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Title:   Author  item
                                            Dubey, Jitenderitem
                                            WHY, KYLE - Animal And Plant Health Inspection Service (APHIS)item
                                            VERMA, SHIV - Non ARS Employeeitem
                                            CHOUDHARY, SHANTI - Non ARS Employeeitem
                                            Kwok, Oliveritem
                                            KHAN, ASSIS - Washington University School Of Medicineitem
                                            BEHINKE, MICHAEL - Washington University School Of Medicineitem
                                            SIBLEY, LAWRENCE - Washington University School Of Medicineitem
                                            FERREIRA, LEANDRA - Non ARS Employeeitem
                                            WEAVER, MELANIE - Pennsylvania Game Commission, Bureau Of Wildlife Managementitem
                                            STEWART, RICHARD - Shippensburg Universityitem
                                            SU, CHUNLEI - University Of Tennessee
  Submitted to: Veterinary Parasitology
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 1/11/2013
                                    Publication  Date: 1/22/2014
                                    Citation: Dubey, J.P., Why, K., Verma, S., Choudhary, S., Kwok, O.C., Khan, A., Behinke, M., Sibley, L., Ferreira, L., Weaver, M., Stewart, R., Su, C. 2014. Genotyping Toxoplasma gondii from wildlife in Pennsylvania and identification of natural recombinants virulent to mice. Veterinary Parasitology. 200(2014):74-84.Interpretive Summary: Toxoplasma gondii is a single-celled parasite of all warm-blooded hosts worldwide. It causes mental retardation and loss of vision in children, and abortion in livestock. Cats are the main reservoir of T. gondii because they are the only hosts that can excrete the resistant stage (oocyst) of the parasite in the feces. Humans become infected by eating under cooked meat from infected animals and food and water contaminated with oocysts. Why some people become sick whereas most remain asymptomatic is unknown. The genetic differences among isolates of Toxoplasma is thought to be one factor for pathogenicity. In the present study, authors found that mouse virulent strains of Toxoplasma are circulating in wildlife in the USA that are part of food chain. The results will be of interest to biologists, and Parasitologists.
                                Technical Abstract: Little is known of the genetic diversity of Toxoplasma gondii circulating in wildlife. Available data indicate a predominance of one T. gondii genotype (haplogroup 12) in wildlife in the USA but it is uncertain if this is due to sampling bias, range of hosts, or geographical restriction. In the present study, we tested coyotes (Canis latrans), red foxes (Vulpes vulpes), white-tailed deer (Odocoileus virginianus), and geese (Branta canadensis) from one state, Pennsylvania for T. gondii infection. Antibodies to T. gondii were found in 92 (34.5%) of 266 coyotes, 49 (62.0%) of 79 white-tailed deer 17 (85.0%) of 20 red fox, and two of two Canada geese tested by the modified agglutination test (cut off titer 1:25). Tissues from 105 seropositive animals were bioassayed in mice, and viable T. gondii was isolated from tissues of 10 of 53 coyotes, 11 of 16 foxes, seven of 49 deer, and one of one goose. All 29 isolates were further propagated in cell culture. DNA isolated from culture-derived tachyzoites of the 29 isolates was characterized initially using 11 PCR-RFLP markers (SAG1, 5’- and 3’-SAG2, alt.SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico). Nine genotypes were revealed, including ToxoDB PCR-RFLP #1 (4 isolates),  #2 (2 isolates), #3 (4 isolates), #4 (6 isolates), #5 (4 isolates), #54 (1 isolate), #141 (1 isolate), #143 (1 isolate), and #216 (6 isolates), indicating high genetic diversity of T. gondii in wildlife in Pennsylvania.  Pathogenicity of six T. gondii isolates (5 of #216 and #141) was determined in outbred Swiss Webster mice. Three of #216 and the #141 isolates were acutely virulent to mice, and the other 2 #216 isolates were intermediately virulent. To determine the extent of genetic variation of these as well as a few recently reported virulent isolates from wildlife in North America, intron sequences were generated. Analysis of intron sequences and PCR-RFLP genotyping results indicated that the #216 isolates are likely derived from recombination of the clonal type I and III lineages. To determine if T. gondii virulence can be predicted by typing, we genotyped a collection of strains using PCR-RFLP markers for polymorphic genes ROP5, ROP16, ROP18 and GRA15, which are known to interact with host immune response. The results showed that there is an association of genotypes of ROP5 and ROP18 with mouse-virulence, however, additional gene(s) may also contribute to virulence in distinct T. gondii genotypes.


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