Isolation and Cryopreservation of Toxoplasma gondii Isolates from Cats and Chickens from Selected Households in the Thika Region, Kenya

Abstract

There is a shortage of information in Africa regarding the genotypic and phenotypic characteristics of the Toxoplasma gondii circulating in domestic cats (Felis catus) and the intermediate hosts such as chicken. The situation is compounded by a lack of collection of well-stored isolates. The present study was aimed at creating a cryobank of T. gondii bradyzoites, tachyzoites, and oocysts. The parasites were isolated from cats and chickens kept in households in the Thika region, Kenya. Eight (8) cat fecal samples positive for T. gondii oocysts and 38 chicken brain tissue cysts (bradyzoites) were obtained and used for propagation in mice before cryopreservation. For each sample from the cats, (two donors) BALB/c mice were infected orally with 1 x 104/ml of oocysts. From each chicken sample, (two donors) BALB/c mice were infected intraperitoneally with 20-30 tissue cysts. On the third day after infection, tachyzoites were harvested from the peritoneal cavity of one donor mouse. The other two infected mice were further monitored for eight weeks, euthanized and the brain tissue harvested for toxoplasma cysts which were purified and cryopreserved. From the mice infected with oocysts from the cats’ samples, 2 (25%) tachyzoites but a higher 8 (100%) isolation was obtained from brain tissue cysts. On the other hand, from chicken samples generated 18 (47.3%) tachyzoites and 38 (100%), tissue cysts were obtained. The isolated oocysts (from cats), tachyzoites, and tissue cysts (from mice) were cryopreserved using 15% glycerol as cryoprotectant and stored in liquid nitrogen (-196oC). After 6 months of cryopreservation, the viability of the isolates was tested using Trypan blue dye exclusion on a manual hemocytometer. Viability (99.5% - 96%) of the cryopreserved samples was maintained for the three toxoplasma stages and there was no significant change (p>0.05) in the viability of the parasites before and after cryopreservation. The cryobank will serve as a repository for subsequent studies on molecular and phenotypic characterization of T. gondii isolates from Kenya.