Toxoplasma gondii infections are prevalent in humans and animals worldwide. In North America and Europe, T. gondii is highly clonal, consisting of three distinct lineages (Types I, II and III), whereas in South America, T. gondii is highly diverse with a few lineages expanded in the population. However, there is limited data on the diversity of T. gondii in Asia. Here we report the genetic characterization of T. gondii isolates from different hosts and geographical locations in China using the multilocus PCR-RFLP. A total of 17 T. gondii isolates from humans (3 strains), sheep (1 strain), pigs (5 strains) and cats (8 strains) were typed at 10 genetic markers including 9 nuclear loci SAG1, SAG2, SAG3, BTUB, GRA6, L358, PK1, c22-8, c29-2 and an apicoplast locus Apico. Four genotypes were revealed, including three previously reported and one new genotype. Three isolates belong to the clonal Type I lineage, one isolate belongs to the clonal Type II lineage, and the rest 13 isolates are grouped into two genotypes. This is the first report of genetic typing of T. gondii isolates from different hosts and geographical locations in China using a number of genetic markers, which has implications for the studies of population genetic structures of T. gondii, as well as for the prevention and control of T. gondii infections in humans and animals in China.
"Ten T. gondii strains collected from different hosts and locations were used for analysis in this study (Table 1). These strains have been genotyped and their genomic DNA has been prepared as described previously [15–17]. "
[Show abstract][Hide abstract] ABSTRACT: Genetic diversity of T. gondii is a concern of many studies, due to the biological and epidemiological diversity of this parasite. The present study examined sequence variation in rhoptry protein 17 (ROP17) gene among T. gondii isolates from different hosts and geographical regions. The rop17 gene was amplified and sequenced from 10 T. gondii strains, and phylogenetic relationship among these T. gondii strains was reconstructed using maximum parsimony (MP), neighbor-joining (NJ), and maximum likelihood (ML) analyses. The partial rop17 gene sequences were 1375 bp in length and A+T contents varied from 49.45% to 50.11% among all examined T. gondii strains. Sequence analysis identified 33 variable nucleotide positions (2.1%), 16 of which were identified as transitions. Phylogeny reconstruction based on rop17 gene data revealed two major clusters which could readily distinguish Type I and Type II strains. Analyses of sequence variations in nucleotides and amino acids among these strains revealed high ratio of nonsynonymous to synonymous polymorphisms (>1), indicating that rop17 shows signs of positive selection. This study demonstrated the existence of slightly high sequence variability in the rop17 gene sequences among T. gondii strains from different hosts and geographical regions, suggesting that rop17 gene may represent a new genetic marker for population genetic studies of T. gondii isolates.
"In this study, ToxoDB#3 (the type II variant) was identified for the first time in cats in China. This type had been founded from sheep in Qinghai province , from birds in Xinjiang Uygur Autonomous Region , from sparrows in Lanzhou, Gansu province , and from pigs in Zhongshan, Guangdong province . ToxoDB#1 (the type II) was reported in humans , but this is the first report of this genotype in cats. "
[Show abstract][Hide abstract] ABSTRACT: Cats are the definitive hosts of Toxoplasma gondii. The distribution of genetic diversity of T. gondii in cats is of importance to understand the transmission of this parasite. The objective of this study was to genetically characterize T. gondii isolates from cats in Yunnan province, southwestern China.
Genomic DNA was extracted from 5-10 g cat tissue samples (brain, tongue, heart, and liver). Using multilocous polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technology, we determined genetic diversity of T. gondii isolates from cats in Yunnan province.Result: In total, 175 stray cats were tested for T. gondii DNA, respectively, 44 (25.14%) of which were found to be positive for the T. gondii B1 gene by PCR amplification. The positive DNA samples were typed at 11 genetic markers, including 10 nuclear markers, namely, SAG1, 5[prime]-3[prime]SAG2, alternative SAG2, SAG3, GRA6, L358, PK1, BTUB, c22-8, c29-2 and an apicoplast locus Apico. Of these, 16 isolates from cats were genotyped with data for more than 9 loci, revealed 5 genotypes in total, of which 11 of 16 samples were identified as ToxoDB#9, two samples may belong to genotye #225, one was Type II, one was ToxoDB#3, and one was ToxoDB#20 (http://toxodb.org/toxo/).
The results of the present study indicated a wide distribution of T. gondii infection in cats in Yunnan province, which may pose significant public health concerns. To our knowledge, the present study is the first report of T. gondii prevalence and genotypes in cats in southwestern China, and the first report of Type II T. gondii from cats in China.
"However , T. gondii isolates from South America are diverse and largely distinct from those of North America and Europe (Ajzenberg et al. 2004; Khan et al. 2006; Lehmann et al. 2006; Pena et al. 2008). In Asia there are few reports from Japan (Smith and Frenkel 2003), Korea (Quan et al. 2008), Vietnam (Dubey et al. 2007b), Iran (Zia-Ali et al. 2007) and China (Dubey et al. 2007c; Zhou et al. 2009) indicating the existence of limited diversity among the isolates. There is a solitary report from India on a RFLP based genotyping targeting the SAG 2 locus of T. gondii isolates (Sreekumar et al. 2003). "
[Show abstract][Hide abstract] ABSTRACT: PCR-RFLP and nucleotide sequencing based genotyping of Toxoplasma gondii Indian isolates (Izatnagar and Chennai isolates and Chennai clone) vis-a vis RH-IVRI strain was conducted by targeting GRA6 as genetic marker. The 791 bp GRA6 product was PCR amplified from the genomic DNA of different T. gondii Indian isolates, including the RH-IVRI strain. Tru1I restriction endonuclease based PCR-RFLP of GRA6 sequence produced polymorphic digestion pattern that discriminated the virulent RH-IVRI strain (as type I) from the moderately virulent local isolates as type III. The PCR amplicon of T. gondii GRA6 from RH-IVRI strain as well as from the local isolates were cloned in cloning vector and custom sequenced. The nucleotide and deduced amino acid sequences of T. gondii isolates were aligned with that of the type I, II and III strains (RH, EVERLEY,
ME49, C56, TONT and NED) available in public domain and analyzed in silico using MEGA version 4.0 software. Nucleotide sequencing and phylogenetic analysis of GRA6 marker from the Indian isolates revealed a close genetic relationship with type III strains of T. gondii. Further, detection of a single nucleotide polymorphism (SNP) at positions 162 and 171 of the GRA6 marker, established the lineage of Indian isolates as type III. This is the first report on characterization of T. gondii lineage as type III in selective chicken population of India based on PCR-RFLP and sequence analysis of GRA6 gene.
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