In-vivo efficacy of toltrazuril on experimentally induced Toxoplasma gondii tissue cysts in lambs: A novel strategy for prevention of human exposure to meat-borne toxoplasmosis

The aim of the present study was to investigate in vivo efficacy of toltrazuril on Toxoplasma gondii tissue cysts following induction of chronic toxoplasmosis in 4-week-old lambs (n=27) by inoculation of 1×105T. gondii ME 49 strain oocysts (day 0). Beginning at the 15th day after inoculation, lambs in Group T20 and Group T40 were given toltrazuril orally 2 times, once every week (Baycox 5%, Bayer Animal Health) at a dose of 20mg/kg and 40mg/kg, respectively. Positive control (PC) lambs were not given any therapy, and 2 clinically healthy non-infected lambs were used as negative controls (Group NC). Two out of 9 lambs in PC group (oocyst inoculated but non-treated) were killed on toltrazuril treatment days (day 15 and 22) to evaluate the tissue cyst presence in their muscles. On day 90, the remaining 25 lambs were necropsied, and samples from the brain and 11 different muscle groups were collected. The tissues were examined for the presence of tissue cysts by histopathology, immunohistochemistry, nested-PCR and percoll gradient centrifugation. Anti-T. gondii antibodies were screened by IFAT throughout the experiment. The increased T. gondii seropositivity beginning from the 15th day of inoculation remained steady at Day 45 and Day 90 in Groups PC while it was significantly lower at Day 90 in toltrazuril receiving groups. In toltrazuril treated groups, histopathological findings included degenerative changes in the cyst wall, complete macrophage invasion to the cysts, and reduction or removal of the cysts in toto. Four out of 9 lambs (44.4%) in both toltrazuril treated group (Group T20 and T40) did not contain tissue cyst in any examined tissues while all positive control animals had T. gondii tissue cysts at least in one muscle group. The toltrazuril treatment efficacy on the cyst presence was determined as 44.4%. The number of the cysts in the musculature was significantly different between non-treated and toltrazuril treated lambs (X2=6.613; p=0.037). For the total number of cysts, the positive control lambs had higher number of cysts compared to both toltrazuril treated lambs (T20 and T40) (X2=5.629; p=0.018 and X2=5.629; p=0.018, respectively) while there were no differences between Group T20 and Group T40 (X2=0.000; p=1.000). According to PCR results, the brain and M. semitendinosus were positive in all 7 control lambs while 12 out of 18 lambs were positive in toltrazuril treated lambs. In conclusion, the results are promising as the toltrazuril treate