Jian-Fa Yang

Seroprevalence of Toxoplasma gondii in horses and donkeys in Yunnan Province, Southwestern China

BackgroundToxoplasma gondii is an intracellular protozoan parasite that infects almost all warm-blooded animals, including humans, with a worldwide distribution. There have been limited reports about the seroprevalence of T. gondii infection in equids around the world and little is known about the seroprevalence of T. gondii in equids in southwestern China, in particular in Yunnan Province. The objective of the present investigation was to estimate the seroprevalence of T. gondii infection in equids in this area.MethodsA total of 399 serum samples (266 from horses and 133 from donkeys) were collected in 2012, and assayed for T. gondii antibodies by Indirect Haemagglutination (IHA) test using a commercially available kit.ResultsA total of 108 (27.1%) equids, including 81 (30.5%) horses and 27 (20.3%) donkeys were positive for T. gondii antibodies, and the seroprevalence ranged from 18.8% to 37.5% among different sampling areas. The seroprevalence was 27.4% and 26.8% for male and female equids, respectively, and the difference was not statistically significant (P > 0.05). The seroprevalence ranged from 21% to 32.9% among different age groups, and the difference was not statistically significant (P > 0.05).ConclusionsThe results of the present survey indicated the existence of high T. gondii seroprevalence in Yunnan Province, southwestern China, which has significant public health concern. Therefore, it is imperative that improved integrated measures be carried out to prevent and control T. gondii infection in equids in the studied region.

First report of Toxoplasma gondii seroprevalence in peafowls in Yunnan Province, Southwestern China

BackgroundToxoplasma gondii is an intracellular protozoan parasite infecting almost all warm-blooded animals, including birds, with a worldwide distribution. Surveys of T. gondii infection in wild birds have been reported extensively in the world, but little is known of T. gondii infection in peafowls worldwide. This study was performed to determine the seroprevalence of T. gondii infection in peafowls in Yunnan Province, southwestern China.MethodsSera from 277 peafowls, including 272 blue peafowls (Pavo cristatus) and 5 green peafowls (Pavo muticus) originated from two geographic areas in Yunnan Province were assayed for T. gondii antibodies using the modified agglutination test (MAT).ResultsSpecific T. gondii antibodies were detected in 35 of 277 (12.64%) peafowls (MAT titer ≥ 1:5). Seropositive birds were found in both species, 33 in 272 blue peafowls and 2 in 5 green peafowls. There was no significant difference in T. gondii seroprevalence between the adolescent birds (6.74%) and the adult birds (6.67%) (P > 0.05). The geographical origins of peafowls was found to be highly associated with T. gondii infection in the present study, a statistically significant difference in T. gondii seropositivity was observed between peafowls from Kunming (31.08%) and those from Xishuangbanna Dai Autonomous Prefecture (5.91%) (OR = 10.956, 95% CI = 1.632-73.545, P = 0.014). Statistical analyses showed that there were no significant interactions between ages and geographical origins of peafowls (P > 0.05).ConclusionsThe results of the present survey indicated that infection of peafowls with T. gondii is widespread in Yunnan Province, which has significant public health concerns and implications for prevention and control of toxoplamosis in this province. To our knowledge, this is the first seroprevalence report of T. gondii infection in China’s southwestern Yunnan Province.

Seroprevalence of Toxoplasma gondii infection in household and stray cats in Lanzhou, northwest China.

BackgroundToxoplasma gondii is an important protozoan parasite infecting humans and almost all warm-blooded animals. As the only definitive host, cats play a crucial role in the transmission of T. gondii infection by shedding parasite oocysts in their feces. However, little information on T. gondii infection in cats was available in Lanzhou, northwest China. This study was performed to determine the seroprevalence of T. gondii infection in household and stray cats in Lanzhou, northwest China.ResultsA total of 221 (179 households and 42 strays) blood samples were collected from clinically healthy cats admitted to several pet hospitals located in Lanzhou City, between November 2010 and July 2011 for the serological detection of T. gondii infection. The majority (207) of these cats represented Chinese Lihua cats. 47 of 221 (21.3%) examined cats were seropositive for T. gondii infection using the modified agglutination test (MAT) at the cut-off of 1:25. The seroprevalence in household and stray cats was assessed to be 15.6% and 45.2%, respectively, and the difference was statistically significant (P < 0.05). The seroprevalence ranged from 15.1% to 25.8% among different age groups, but the differences were not statistically significant (P > 0.05). Studies showed that there was no relationship between seroprevalence and the gender (P > 0.05).ConclusionsThe present survey indicated the high seroprevalence of T. gondii in cats in Lanzhou, northwest China, which poses a threat to animal and human health. Therefore, measures should be taken to control and prevent toxoplasmosis of cats in this area.

Genetic characterization of Toxoplasma gondii in Yunnan black goats (Capra hircus) in southwest China by PCR-RFLP

BackgroundToxoplasma gondii is a protozoan parasite that infects almost all warm-blooded animals and human beings. Goats are one of the susceptible animals to T. gondii. However, little is known of genetic diversity of T. gondii in Yunnan black goats in China. The objective of this present study was to determine the genotypes of T. gondii isolates from black goats in Yunnan province, southwest China.MethodsGenomic DNA was extracted from liver (n = 403), lung (n = 403) and lymph nodes (n = 250) of Yunnan black goats and assayed for T. gondii infection by semi-nested PCR of B1 gene. Then, the positive DNA samples were typed at 10 genetic markers using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technology. These markers include 9 nuclear loci, namely, SAG1, SAG2 (5’-SAG2 and 3’-SAG2, alternative SAG2), SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and an apicoplast locus Apico.ResultsOut of 403 tested samples, 20 (4.96%) DNA samples were T. gondii positive by amplification of B1 gene. Among them, 2 isolates were genotyped at all loci, and 6 isolates were genotyped for 8 or more loci. In total, seven samples belong to ToxoDB PCR-RFLP genotype#10 (Type I), and one belongs to genotype ToxoDB #9.ConclusionsTo our knowledge, this is the first report of ToxoDB#9 and ToxoDB#10 T. gondii in Yunnan black goats in China. These results revealed a wide distribution of these T. gondii in Yunnan black goats in China, which has important implications for public health.

Variability in intron sequences of housekeeping and antigen-coding genes among Schistosoma japonicum isolates in mainland China.

The accurate characterization of Schistosoma japonicum has important implications for analyzing genetic variation and would provide basic data for disease control. Previous studies using proteins, coding sequences, and especially antigen-coding genes showed lower genetic variation among S. japonicum isolates from mainland China. Therefore, the present study focused on variations in intron sequences of housekeeping and antigen-coding genes, which may be more informative for genetic analysis. We compared sequence variation between introns of two housekeeping genes and two antigen-coding genes. All 4 genes were polymorphic among all the S. japonicum isolates in mainland China, with 103, 158, 47, and 19 polymorphic (segregating) sites per kilobase in intron sequences of Actin, FBPA, 22.6kDa antigen and GST-26, respectively. Introns of housekeeping genes were slightly more polymorphic than coding and non-coding regions of antigen-coding genes examined in the present study within or among lake/marshland and mountainous types. Phylogenetic analysis based on sequences of single gene or combined sequences of multiple genes showed no specific clustering comprising parasites from single geographical or endemic regions. These results demonstrated that introns of housekeeping and antigen-coding genes were polymorphic, but the intron sequences examined in the present study were not suitable markers for examining genetic relationship among different isolates from endemic regions in mainland China.

Prevalences of Schistosoma japonicum infection in reservoir hosts in south-western China.

The human schistosomiases are major communicable parasitic diseases that, in the developing world, have public-health and socio–economic importance. Together, they affect approximately 200 million people worldwide, of whom approximately 20 million suffer from severe illness (Gryseels et al., 2006; Ribeiro-dos-Santos et al., 2006; Friedman et al., 2007). In mainland China, human schistosomiasis caused by Schistosoma japonicum was once a devastating disease that killed millions of people (Ross et al., 2001; Utzinger et al., 2005). Despite five decades of progress in the control and prevention of this problem in China, schistosomiasis is still regarded as the country’s most important parasitic zoonosis and one of the three most important communicable diseases (Guo and Yu, 2005; Zhou et al., 2005; Zhou et al., 2008). Of the three schistosome species that cause most cases of human schistosomiasis, S. japonicum is recognised as the most difficult to control because several common mammals serve as its ‘reservoir’ hosts (Utzinger et al., 2005). In China, while many wild mammals serve as natural hosts for S. japonicum, buffaloes and cattle are commonly infected and act as the main reservoir hosts for human infection (Li and Lin, 2007). As well as attracting large numbers of tourists, the province of Yunnan, in south– western China, holds more ethnic minorities than any other Chinese province. This area remains endemic for human schistosomiasis, despite the considerable progress made nationally in the control and prevention of the disease (Chen, 1999; Dong et al., 2008). Within the last decade, 15 of the 18 counties where the disease was once common in Yunnan met the national criteria for either transmission interruption (12 counties) or transmission control (Dali, Heqing and Nanjian), leaving only three counties in the province (Weishan, Eryuan and Yongsheng) where the disease remained poorly controlled. Unfortunately, over the last few years, schistosomiasis has re-emerged in some counties (such as Dali, Heqing and Nanjian) where control once appeared effective, probably because of increases in the mobility of the human populations and poor socio–economic conditions. The main aim of the present study was to evaluate the current problem posed by S. japonicum in all 18 counties of Yunnan that were formerly endemic for the parasite, by determining the prevalences of infection in the local livestock. The data collected should provide a foundation for the improved control of S. japonicum infection in the human populations and livestock of Yunnan.

Characterization of Fasciola Samples by ITS of rDNA Sequences Revealed the Existence of Fasciola hepatica and Fasciola gigantica in Yunnan Province, China

Abstract: On mainland China, liver flukes of Fasciola spp. (Digenea: Fasciolidae) can cause serious acute and chronic morbidity in numerous species of mammals such as sheep, goats, cattle, and humans. The objective of the present study was to examine the taxonomic identity of Fasciola species in Yunnan province by sequences of the first and second internal transcribed spacers (ITS-1 and ITS-2) of nuclear ribosomal DNA (rDNA). The ITS rDNA was amplified from 10 samples representing Fasciola species in cattle from 2 geographical locations in Yunnan Province, by polymerase chain reaction (PCR), and the products were sequenced directly. The lengths of the ITS-1 and ITS-2 sequences were 422 and 361–362 base pairs, respectively, for all samples sequenced. Using ITS sequences, 2 Fasciola species were revealed, namely Fasciola hepatica and Fasciola gigantica. This is the first demonstration of F. gigantica in cattle in Yunnan Province, China using a molecular approach; our findings have implications for studying the population genetic characterization of the Chinese Fasciola species and for the prevention and control of Fasciola spp. in this province.

Genetic diversity among Toxoplasma gondii isolates from different hosts and geographical locations revealed by analysis of ROP13 gene sequences

Toxoplasma gondii can infect almost all the warm-blooded animals and human beings, causing serious public health problems and economic losses worldwide. Rhoptry protein 13 (ROP13) plays some roles in the invasion process of T. gondii . In this study, sequence variation in ROP13 gene among 14 T. gondii isolates from different geographical locations and hosts was examined. The ROP13 gene was amplified from individual isolates and sequenced. Results show that the length of the ROP13 sequences was 1203 bp. In total, there were 44 variable nucleotide positions in the ROP13 sequences, and sequence variations were 0.1 to 2.0% among the 14 examined T. gondii isolates, representing higher rate in transversion than in transition. Intra-specific nucleotide variations were mainly at the second codon positions. Phylogenetic analysis of the 14 examined T. gondii isolates indicate that the ROP13 sequence was not a suitable genetic marker to differentiate T. gondii isolates of different genotypes from different hosts and geographical regions. Low variation in ROP13 gene sequence may suggest that ROP13 gene could represent a good vaccine candidate against toxoplasmosis. Key words: Toxoplasma gondii , toxoplasmosis, rhpotry protein 13 (ROP13), sequence variation, phylogenetic analysis.

Prevalence, genotypes, and risk factors of Enterocytozoon bieneusi in Asiatic black bear ( Ursus thibetanus ) in Yunnan Province, Southwestern China

Microsporidiosis is an important zoonotic disease, even leading to severe diarrhea. However, no information about prevalence and genotypes of Enterocytozoon bieneusi infection in Asiatic black bears in southwestern China is available. The objectives of the present study were to investigate the prevalence of E. bieneusi and to characterize their genotypes using the nested PCR amplification of the internal transcribed spacer region of the ribosomal RNA gene cluster and multilocus sequence typing (MLST). The overall prevalence of E. bieneusi was 19.75% (80/405) and the rate of E. bieneusi in Xishuangbanna (33.33%) was significantly higher than that in any other regions (Honghe, 17.65%; Dehong, 13.04%; Kunming, 0; P = 0.01). Sequence analysis revealed that 4 known genotypes (D, n = 2; SC02, n = 10; SC01, n = 5; and CHB1, n = 4) and 13 novel genotypes (designed MJ1–MJ13) were identified. When 17, 5, 14, and 34 sequences at loci MS1, MS3, MS4, and MS7 via MLST analyses, representing 4, 4, 5, and 10 genotypes, respectively, were completed, one multilocus genotype (MLG novel-ABB1) was identified. This is the first report of E. bieneusi in Asiatic black bear in Yunnan province, Southwestern China. The results indicated the potential zoonotic risk of this parasite through the Asiatic black bear in this region and provided foundation data for preventing and controlling E. bieneusi infection of many other animals and humans in these regions.

Prevalence and multi-locus genotypes of Enterocytozoon bieneusi in black-boned sheep and goats in Yunnan Province, southwestern China

Enterocytozoon bieneusi is a common pathogen in sheep and goats worldwide, threatening husbandry development and public health. However, there were few reports on native breeds of sheep and goats in China. In the present study, we investigated the prevalence and multi-locus genotypes (MLGs) of E. bieneusi in two important local breeds, black-boned sheep and goats, in Yunnan province, southwestern China. Of 661 faecal specimens collected from five counties of Yunnan province, 70 (10.59%) were positive for E. bieneusi infection. Infections were found in both black-boned sheep and goats, but no significant difference in prevalence was detected between two species. E. bieneusi was found in all age groups of black-boned sheep, while no infections were observed in 0-2-month black-boned goats. 12 genotypes were identified based on sequence variations of the internal transcribed spacer, including 10 known genotypes (BEB6, COS I, SX1, CM21, CHG3, CHG1, CHS5, EbpC, COS II, PigEb4) and two novel genotypes (named as YNS1 and YSM1), with BEB6 as the prevalent genotype both in black-boned sheep and goats. Phylogenetic analysis indicated that three genotypes of them, namely EbpC, CHS5 and PigEb4, were clustered into the zoonotic group 1, while the remaining nine genotypes were belonged to the host-adapted group 2. Multi-locus sequence typing (MLST) revealed 12 distinct multilocus genotypes (MLGs), with 11 MLGs detected in black-boned sheep and only one found in one black-boned goat. These findings expanded the reservoirs of E. bieneusi and provided fundamental data for controlling E. bieneusi infection in native small ruminants as well as other hosts in this province.