Investigation Of Toxoplasma Gondii And Sarcocystis Sp. In Captive Cheetahs In Zoos

Toxoplasma gondii causes lifelong infection in both definitive hosts and intermediate hosts.Cats can excrete oocysts into the environment.Clinical cases of toxoplasmosis in captive cheetahs were reported.However,there are few reports of viable T.gondii strains isolated from cheetahs.Sarcocystis sp.has an obligatory prey-predator 2-host life cycle.Asexual stages develop only in the intermediate host,which,in nature,is often a prey animal.Sexual stages develop only in the definitive host.Intermediate and definitive hosts vary for each species of Sarcocystis sp.Generally,herbivorous animals are intermediate hosts,and carnivorous or omnivorous animals are definitive hosts.In this study,the cheetah tissue and serum samples were collected from 2017 to 2022,which were investigated for infection of T.gondii and Sarcocystis sp.,and to provide reference value for the prevention and control parasites infections in cats.We collected tissues and sera from seven cheetahs in zoos in Henan Province,T.gondii infection was investigated by serological,molecular assay and bioassay in cheetahs.100%(7/7)of the cheetahs showed T.gondii antibody by MAT(cut-off:1:25).T.gondii DNA was detected in 67%(4/6)of cheetahs.Two viable T.gondii strains were successfully isolated from the striated muscles of two cheetahs by mice bioassay.There were designated as TgCheetahCHnl and TgCheetahCHn2.T.gondii TgCheetahCHnl belonged to the genotype ToxoDB#319,alleles of ROP18/ROP5 were 3/4.TgCheetahCHn2 belonged to ToxoDB#9,alleles of ROP18/ROP5 were 3/5.TgCheetahCHnl was intermediate virulence,TgCheetahCHn2 was avirulent for Swiss mice,which matched the ROP18 and ROP5 gene alleles.Few brain tissue cysts(0-50)were observed in the mice after inoculation with TgCheetahCHnl or TgCheetahCHn2.Both of TgCheetahCHnl and TgCheetahCHn2 were avirulent for rabbits and piglets.Cheetahs in the zoos may potentially transmit T.gondii to other animals and humans.Tissue samples were collected from six cheetahs.The infection,morphology and ultrastructure of Sarcocystis sp.were studied by muscle squash,histological examination,electron microscopy and PCR.The results showed that the total positive rate of 67%(4/6).They were all thin-walled,ultrastructure confirmed that they were all Sarcocystis felis under light and electron microscopy and by 18S rRNA,ITS-1 and COX1.This is the first report of S.felis infection in a captive cheetah in China.Based on the phylogenetic tree analysis,it was suggested that carnivorous birds might be the definitive host of S.felis.In this study,the positive rate of T.gondii and Sarcocystis sp.infection in captive cheetahs in zoos was found to be high.The acquisition of cheetah-derived isolates with genotypes ToxoDB#319(novel genotype)and ToxoDB#9(Chinese1)enriches the genetic diversity and host diversity of T.gondii in China.It is the first time to report that cheetahs infected with S.felis in captivity in China,which provides direct evidence that the cheetah is an intermediate natural host of T.gondii and S.felis.