina martensi martensi(F.martensi martensi)and genotype variation of echinostome metacercariae by using random amplified polymorphic DNA(RAPD)analysis.Methods:Filopaludina sp.snails were collected from eight localities of Lamphun Province,Northern Thailand and examined for echinostome metacercariae.RAPD-PCR was used lo analyze genotype variation of echinostome metacercariae.Results:A total of 3 226 F.dorliaris and F.martensi martensi snails were collected from eight localities.The overall prevalences of echinostome metacercariae in F.dorliaris and F.martensi martensi were 40.89%and 36.27%,while the intensity of infection was 20.37 and 12.04,respectively.The dendrogram constructed base on RAPD profiles,4 well supported domains were generated;(i)group of metacercariae from Ban Hong,Mae Ta,Meaung,Pa Sang,Toong Hua Chang,and Weang Nong that were clustered in the group of E.revolutum,(ii)Ban Thi,(iii)Lee,and(iv)3 adults of an out group.Condusioiis:This research demonstrated RAPD profiling has been a useful tool to detect DNA polymorphisms to determine genetic relationship between echinostome metacercariae in Lamphun Province,Northern Thailand.

Molecular phylogeny of trematodes in Family Heterophyidae based on mitochondrial cytochrome c oxidase subunit Ⅰ(mCO Ⅰ)

Objective:To analyze a phylogenetic tree for understanding the molecular systematic of trematode in Pamily Heterophyidae.which are highly distributed in Thailand.Methods:Based on thirteen sequences of mitochondrial cytochrome C oxidase subunit 1(mCO I) gene from six genera of heterophyid trernatodes,riz.Haplorchis,Stellantchasmus,Centrocestus,Metagonimus,Pygidopsis,and Haplnrchoides were aligned automatically using the Clustal × 2.0 program.A phylogenetic tree was constructed by maximum likeiughood(ML) and neighbor-joining(NJ)methods,with 1000 bootstrap using the 5.0 program.Results:The phylogenetic relationship from both methods was similar and separated into three groups consisting of Haplorchoides pumilio group,Haplorchoides taichui group and another heterophyid genera.Comclusions:The sequence data of mtCOl can he used to investigate the phylogenetic relationships of trernatodes at the genus level.Each clade of different genera of heterophyid trernatodes can be separated into sister groups that correlated with the morphological characteristic,kind of secondary intermediate host and geographic distribution.

Prevalence of cercarial infections in freshwater snails and morphological and molecular identification and phylogenetic trends of trematodes

Objective:To investigate the prevalence of cercarial infections in freshwater snails from several water sources in Nakhon Nayok,Nonthaburi,and Pathum Thani provinces of Central Thailand,and to reconstruct a phylogenetic tree for improved understanding of the relationships in the cercarial stage.Methods:The snail specimens were collected from 34 total sampling sites and investigated for cercarial infections using the crushing method.The cercarial specimens were classified and used for the phylogenetic tree analysis using the Internal Transcribed Spacer 2(ITS2).Results:A total of 1921 snail specimens were classified into five families and seven species.The results showed that four snail species were identified as intermediate hosts of the larval stages of trematodes,with an overall prevalence of infection of 2.45%(47/1921).The infected snail specimens included five groups of the cercarial type:cercariaeum cercariae,echinostome cercaria,megalurous cercaria,parapleurolophocercous cercaria,and xiphidiocercariae.This is particularly true of xiphidiocercariae,which was found to be the dominant type among cercarial infections in bithyniid snails by approximately 38.00%.With regard to molecular identification,the phylogenetic tree was reconstructed using the neighbor-joining method with 10000 bootstraps and separated the trematodes into three clades:Echinostomatoidea,Microphalloidea and Opisthorchioidea.Conclusions:The study reveals a high prevalence of cercarial infection for each cercarial type and maturation into a definite trematode genus and delineates morphological characteristics and evolutionary trends among each larval trematode in Nakhon Nayok,Nonthaburi and Pathum Thani provinces.In addition,the ITS2 sequence data of cercariae could be used to examine classification of these species at the family level.

Investigation of cryptic diversity and occurrence of echinostome metacercariae infection in Anentome helena (von dem Busch, 1847)

Objective: To investigate the parasitic infection of Anentome helena(A. helena) and determine the validity of species boundaries for A. helena by combining molecular phylogeny and morphological approaches. Methods: A total of 325 individuals of A. helena were collected throughout northern Thailand. Shells were measured and compared by t-test. Radulae were investigated by using light and scanning electron microscope. Two partial mitochondrial DNA sequences of COI and 16S rRNA from 36 specimens of A. helena and related species were used to test the validity of the morphospecies. Phylogenetic trees were constructed using neighbour joining, maximum likelihood and Bayesian inference. Infection of A. helena with trematode larva was examined and observed. Results: Morphological examination of A. helena revealed 2 distinct morphospecies. Genetic divergences supported the separation of the two morphotypes into two distinct groups. Both individual and combined analyses of the two nucleotide fragments revealed two phylogroups that corresponded with shell and radula characteristics. In addition, A. helena was found infected with 37-collar spined echinostome metacercariae. The prevalence and intensity of metacercariae was highest in the San Kamphaeng district, Chiang Mai province, with 7.5% and 1.670±0.577, at comprehensive taxonomic respectively. Conclusions: These findings suggest thision of this unrecognised species complex is needed. This study represents the preliminary step to reveal new data on the recent distribution of trematode infection in A. helena. This information may be useful for developing conservation management of the snail and the practice of targeted regimes to reduce anthelmintic resistance in the future.