OBJECTIVETo identify the egg antigens related to the formation of hepatic granulomas and fibrosis of Schistosomiasis japonica.

METHODSThe egg cDNA library of Schistosoma japonicum (S. japonicum) was constructed and screened by immunological methods with the pooled sera of advanced schistosomiasis patients. The inserted foreign DNA fragments of positive clones were sequenced. The sequence data were analyzed using Wdnasis 2.5 and compared with Genebank data using blast software.

RESULTSEighty-one clones containing recombinant DNA fragments were obtained from the egg cDNA library of S. japonicum by immunological screening. The DNA sequences of all clones belonged to the miracidial antigen family. The longest cDNA fragment was 1604 bp, which contained an open reading frame of 351 bp, which encoded a protein of 1 2913.35 daltons.

CONCLUSIONThe cDNA sequence of the miracidial antigen of S. japonicum (Chinese strain) was obtained for the first time.

Animals ; Antigens, Helminth ; genetics ; Base Sequence ; China ; DNA, Complementary ; analysis ; Ovum ; Schistosoma japonicum ; genetics ; immunology

Mitochondrial genomes have been extensively studied for phylogenetic purposes and to investigate intra- and interspecific genetic variations. In recent years, numerous groups have undertaken sequencing of platyhelminth mitochondrial genomes. Haplorchis taichui (family Heterophyidae) is a trematode that infects humans and animals mainly in Asia, including the Mekong River basin. We sequenced and determined the organization of the complete mitochondrial genome of H. taichui. The mitochondrial genome is 15,130 bp long, containing 12 protein-coding genes, 2 ribosomal RNAs (rRNAs, a small and a large subunit), and 22 transfer RNAs (tRNAs). Like other trematodes, it does not encode the atp8 gene. All genes are transcribed from the same strand. The ATG initiation codon is used for 9 protein-coding genes, and GTG for the remaining 3 (nad1, nad4, and nad5). The mitochondrial genome of H. taichui has a single long non-coding region between trnE and trnG. H. taichui has evolved as being more closely related to Opisthorchiidae than other trematode groups with maximal support in the phylogenetic analysis. Our results could provide a resource for the comparative mitochondrial genome analysis of trematodes, and may yield genetic markers for molecular epidemiological investigations into intestinal flukes.
Animals ; Asia ; Codon, Initiator ; DNA, Mitochondrial/chemistry/genetics ; Gene Order ; Genes, Helminth ; *Genome, Mitochondrial ; Heterophyidae/*genetics/isolation & purification ; Humans ; Molecular Sequence Data ; Sequence Analysis, DNA

The present study was performed to describe 2 human cases infected by the horsehair worm, Parachordodes sp., in Japan. Two gordiid worms were collected in the vomit and excreta of an 80-year-old woman in November 2009 in Kyoto city, and in the mouth of 1-year-old boy in December 2009 in Nara city, Japan, respectively. Both worms were males having bifurcated posterior ends and male gonads in cross sectional specimens. They were identified as Parachordodes sp. (Nematomorpha: Chordodidae) based on the characteristic morphologies of cross sections and areoles in the cuticle. DNA analysis on 18S rRNA partial sequence arrangements was also carried out and both worms were assumed to be close to the genus Paragordionus based on tree analysis, and far from Gordius sp. which has already been reported in humans in Japan. DNA sequencing of the Parachordodes worm does not appear on the database; therefore, more information on the gene sequences of the genus Parachordodes from humans, animals, or intermediates is required.
Aged, 80 and over ; Animals ; Cluster Analysis ; DNA, Helminth/chemistry/genetics ; DNA, Ribosomal/chemistry/genetics ; Female ; Helminthiasis/*diagnosis/*parasitology/pathology ; Helminths/anatomy & histology/classification/genetics/*isolation & purification ; Humans ; Infant ; Japan ; Male ; Microscopy ; Phylogeny ; RNA, Ribosomal, 18S/genetics ; Sequence Analysis, DNA

To gain information on retrotransposons in the genome of Paragonimus westermani, PCR was carried out with degenerate primers, specific to protease and reverse transcriptase (rt) genes of long-terminal-repeat (LTR) retrotransposons. The PCR products were cloned and sequenced, after which 12 different retrotransposon-related sequences were isolated from the trematode genome. These showed various degrees of identity to the polyprotein of divergent retrotransposon families. A phylogenetic analysis demonstrated that these sequences could be classified into three different families of LTR retrotransposons, namely, Xena, Bel, and Gypsy families. Of these, two mRNA transcripts were detected by reverse transcriptase-PCR, showing that these two elements preserved their mobile activities. The genomic distributions of these two sequences were found to be highly repetitive. These results suggest that there are diverse retrotransposons including the ancient Xena family in the genome of P. westermani, which may have been involved in the evolution of the host genome.
Amino Acid Sequence ; Animals ; Cloning, Molecular ; DNA, Helminth/analysis ; *Evolution, Molecular ; *Genome ; Molecular Sequence Data ; Paragonimus/*genetics ; Phylogeny ; RNA-Directed DNA Polymerase/chemistry/genetics ; Retroelements/*genetics ; Sequence Alignment ; Sequence Analysis, DNA ; Terminal Repeat Sequences/*genetics

The evolutionary course of the CsRn1 long-terminal-repeat (LTR) retrotransposon was predicted by conducting a phylogenetic analysis with its paralog LTR sequences. Based on the clustering patterns in the phylogenetic tree, multiple CsRn1 copies could be grouped into four subsets, which were shown to have different integration times. Their differential sequence divergences and heterogeneous integration patterns strongly suggested that these subsets appeared sequentially in the genome of C. sinensis. Members of recently expanding subset showed the lowest level of divergence in their LTR and reverse transcriptase gene sequences. They were also shown to be highly polymorphic among individual genomes of the trematode. The CsRn1 element exhibited a preference for repetitive, agenic chromosomal regions in terms of selecting integration targets. Our results suggested that CsRn1 might induce a considerable degree of intergenomic variation and, thereby, have influenced the evolution of the C. sinensis genome.
Animals ; Clonorchis sinensis/*genetics ; DNA, Helminth/analysis/genetics ; *Evolution, Molecular ; Gene Dosage ; *Genome ; Phylogeny ; Polymorphism, Genetic ; RNA-Directed DNA Polymerase ; Retroelements/*genetics ; Sequence Analysis, DNA ; Terminal Repeat Sequences/*genetics

Fascioliasis, a food-borne trematode zoonosis, is a disease primarily in cattle and sheep and occasionally in humans. Water dropwort (Oenanthe javanica), an aquatic perennial herb, is a common second intermediate host of Fasciola, and the fresh stems and leaves are widely used as a seasoning in the Korean diet. However, no information regarding Fasciola species contamination in water dropwort is available. Here, we collected 500 samples of water dropwort in 3 areas in Korea during February and March 2015, and the water dropwort contamination of Fasciola species was monitored by DNA sequencing analysis of the Fasciola hepatica and Fasciola gigantica specific mitochondrial cytochrome c oxidase subunit 1 (cox1) and nuclear ribosomal internal transcribed spacer 2 (ITS-2). Among the 500 samples assessed, the presence of F. hepatica cox1 and 1TS-2 markers were detected in 2 samples, and F. hepatica contamination was confirmed by sequencing analysis. The nucleotide sequences of cox1 PCR products from the 2 F. hepatica-contaminated samples were 96.5% identical to the F. hepatica cox1 sequences in GenBank, whereas F. gigantica cox1 sequences were 46.8% similar with the sequence detected from the cox1 positive samples. However, F. gigantica cox1 and ITS-2 markers were not detected by PCR in the 500 samples of water dropwort. Collectively, in this survey of the water dropwort contamination with Fasciola species, very low prevalence of F. hepatica contamination was detected in the samples.
Animals ; Base Sequence ; Cluster Analysis ; DNA, Helminth/chemistry/genetics ; DNA, Ribosomal Spacer/chemistry/*genetics ; Electron Transport Complex IV/*genetics ; Fasciola hepatica/*genetics/*isolation & purification ; Korea ; Molecular Sequence Data ; Oenanthe/*parasitology ; Phylogeny ; Sequence Analysis, DNA ; Sequence Homology, Nucleic Acid

Diphyllobothrium nihonkaiense was first described by Yamane in 1986 but the taxonomical features have been obscure due to lack of critical morphologic criteria in its larval and adult stages. In Korea, this tapeworm had long been known as Diphyllobothrium latum. In this study, we observed 62 specimens collected from Korean residents and analyzed them by morphological features and nucleotide sequences of mitochondrial cox1 gene as well as the ITS1 region. Adult tapeworms were examined after carmine or trichrome stain. Longitudinal sections of the gravid proglottids showed an obtuse angle of about 150 degree between the cirrus sac and seminal vesicle. This angle is known as a major differential point compared with that of D. latum. Nucleotide sequence differences between D. latum and the specimens from Koreans represented 17.3% in mitochondrial DNA cox1 gene. Sequence divergence of ITS1 among 4 Korean isolates was 0.3% and similarity was 99.7% with D. nihonkaiense and D. klebanovskii. All of the Korean specimens analyzed in this study were identified as being D. nihonkaiense (n = 62). We propose its Korean name as "Dong-hae-gin-chon-chung" which means 'long tapeworm of the East Sea' for this newly analyzed diphyllobothriid tapeworm in Korea.
Adult ; Aged ; Animal Structures/anatomy & histology ; Animals ; Cluster Analysis ; Cyclooxygenase 1/genetics ; DNA, Helminth/chemistry/genetics ; DNA, Intergenic/chemistry/genetics ; DNA, Ribosomal/chemistry/genetics ; Diphyllobothriasis/parasitology ; Diphyllobothrium/*anatomy & histology/classification/*genetics/isolation & purification ; Female ; Helminth Proteins/genetics ; Humans ; Korea ; Male ; Microscopy ; Microscopy, Electron, Scanning ; Mitochondrial Proteins/genetics ; Phylogeny ; Sequence Analysis, DNA ; Sequence Homology

Hydatid cyst caused by Echinococcus granulosus is one of the most important parasitic diseases around the world and many countries in Asia, including Iran, are involved with this infection. This disease can cause high mortality in humans as well as economic losses in livestock. To date, several molecular methods have been used to determine the genetic diversity of E. granulosus. So far, identification of E. granulosus using real-time PCR fluorescence-based quantitative assays has not been studied worldwide, also in Iran. Therefore, the aim of this study was to investigate the genetic diversity of E. granulosus from center of Iran using real-time PCR method. A total of 71 hydatid cysts were collected from infected sheep, goat, and cattle slaughtered in Isfahan, Iran during 2013. DNA was extracted from protoscolices and/or germinal layers from each individual cyst and used as template to amplify the mitochondrial cytochrome c oxidase subunit 1 gene (cox1) (420 bp). Five cattle isolates out of 71 isolates were sterile and excluded from further investigation. Overall, of 66 isolates, partial sequences of the cox1 gene of E. granulosus indicated the presence of genotypes G1 in 49 isolates (74.2%), G3 in 15 isolates (22.7%), and G6 in 2 isolates (3.0%) in infected intermediate hosts. Sixteen sequences of G1 genotype had microgenetic variants, and they were compared to the original sequence of cox1. However, isolates identified as G3 and G6 genotypes were completely consistent with original sequences. G1 genotype in livestock was the dominant genotype in Isfahan region, Iran.
Animals ; Cattle ; Cluster Analysis ; DNA, Helminth/chemistry/genetics ; Echinococcosis/parasitology/*veterinary ; Echinococcus granulosus/*classification/*genetics/isolation & purification ; Electron Transport Complex IV/genetics ; *Genetic Variation ; Genotype ; Goats ; Iran ; Phylogeny ; Real-Time Polymerase Chain Reaction ; Sequence Analysis, DNA ; Sheep

The phylogenetic relationships of the 3 Neodiplostomum spp. (Digenea: Neodiplostomidae) occurring in Korea (N. seoulense, N. leei, and N. boryongense) were analyzed using the partial mitochondrial cytochrome c oxidase subunit 1 (CO1) gene. The adult flukes were recovered from Sprague-Dawley rats (N. seoulense) and newborn chicks (N. leei and N. boryongense) experimentally infected with the neodiplostomula from the grass snake, Rhabdophis tigrinus tigrinus. The genomic DNA was amplified using specific primers, and the sequence of CO1 was obtained. According to the results, the pairwise similarity was 96.1% between N. boryongense and N. seoulense, but was 95.0% between N. boryongense and N. leei and 94.2% between N. leei and N. seoulense. The results demonstrated a closer phylogenetic relationship between N. seoulense and N. boryongense. This high relationship of N. seoulense and N. boryongense may be related to their similar morphologic features including the limited distribution of vitellaria and the presence of a genital cone. N. leei is distinct on the other hand with an extensive distribution of vitellaria and the absence of a genital cone.
Animals ; Base Sequence ; Chickens ; Cluster Analysis ; Colubridae/*parasitology ; DNA, Helminth/chemistry/genetics ; Electron Transport Complex IV/*genetics ; Female ; Korea ; Molecular Sequence Data ; Phylogeny ; Rats, Sprague-Dawley ; Sequence Analysis, DNA ; Sequence Homology, Nucleic Acid ; Trematoda/*classification/*genetics

Fasciola hepatica is a trematode that causes zoonosis mainly in cattle and sheep and occasionally in humans. Fascioliasis has been reported in Korea; however, determining F. hepatica infection in snails has not been done recently. Thus, using PCR, we evaluated the prevalence of F. hepatica infection in snails at 4 large water-dropwort fields. Among 349 examined snails, F. hepatica-specific internal transcribed space 1 (ITS-1) and/or ITS-2 markers were detected in 12 snails and confirmed using sequence analysis. Morphologically, 213 of 349 collected snails were dextral shelled, which is the same aperture as the lymnaeid snail, the vectorial host for F. hepatica. Among the 12 F. hepatica-infected snails, 6 were known first intermediate hosts in Korea (Lymnaea viridis and L. ollula) and the remaining 6 (Lymnaea sp.) were potentially a new first intermediate host in Korea. It has been shown that the overall prevalence of the snails contaminated with F. hepatica in water-dropwort fields was 3.4%; however, the prevalence varied among the fields. This is the first study to estimate the prevalence of F. hepatica infection using the vectorial capacity of the snails in Korea.
Animals ; Base Sequence ; DNA, Helminth/chemistry/genetics ; DNA, Ribosomal Spacer/chemistry/genetics ; Fasciola hepatica/anatomy & histology/genetics/*isolation & purification ; Molecular Sequence Data ; Oenanthe/growth & development ; *Polymerase Chain Reaction ; Republic of Korea ; Sequence Analysis, DNA ; Snails/growth & development/*parasitology