Histologically, immunohistochemically, ultrastructurally, and molecularly confirmed neosporosis abortion in an aborted equine fetus.

Neosporosis is a common cause of abortion in cattle worldwide but is rare in horses. Here, the first case of histologically, ultrastructurally, immunohistochemically, and molecularly confirmed equine abortion caused by neosporosis is reported. Samples of lung, heart, liver, skeletal muscle, tongue, brain, and the placenta from a female fetus aborted at 280 days of gestation were fixed in formalin and submitted for diagnosis. Histologically, there was disseminated neosporosis with severe lesions in lungs, liver and the heart. Protozoal tachyzoites in all tissues reacted with polyclonal anti-Neospora caninum rabbit antibodies. Transmission electron microscopic observation on lung tissue revealed tachyzoites consistent with Neospora, including many rhoptries. Polymerase-chain reaction (PCR) using primers designed to amplify the rRNA gene internal transcribed spacer 1 (ITS1) of the Sarcocystidae was performed on DNA extracted from fetal tissues. Comparison of the ITS1 amplified from the foal tissue to sequences available in GenBank revealed 100% sequence identity to the ITS1 from three isolates of Neospora hughesi.

Epidemiological Investigation of Toxoplasma gondii Infections in Commercial Sheep Flock in an Endemic Area for Ocular Toxoplasmosis in Southern Brazil.

INTRODUCTION: Toxoplasma gondii is a widely distributed parasite and of great importance to human and animal health. METHODS: The objective of this study was to assess the prevalence of T. gondii antibodies and risk factors associated with the infection in sheep in the Northwest region of the State of Rio Grande do Sul, Brazil; this region has a very high rate of human ocular toxoplasmosis. Ovine sera were tested by the modified agglutination test (cut-off 1:25). RESULTS: T. gondii antibodies were detected in 70.2% (224 of 319). According to the logistic regression, the most significant factors associated were age and cat access to food stock facility. CONCLUSION: Preventive measures are discussed to reduce the risk of transmission of this zoonosis.

Isolation and Genetic Characterization of Toxoplasma gondii from Tissues of Wild Turkeys (Meleagris gallopavo) in Pennsylvania.

Toxoplasmosis in wild turkeys (Meleagris gallopavo) is of epidemiological interest because turkeys feed from the ground, and detection of infection in turkeys indicates contamination by oocysts in the environment. During the 2018 spring hunting season in Pennsylvania, fresh (unfixed, not frozen) samples were obtained from 20 harvested wild turkeys and tested for Toxoplasma gondii infection. Hearts from all wild turkeys and skeletal muscle from 1 were bioassayed for T. gondii by inoculation in outbred Swiss Webster (SW) and interferon-gamma gene knockout (KO) mice. Antibodies to T. gondii were detected in 1:5 dilution of neat serum from 5 of 15 wild turkeys and in fluid from the heart of 1 of 4 wild turkeys with the modified agglutination test (MAT); neat serum was not available from 4 wild turkeys. Viable T. gondii was isolated from hearts of 5 wild turkeys, 1 with MAT of 1:10, 1 with MAT of 1:5, and 3 seronegative (MAT < 1:5). Toxoplasma gondii was isolated from both heart and skeletal muscle in the 1 wild turkey that had skeletal muscle submitted. The KO mice inoculated with tissue from all 5 infected wild turkeys died or were euthanatized when ill, 7-21 days post-inoculation (PI). Tachyzoites were detected in lungs of all KO mice, and the T. gondii strains were successfully propagated in cell culture. The SW mice inoculated with tissues of wild turkeys remained asymptomatic, and tissue cysts were seen in the brains of infected mice when euthanatized in good health at 46 days PI; 1 of the 2 SW mice inoculated with the heart of 1 turkey died on day 26, and tachyzoites were detected in its lung. Genetic typing on DNA extracted from culture-derived tachyzoites using the PCR restriction fragment length polymorphism with 10 genetic markers (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico) revealed that 4 isolates belonged to ToxoDB PCR-RFLP genotype #5 and 1 was genotype #216.

Characterization of the zoonotic potential of Toxoplasma gondii in horses from Rio de Janeiro State.

The aim of this study was to perform a survey on the prevalence of anti-Toxoplasma gondii antibodies in horses from Rio de Janeiro State, Brazil. From 2012 to 2013, a total of 624 blood samples were collected from horses from the eight regions comprising Rio de Janeiro State (Baixadas Litorâneas, Serrana, Norte Fluminense, Noroeste Fluminense, Centro-Sul, Metropolitana, Médio Paraíba, and Costa Verde). All sera samples were tested for anti-T. gondii antibodies by performing the modified agglutination test with a cut-off of 1:25. Positive serology for T. gondii was detected in 22.8% (142/624) of the horses studied. Seropositivity was detected in all regions sampled; furthermore, statistical significance was observed when all locations were compared at once. The Médio Paraíba region had the highest number of positive animals 54.76% (23/42) in the Bonferroni correction among Costa Verde, Centro Sul, Metropolitana, and Serrana (p<0.001). Seropositivity was 39.58% (38/96) in Norte Fluminense, which was the second most prevalent region. The results indicated that the T. gondii parasite is widely distributed in horses in Rio de Janeiro State and represents a risk to public and animal health. These findings emphasize the need to increase control and prevention of this disease in horses.

Toxoplasma gondii transmission by artificial insemination in sheep with experimentally contaminated frozen semen.

Toxoplasma gondii is a parasite considered one of the major causes of reproductive problems in sheep. Furthermore, the presence of the agent in ram semen urges the possibility of sexual transmission in this species. The aim of this study was to evaluate if ram's frozen semen spiked with T. gondii tachyzoites would be able to cause infection in sheep by laparoscopic artificial insemination (AI). Nine ewes tested seronegative to anti-T. gondii antibodies by the modified agglutination test (MAT) were superovulated and inseminated to collect embryos. Animals were divided into two groups: G1 (n = 5), ewes inseminated with semen containing 4 × 107 tachyzoites; and G2 (n = 4), ewes inseminated with tachyzoite-free semen (control group). To confirm infection, ewe's blood samples were collected on days -14, -7, 0, 7, 14, 21, 28, 35, 49 and 57 after AI for analysis by MAT and PCR. Tissue samples of these ewes were also collected for histopathology and immunohistochemistry (IHC). Seven days after AI, all ewes of group G1 had specific antibodies to T. gondii, while those of G2 were negative. Toxoplasma gondii DNA was detected in the blood of one ewe and parasites were observed in tissues of all five animals inseminated with contaminated semen, indicating that semen freezing protocol does not affect T. gondii transmission by artificial insemination in sheep.

Increased incidence of DNA amplification in follicular than in uterine and blood samples indicates possible tropism of Neospora caninum to the ovarian follicle.

This study evaluated the presence of Neospora caninum in ovarian follicle aspirates and uterine flushes obtained from N. caninum seropositive dairy cows. Ninety-two cows that aborted within the previous 90 days were sampled to determine the presence of antibodies against N. caninum. Thirteen seropositive cows were chosen for collection of blood leukocytes, uterine flushes (UF; n=12) and follicular aspirates (OPU; n=13). Samples were centrifuged and the cellular sediment from the follicular fluid, uterine flushes and blood leukocytes were used for DNA extraction and PCR. Follicular aspirates had the highest frequency of DNA amplification for N. caninum (p<0.05, 92.3%; 12/13). Whereas uterine (4/12) and blood leukocyte (5/13) samples had similar (p>0.05) rate of positive results. Nonetheless, there was no agreement between blood leukocytes and follicular samples taken from the same animal (Cohen Kappa=-0.16). Similarly, blood leukocytes and uterine results had moderate agreement between them (Cohen Kappa=0.47). This study indicates that N. caninum is present in the ovarian follicle and uterus of seropositive cows, suggesting a possible risk of neosporosis transmission between females during oocyte and embryo collection and transfer. Hence, precautions should be taken to minimize the risk of N. caninum transmission. Furthermore, the high incidence of positive results in follicle samples, that exceeded that of their paired blood leukocytes, suggests a possible tropism of N. caninum for the ovarian follicle.