Apoptosis ; Down-Regulation ; Electron Transport Complex IV ; chemistry ; genetics ; metabolism ; Humans ; Mitochondria ; metabolism ; Mutation ; Neoplasms ; genetics ; metabolism ; pathology

To determine the molecular phylogenic location of Plagiorchis muris, 28S D1 ribosomal DNA (rDNA) and mitochondrial cytochrome C oxidase subunit I (mtCOI) were sequenced and compared with other trematodes in the family Plagiorchiidae. The 28S D1 tree of P. muris was found to be closely related to those of P. elegans and other Plagiorchis species. And, the mtCOI tree also showed that P. muris is in a separate clade with genus Glypthelmins. These results support a phylogenic relationship between members of the Plagiorchiidae, as suggested by morphologic features.
Animals ; Base Sequence ; DNA, Helminth/chemistry/genetics ; DNA, Mitochondrial/chemistry/genetics ; Electron Transport Complex IV/chemistry/*genetics ; Molecular Sequence Data ; Phylogeny ; Polymerase Chain Reaction ; RNA, Ribosomal, 28S/chemistry/*genetics ; Sequence Alignment ; Support, Non-U.S. Gov't ; Trematoda/classification/*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

Mitochondrial cytochrome oxidase III (COIII) of duck was successfully amplified by PCR-mtDNA with duck muscle DNA as the template (GenBank Accession No. DQ655706). Cloning sequence analysis shows that the 784 bp nucleotides of COIII gene were contained. Through homology analysis, we confirmed that the cytochrome oxidase III (COIII) was relatively conservative. The method of PCR-mtDNA can be designed to detect the components of duck origin. And then, the method of PCR can be applied to amplify with the muscle DNA of various animal and feedstuff as the template, repeated verification, the primer (P3, P4) with strong specificity and good stability is screened, which can only amplify the sequence of duck. The special sequence contains 226 bp, the amplified product of 226 bp was sequenced and analyzed, it showed 100% homology with duck mtDNA COIII gene, which proved the accuracy of the special primer. The test that used different concentration of DNA with P3 and P4 is the sensitive experiment by PCR. The result showed that the primer has much specialty and rather sensitivity. So it is a way to detect the duck origin in the muscle of various animal and feedstuff.
Animal Feed ; analysis ; Animals ; DNA Primers ; genetics ; metabolism ; DNA, Mitochondrial ; genetics ; metabolism ; Ducks ; genetics ; Electron Transport Complex IV ; genetics ; Mitochondria, Muscle ; genetics ; Muscle, Skeletal ; chemistry ; Polymerase Chain Reaction ; methods ; Sensitivity and Specificity

Paragonimiasis is an important food-borne parasitic zoonosis caused by infection with lung flukes of the genus Paragonimus. Of the 7 members of the genus known in Thailand until recently, only P. heterotremus has been confirmed as causing human disease. An 8th species, P. pseudoheterotremus, has recently been proposed from Thailand, and has been found in humans. Molecular data place this species as a sister species to P. heterotremus, and it is likely that P. pseudoheterotremus is not specifically distinct from P. heterotremus. In this study, we collected metacercariae of both nominal species (identification based on metacercarial morphology) from freshwater crabs from Phetchabun Province in northern Thailand, Saraburi Province in central Thailand, and Surat Thani Province in southern Thailand. In addition, we purchased freshwater crabs imported from Myanmar at Myawaddy Province, western Thailand, close to the Myanmar-Thailand border. The DNAs extracted from excysted metacercariae were PCR-amplified and sequenced for ITS2 and cox1 genes. The ITS2 sequences were nearly identical among all samples (99-100%). Phylogenies inferred from all available partial cox1 sequences contained several clusters. Sequences from Indian P. heterotremus formed a sister group to sequences from P. pseudoheterotremus-type metacercariae. Sequences of P. heterotremus from Thailand, Vietnam, and China formed a separate distinct clade. One metacercaria from Phitsanulok Province was distinct from all others. There is clearly considerable genetic variation in the P. heterotremus complex in Thailand and the form referred to as P. pseudoheterotremus is widely distributed in Thailand and the Thai-Myanmar border region.
Animals ; Cluster Analysis ; DNA, Ribosomal Spacer/chemistry/genetics ; Electron Transport Complex IV/genetics ; *Genetic Variation ; Metacercariae/genetics/isolation & purification ; Molecular Sequence Data ; Myanmar ; Paragonimus/*classification/*genetics/isolation & purification ; Phylogeny ; Polymerase Chain Reaction ; Sequence Analysis, DNA ; Sequence Homology ; Shellfish/parasitology ; Thailand

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

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

DNA barcoding is a powerful approach for characterizing species of organisms, especially those with almost identical morphological features, thereby helping to to establish phylogenetic relationships and reveal evolutionary histories. In this study, we chose a 648-bp segment of the mitochondrial gene, cytochrome c oxidase subunit 1 (COI), as a standard barcode region to establish phylogenetic relationships among brine shrimp (Artemia) species from major habitats around the world and further focused on the biodiversity of Artemia species in China, especially in the Tibetan Plateau. Samples from five major salt lakes of the Tibetan Plateau located at altitudes over 4,000 m showed clear differences from other Artemia populations in China. We also observed two consistent amino acid changes, 153A/V and 183L/F, in the COI gene between the high and low altitude species in China. Moreover, indels in the COI sequence were identified in cyst and adult samples unique to the Co Qen population from the Tibetan Plateau, demonstrating the need for additional investigations of the mitochondrial genome among Tibetan Artemia populations.
Animals ; Artemia ; classification ; genetics ; Base Sequence ; China ; DNA, Mitochondrial ; chemistry ; genetics ; Electron Transport Complex IV ; genetics ; Genetic Variation ; Molecular Sequence Data ; Phylogeny ; Selection, Genetic ; Sequence Analysis, DNA ; Sequence Homology, Nucleic Acid ; Tibet

Among Paragonimus species, P. paishuihoensis is one of the most mysterious and poorly understood species. Metacercariae are characterized by having a unique dendritically branched excretory bladder. However, the morphology of the adult worm remains unknown. To date, metacercariae of this species have been reported only in China and Thailand. In this study, we first found P. paishuihoensis metacercariae in freshwater crabs, Potamon lipkei, in Hinheub District, Vientiane, Lao PDR, with a prevalence of 77.7% and the average intensity of 10.3 (range 1-28) metacercariae per crab. The molecular data based on ITS2 and CO1 markers indicated that P. paishuihoensis from Laos and Thailand were almost completely identical and were close to members of the Paragonimus bangkokensis/Paragonimus harinasutai complex. Attempts to infect experimental animals (cats, dogs, and rats) with P. paishuihoensis were unsuccessful, suggesting that these animals might be unsuitable definitive hosts for the species. Further studies are necessary to elucidate the taxonomic status and life cycle of P. paishuihoensis.
Animals ; Brachyura/*parasitology ; Cluster Analysis ; DNA, Ribosomal Spacer/chemistry/genetics ; Electron Transport Complex IV/genetics ; Fresh Water ; Laos ; Metacercariae/*isolation & purification ; Molecular Sequence Data ; Paragonimus/*isolation & purification ; Phylogeny ; Polymerase Chain Reaction ; Prevalence ; Sequence Analysis, DNA ; Sequence Homology

In order to provide a new method for the identification of Placenta hominis, the COI barcode has been employed to identify the P. hominis medicinal materials and its adulterants. Genomic DNA was extracted from the experimental samples. The COI sequences were amplified and sequenced bi-directionally. Sequence assembly and consensus sequence generation were performed using the CodonCode Aligner. NJ tree was constructed by MEGA6.0 software. COI sequences can be successfully obtained from all experimental samples. The intra-specific variation and inter-specific divergence were calculated. The average intra-specific K2P distance of P. hominis was 0.001 and the maximum intra-specific distance was 0.008. The cluster dendrogram constructed can be seen that the same genus is together, and distinguished from its adulterants. It is concluded that P. hominis and its adulterants can be correctly identified by DNA barcoding method.
Animals ; Cattle ; DNA Barcoding, Taxonomic ; methods ; Drug Contamination ; prevention & control ; Electron Transport Complex IV ; genetics ; Female ; Humans ; Medicine, Chinese Traditional ; Molecular Sequence Data ; Phylogeny ; Placenta ; chemistry ; enzymology ; Pregnancy ; Quality Control ; Sheep ; Swine