Epidemiological Survey and Phylogenetic Characterization of Cysticercus tenuicollis Isolated from Tibetan Pigs in Tibet, China.
ABSTRACT: Cysticercus tenuicollis, commonly known as "water bell," is a larva of Taenia hydatigena, which is the most significant parasite of pigs. However, until now very few information is available regarding the prevalence and genetic characterization of the Cysticercus tenuicollis in Tibetan pigs. Therefore, the aim of this study was to investigate the prevalence and phylogenetic analysis of Cysticercus tenuicollis in Tibetan pigs. For this purpose, the COX2 gene of Cysticercus tenuicollis was amplified and sequenced for the first time in Tibetan pigs. The overall prevalence of Cysticercus tenuicollis was 43.93% in Tibetan pigs, with further distribution of 42.86% in 2014 and 45.35% in 2015. In Tibetan male and female pigs, the prevalence of Cysticercus tenuicollis was 43.39% and 44.56%, respectively. The prevalence of Cysticercus tenuicollis in different growing stages (juveniles, subadults, and adults) varied from 30.20% to 63.79%. The phylogenetic analysis of the Cysticercus tenuicollis isolates showed very close resemblance to 16 reference strains, isolates from Gansu, Hunan, and Sichuan provinces of China. To the best of our knowledge, this is the first report on the prevalence and genetic characterization of Cysticercus tenuicollis derived from Tibetan pigs. The data of present study provides baseline information for controlling cysticerci infections in pigs in Tibetan Plateau, China.
Project description:Taenia multiceps and Taenia hydatigena are widely distributed tapeworms of canids. Due to a lack of genetic information on these two parasites in China, in this study we analyzed six coenurus cerebralis and two cysticercus tenuicollis cysts from goats or sheep in Inner Mongolia, northern China by amplifying three mitochondrial genes (cox1, nad4, and cytb). Two haplotypes were obtained at each locus for either of the two Taenia cestode species, with ten nucleotide sequences being novel. The degrees of genetic variations were 1.18%, 0.61% and 0.52% for coenurus cerebralis, and 0.24%, 0.46% and 0.35% for cysticercus tenuicollis at the cox1, nad4 and cytb loci, respectively. This is the first molecular description of animal-derived metacestodes of T. multiceps and T. hydatigena in Inner Mongolia, China. Novel nucleotide sequences might reflect endemic genetic characterization of the two cestodes. The present data are useful to explore the biological and epidemiological significance of intra-specific variations within both Taenia cestodes.
Project description:Introduction:Cysticercosis caused by the larval stage of Taenia hydatigena is economically the most important endemic parasitic disease in Iraq. Few data are available relating to the genetic divergence of this helminth. This study aimed to molecularly characterise Cysticercus tenuicollis isolates from sheep in Sulaymaniyah province, Iraq. Material and Methods:DNA extraction and amplification of specimens of C. tenuicollis from 46 sheep were conducted by PCR for the mitochondrial 12S rRNA gene. The 19 amplicons were subjected to purification and partial sequencing. Results:Five 12S rRNA nucleotide sequence haplotypes were found. The pairwise nucleotide difference between haplotypes of 12S rRNA gene ranged from 0.2% to 0.7%. Four out of the five haplotypes of C. tenuicollis contained one to two base mutations and were discovered in Iraq for the first time, and this may be a unique mutation globally which has not been recorded previously. Three newly recorded haplotypes contained only one single mutation, and the other one contained two mutations. Phylogenetic analysis showed that all isolated strains were closely related to Iranian sheep isolates. Conclusions:Four new strains of T. hydatigena were discovered for the first time in the study area.
Project description:Similar to other metazoan pathogens, cysticercus undergoes transcriptional and developmental regulation during its complex lifecycle and host interactions. DNA methylation as a mechanism to control these processes has, to date, been discounted in this parasite. Given the new availability of high-resolution methylation detection methods, here we show the first evidence for cytosine methylation in the Taenia solium genome. Transcriptional coregulation of novel DNA methyltransferase (Dnmt2), DNA Methyltransferase 3 (DNMT3) and methyl-CpG-binding domain proteins mirrors the detection of cytosine methylation abundance and implicates the presence of functional DNA methylation machinery. Here, using MethylC-seq, we present the first study to confirm the existence of DNA methylation in the cysticercus of Taenia solium genome. we conservatively estimate that 0.11% of genomic cytosines are methylcytosines, most(56%) of which occur in CpA dinucleotides.in non-expressed gene cohorts mCG hypermethlated in gene bodies. and DNA methylation level with a negtive correlation with expression, suggesting it has a positive role in gene transcription. We find that transposable elements are hypomethylated, but in contrast, ribosomal DNAs are densely methylated. This work contributes to our understanding of epigenetics in platyhelminth such as cysticercus, which is an important human parasite. raises the possibility that targeting DNA methylation processes may be a useful strategy in therapeutics of cysticercosis. Finally, our work constitutes the first integrated report, to our knowledge, of DNA methylation in a cysticercus, demonstrates a strategy for sequencing the epigenomes of had methylation not on CpG. provide a foundation for future studies exploring this key epigenetic modification in human cysticercus disease and development. Overall design: Whole genome bisulfite sequencing and RNA-seq of cysticercus
Project description:BACKGROUND:Taenia pisiformis is one of the most common intestinal parasites in canines, and leads to serious economic losses in the rabbit breeding industry. Exosome-like vesicles from parasites play crucial roles in host-parasite interactions by transferring cargo from parasites to host cells and by modulating host immunological response through inducing production of host-derived cytokines. Nevertheless, the mechanism by which exosome-like vesicles from T. pisiformis cysticercus regulate the macrophage immune response remains unknown. METHODS:Using ultracentrifugation, we isolated exosome-like vesicles from excretory/secretory products (ESP) of T. pisiformis cysticercus. The morphology and size of purified vesicles were confirmed by transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). The components of proteins and miRNAs within these vesicles were identified by proteomic analysis and high-throughput small RNA sequencing. The biological function of targets of exosomal miRNAs was predicted by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Moreover, the expression of Th1- and Th2-type immune response associated cytokines in RAW264.7 macrophages were evaluated by qPCR and ELISA. We found that exosome-like vesicles were typical cup-shaped vesicles with diameters from 30 to 150 nm. A total of 87 proteins were identified by proteomic analysis, including proteins prominently associated with exosome-like vesicles biogenesis and vesicle trafficking. 41 known miRNAs and 18 novel miRNAs were identified in the exosome-like vesicles. Eleven selected miRNAs, including 7 known miRNAs (miR-71-5p, miR-10a-5p, miR-let-7-5p, miR-745-3p, miR-219-5p, miR-124-3p and miR-4989-3p) and 4 novel miRNAs (novel-mir-3, novel-mir-7, novel-mir-8 and novel-mir-11) were validated to exist in metacestiodes and exosome-like vesicles of T. pisiformis cysticercus by qPCR. The functions of most targets of exosomal miRNAs were mainly associated with signal transduction and the immune system. Additionally, T. pisiformis cysticercus-derived vesicles induced the production of IL-4, IL-6, IL-10, IL-13 and Arg-1, but downregulated the expression of IL-12, IFN-? and iNOS in RAW264.7 macrophages. CONCLUSIONS:We demonstrated that proteins and miRNAs enclosed within exosome-like vesicles from T. pisiformis cysticercus have immunomodulatory functions. Furthermore, exosome-like vesicles were shown to induce the macrophage Th2-type immune response in vitro. Our study suggests that exosome-like vesicles play an important role in the interaction between cysticerci and their hosts.
Project description:Taenia hydatigena is a non-zoonotic cestode that has canines as definitive hosts and ruminants and pigs as intermediate hosts. In pigs, its presence causes cross-reactivity in serological testing for Taenia solium cysticercosis. Therefore, knowledge on the occurrence of T. hydatigena is paramount for validly estimating the seroprevalence of T. solium cysticercosis in pigs. In a cross-sectional abattoir study, we estimated the prevalence of T. hydatigena in pigs slaughtered in Koudougou, Burkina Faso. Carcasses of 452 pigs were examined by investigators for perceived and suspected T. hydatigena cysticercus lesions in the abdominal cavity or on the surface of abdominal organs. Routine meat inspection was performed by local inspectors to identify T. solium cysticerci. All lesions were subjected to PCR-RFLP analysis in order to differentiate Taenia spp. Additionally, individual blood samples were examined for the presence of circulating cysticercus antigens using the B158/B60 Ag-ELISA. Perceived T. hydatigena cysticerci were found in 13 pigs, whereas meat inspectors found seven carcasses infected with T. solium cysticerci. All were confirmed by molecular analysis. Of pigs with other suspected lesions, mostly located in the liver, 27 and six were found to harbour T. hydatigena and T. solium cysticerci, respectively. Overall, 8.8% of pigs (40/452) were found infected with T. hydatigena and 2.9% (13/452) with T. solium. Of these positive pigs, one was found infected with both Taenia spp. (0.2%, 1/452). Blood samples of 48.5% of pigs (219/452) were positive in the Ag-ELISA. Pigs with confirmed cysts of T. hydatigena and T. solium had a positive Ag-ELISA result in 57.5% (23/40) and 61.5% (8/13) of cases, respectively. The observed T. hydatigena prevalence in this study is relatively high in comparison to other studies in Africa. Estimates of the occurrence of active porcine T. solium infection using the B158/B60 Ag-ELISA should therefore be adjusted for the presence of T. hydatigena. The low level of T. solium infection detected upon meat inspection in this study is likely an underestimation of the true prevalence since routine meat inspection shows poor sensitivity and pigs perceived to be infected based on tongue palpation are rarely sent to official abattoirs.
Project description:Tetraparvovirus, formerly known as Partetravirus, is a newly discovered genus in the family Parvoviridae that is considered phylogenetically distinct from other parvoviruses. However, nothing is known about the prevalence of Tetraparvovirus in special livestock living on the Qinghai-Tibet Plateau of China, such as Tibetan pigs and Tibetan sheep. A pair of special primers was designed based on the conserved regions in the genome of ungulate tetraparvovirus 2 (P-PARV4) and ungulate tetraparvovirus 4 (O-PARV4) and was used to detect P-PARV4 in domestic pigs and Tibetan pigs and O-PARV4 in ovines and Tibetan sheep. The results showed a 15.59 and 9.38% prevalence of P-PARV4 in domestic pigs (18.96% in Gansu Province and 11.76% in Qinghai Province) and Tibetan pigs (14.28% in Gansu Province and 4.44% in Qinghai Province), respectively, and a 7.31 and 5.20% prevalence of O-PARV4 in ovines (6.61% in Gansu Province and 8.00% in Qinghai Province) and Tibetan sheep (4.55% in Gansu Province and 5.50% in Qinghai Province), respectively. The prevalence of P-PARV4 was 14.76% (31/210) for ?1-month-old pigs and 10.58% (20/189) for >?1-month-old pigs, and the positive rates of O-PARV4 were 7.65% (18/235) for ?1-month-old sheep and 5.05% (11/218) for >?1-month-old sheep. The phylogenetic analysis of NS1, VP1, VP2 and the whole PARV4-related provirus genome demonstrated that both P-PARV4 and O-PARV4 sequences in this study were more closely related to the sequences of other strains discovered in the same genus of animals. The identity analyses for the full-length VP2 genomes of O-PARV4 revealed 98.84-100.00% sequence identity among the 7 strains and the previously reported strain, which was 98.60-99.28% for P-PARV4. In the present study, for the first time, we have provided comprehensive information regarding the widespread infection of P-PARV4 and O-PARV4 in special livestock on the Qinghai-Tibet Plateau in China. Our present findings highlight the importance of epidemiologic surveillance to limit the spread of Tetraparvovirus in livestock at high altitudes in China.
Project description:BACKGROUND:Cryptosporidium spp. are important zoonotic pathogens infecting a wide range of vertebrate hosts, and causing moderate to severe diarrhea in humans. Cryptosporidium infections are frequently reported in humans and animals worldwide, but little research has been done on local pig breeds such as Tibetan pigs and Yunan Black pigs and imported pig breeds such as Landrace pigs in China. Therefore, a total of 1089 pig fecal samples from four intensive farms in four areas of China, including Tibetan pigs from Gongbujiangda County (n = 180) and Mainling County (n = 434), Tibet, Yunan Black pigs from Sanmenxia, Henan Province (n = 246), and Landrace pigs from Kaifeng, Henan Province (n = 229), and were screened for the presence of Cryptosporidium with microscopy and nested PCR amplification of the small subunit rRNA gene. RESULTS:The total infection rate of Cryptosporidium in 1089 fecal samples of three different pig breeds was 2.11% (23/1089), and the infection rates of Tibetan pigs, Yunan Black pigs, and Landrace pigs were 0.49% (3/614), 0.41% (1/246), and 8.30% (19/229), respectively. The prevalence of Cryptosporidium infection was significantly higher in weaned piglets (1-2 months) (4.36%, 21/482) than in younger and older age groups (p < 0.01). Sequence analysis of positive samples revealed that there was no mixed infection in our study population, which included 12 cases of C. suis mono-infections (52.17%, 12/23) and 11 cases of C. scrofarum mono-infections (47.83%, 11/23). C. suis was identified in one pre-weaned piglet (< 1 month) and 11 weaned piglets (1-2 months), while C. scrofarum was only detected in 10 weaned piglets (1-2 months) and one finished pig (> 2 months). CONCLUSIONS:This is the first report on the identification of Cryptosporidium spp. in Tibetan pigs, and our findings also elucidate the occurrence and distribution of Cryptosporidium in three different pig breeds in Tibet and Henan, China. More molecular epidemiological studies are required to better clarify the prevalence and public health significance of Cryptosporidium in different pigs.
Project description:A specimen of intestinal glycoprotein isolated from the pig and two samples of dextran, all of which are polydisperse (that is, the preparations may be regarded as consisting of a continuous distribution of molecular weights), have been examined in the ultracentrifuge under meniscus-depletion conditions at equilibrium. They are compared with each other and with a glycoprotein from Cysticercus tenuicollis cyst fluid which is almost monodisperse. The quantity c(-(1/3)) (c=concentration) is plotted against xi (the reduced radius); this plot is linear when the molecular-weight distribution approximates to the ;most probable', i.e. when M(n):M(w):M(z): M((z+1))....... is as 1:2:3:4: etc. The use of this plot, and related procedures, to evaluate qualitatively and semi-quantitatively molecular-weight distribution functions where they can be realistically approximated to Schulz distributions is discussed. The theoretical basis is given in an Appendix.
Project description:Taenia solium infection causes severe neurological disease in humans. Even though infection and exposure to swine cysticercosis is scattered throughout endemic villages, location of the tapeworm only explains some of the nearby infections and is not related to location of seropositive pigs. Other players might be involved in cysticercosis transmission. In this study we hypothesize that pigs that carry nematodes specific to dung beetles are associated with cysticercosis infection and/or exposure. We carried out a cross-sectional study of six villages in an endemic region in northern Peru. We euthanized all pigs (326) in the villages and performed necropsies to diagnose cysticercosis. For each pig, we counted cysticerci; measured anti-cysticercus antibodies; identified intestinal nematodes; tabulated distance to nearest human tapeworm infection; and recorded age, sex, productive stage, and geographic reference. For the purpose of this paper, we defined cysticercosis infection as the presence of at least one cysticercus in pig muscles, and cysticercosis exposure as seropositivity to anti-cysticercus antibodies with the presence of 0-5 cysticerci. Compared to pigs without nematode infections, those pigs infected with the nematode Ascarops strongylina were significantly associated with the presence of cysticerci (OR: 4.30, 95%CI: 1.83-10.09). Similarly, pigs infected with the nematode Physocephalus sexalatus were more likely to have cysticercosis exposure (OR: 2.21, 95%CI: 1.50-3.28). In conclusion, our results suggest that there appears to be a strong positive association between the presence of nematodes and both cysticercosis infection and exposure in pigs. The role of dung beetles in cysticercosis dynamics should be further investigated.
Project description:Porcine deltacoronavirus (PDCoV) is a recently discovered RNA virus that belongs to the family Coronaviridae and genus Deltacoronavirus. This virus causes enteric disease in piglets that is characterized by enteritis and diarrhoea. In our present investigation, 189 diarrhoeic samples were collected between July 2016 and May 2017 from Tibetan pigs inhabiting in three different provinces surrounding the Qinghai-Tibet Plateau of China. We then applied the molecular-based method of reverse transcription polymerase chain reactions (RT-PCRs) to detect the presence of PDCoV in collected samples, and RT-PCR indicated that the prevalence of PDCoV was 3.70% (7/189) in Tibetan pigs. Four of 7 PDCoV-positive pigs were monoinfections of PDCoV, three samples were co-infections of PDCoV with porcine epidemic diarrhoea virus (PEDV), and 52 (27.51%) samples were positive for PEDV. Four strains with different full-length genomes were identified (CHN/GS/2016/1, CHN/GS/2016/2, CHN/GS-/2017/1 and CHN/QH/2017/1), and their genomes were used to analyse the characteristics of PDCoV currently prevalent in Tibetan pigs. We found a 3-nt insertion in the spike gene in four strains in Tibetan pigs. Phylogenetic analysis of the complete genome and spike and nucleocapsid gene sequences revealed that these strains shared ancestors with the strain CHN-AH-2004, which was found in pigs from the Anhui province of China mainland. However, PDCoV strains from Tibetan pigs formed different branches within the same cluster, implying continuous evolution in the field. Our present findings highlight the importance of epidemiologic surveillance to limit the spread of PDCoV in livestock at high altitudes in China.