Genetic Characterization of GRA6 Genes from Toxoplasma gondii from Pigs in Okinawa, Japan

Zakimi, Satoshi; Kyan, Hisako; Oshiro, Mamoru; Sugimoto, Chihiro; Xu, Xuan; Fujisaki, Kozo

doi:10.1292/jvms.68.1105

Cited by 27 publications

(16 citation statements)

References 15 publications

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“…Another GRA6 allele was found in type x, a genotype described in Californian sea otters (10,42,43). More recently, Zakimi et al (57) found nine new sequences among 29 isolates from Japanese pigs. Most studies with GRA6 as a genetic marker have been conducted by PCR-RFLP (8, 14-20, 24, 43, 57).…”

Section: Discussionmentioning

confidence: 94%

“…The coding region of the GRA6 locus has been used as a marker for the genotyping of Toxoplasma (8, 15-20, 24, 33, 34, 40, 43, 47, 53). Sequencing of GRA6 detected a high degree of polymorphism (24,57). Single nucleotide polymorphisms (SNP) in the region encoding GRA6 can be detected by methods based on PCR followed by restriction fragment length polymorphism (PCR-RFLP): digestion of the amplification products with a single endonuclease (MseI) can differentiate genotypes I, II, and III (24).…”

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confidence: 99%

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Selection of Polymorphic Peptides from GRA6 and GRA7 Sequences of Toxoplasma gondii Strains To Be Used in Serotyping

Sousa

Ajzenberg

Marle

et al. 2009

Clin Vaccine Immunol

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The evaluation of Toxoplasma gondii isolates obtained from geographical environments other than Europe and North America revealed the existence of atypical strains that are not included in the three archetypal clonal lineages (lineages I, II, and III). GRA6 and GRA7 are polymorphic genes that have been used for the genotyping of Toxoplasma. The coding regions of GRA6 and GRA7 from 49 nonarchetypal strains were sequenced and compared with the sequences of type I, II, and III reference strains. Eighteen and 10 different amino acid sequences were found for GRA6 and GRA7, respectively. The polymorphisms found between the different sequences were analyzed, with the objective of defining peptides to be used for the serotyping of Toxoplasma infections. Two peptides specific for clonal lineages I and III (peptides GRA7I and GRA7III, respectively) were selected from the GRA7 locus. Three peptides specific for some atypical strains (peptides Am6, Af6, and Am7) were selected from both the GRA6 and the GRA7 loci. Serum samples from humans infected with Toxoplasma strains of known genotypes were serotyped with the selected peptides. Peptide GRA7III seems to be a good candidate for the serotyping of infections caused by type III strains. Peptide GRA7I had a very low sensitivity. Peptides Am6 and Af6 had low specificities, since they reacted with serum samples from patients infected with strains belonging to the three archetypal lineages. Although peptide Am7 was specific, it had low sensitivity.The vast majority of Toxoplasma gondii isolates from human patients and domestic animals in Europe and North America belong to three archetypal clonal lineages, namely, types I, II, and III (1,11,30). However, nonarchetypal strains with atypical genotypes have recently been described in unusual hosts such as sea otters (10,42,43) and in tropical areas such as South America and Africa (2,32,39,46,54). Genotyping studies that distinguish different types of strains are important to gain knowledge of the biodiversity of the parasite in order to understand the molecular epidemiology of Toxoplasma and to highlight the correlation between the genotype of the parasite and the pathogenesis of human toxoplasmosis.The dense granules (GRA) are parasitic organelles involved in cell invasion and in the intracellular survival of the parasite. GRA proteins are expressed by the three stages of T. gondii: the tachyzoite, bradyzoite (38), and sporozoite (55) stages. GRA6 is a GRA antigen of 32 kDa described for the first time by Lecordier et al. (38). In extracellular parasites, GRA6 exists in dense secretor granules mostly as soluble proteins. Like the other GRA proteins, GRA6 is involved in host cell invasion. GRA6 is a glycine-rich protein and behaves like an integral membrane protein within the parasitophorous vacuole (36,41). GRA6 is considered a good marker of acute infection (27,28,52). However, the immune response to GRA6 is very heterogeneous (25).GRA7 is a GRA antigen of 29 kDa (26, 31). Like GRA6, it is involved in host cell invasion. This pro...

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Section: Discussionmentioning

confidence: 94%

mentioning

confidence: 99%

Selection of Polymorphic Peptides from GRA6 and GRA7 Sequences of Toxoplasma gondii Strains To Be Used in Serotyping

Sousa

Ajzenberg

Marle

et al. 2009

Clin Vaccine Immunol

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“…The highest number of strains originates from fattening pigs bred in outdoor farms. the world with types II and III predominant in Europe and North America (Mondragon et al 1998;Klun et al 2011;Dubey et al 2012;Turcekova et al 2013), and atypical strains more characteristic for South America and Asia (Zakimi et al 2006;Belfort-Neto et al 2007;Zhou et al 2010b;Bezerra et al 2012).…”

Section: Discussionmentioning

confidence: 99%

Toxoplasma gondii infection in pork produced in France

et al. 2016

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The aim of this study was to assess the seroprevalence of the Toxoplasma gondii parasite in pork produced in France, and to determine infection risk factors. An innovative survey was designed based on annual numbers of slaughtered pigs from intensive and outdoor farms in France. A total of 1549 samples of cardiac fluids were collected from pig hearts to determine seroprevalence using a Modified Agglutination Test. Of those, 160 hearts were bio-assayed in mice to isolate live parasites. The overall seroprevalence among fattening pigs was 2·9%. The adjusted seroprevalence in pigs from intensive farms was 3·0%; the highest in sows (13·4%); 2·9% in fattening pigs and 2·6% in piglets. Adjusted seroprevalence in fattening animals from outdoor farms was 6·3%. Strains were isolated from 41 animals and all were genotyped by Restriction Fragment Length Polymorphism as type II. Risk-factor analysis showed that the risk of infection was more than three times higher for outdoor pigs, and that sows' risk was almost five times higher than that of fattening animals. This study provides further evidence of extensive pork infection with T. gondii regardless of breeding systems, indicating that farm conditions are still insufficient to guarantee 'Toxoplasma-free pork'.

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“…They indicated that all isolates were type II by using 11 markers [ 20 ]. According PCR-RLFP of the GRA6 gene, the type I, II, and III of Toxoplasma were reported from pigs in Japan [ 34 ], the type II and III from humans, chicken, duck, and cat in Iran [ 23 ] and type II and III from humans in Cyprus [ 35 ].…”

Section: Discussionmentioning

confidence: 99%

Isolation and Genotyping of <i>Toxoplasma gondii</i> Strains in Ovine Aborted Fetuses in Khorasan Razavi Province, Iran

2016

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Toxoplasmosis is an important zoonotic disease that can cause abortion in humans and animals. The aim of this study was isolation and subsequent genotyping of Toxoplasma gondii isolates in ovine aborted fetuses. During 2012-2013, 39 ovine aborted fetuses were collected from sheep flocks in Khorasan Razavi Province, Iran. The brain samples were screened for detection of the parasite DNA by nested PCR. The positive brain samples were bioassayed in Webster Swiss mice. The serum samples of mice were examined for T. gondii antibodies by IFAT at 6 weeks post inoculation, and T. gondii cysts were searched in brain tissue samples of seropositive mice. The positive samples were genotyped by using a PCR-RLFP method. Subsequently, GRA6 sequences of isolates were analyzed using a phylogenetic method. The results revealed that T. gondii DNA was detected in 54% (20/37, 95% CI 38.4-69.0%) brain samples of ovine aborted fetuses. In bioassay of mice, only 2 samples were virulent and the mice were killed at 30 days post inoculation, while the others were non-virulent to mice. The size of cysts ranged 7-22 µm. Complete genotyping data for GRA6 locus were observed in 5 of the 20 samples. PCR-RLFP results and phylogenetic analysis revealed that all of the isolated samples were closely related to type I. For the first time, we could genotype and report T. gondii isolates from ovine aborted fetuses in Khorasan Razavi Province, Iran. The results indicate that the T. gondii isolates are genetically related to type I, although most of them were non-virulent for mice.

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Genetic Characterization of GRA6 Genes from Toxoplasma gondii from Pigs in Okinawa, Japan

Cited by 27 publications

References 15 publications

Selection of Polymorphic Peptides from GRA6 and GRA7 Sequences of Toxoplasma gondii Strains To Be Used in Serotyping

Selection of Polymorphic Peptides from GRA6 and GRA7 Sequences of Toxoplasma gondii Strains To Be Used in Serotyping

Toxoplasma gondii infection in pork produced in France

Isolation and Genotyping of <i>Toxoplasma gondii</i> Strains in Ovine Aborted Fetuses in Khorasan Razavi Province, Iran

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