Toxoplasma gondii infection in goats in Pakistan: risk factors and public health significance

Background Toxoplasma gondii is an intracellular protozoan parasite infecting warm-blooded animals, including humans. Infection can be associated with fetal mortality and a spectrum of other symptoms. Infection in humans mainly occurs through ingesting undercooked or raw animal products (meat and potentially milk) from infected animals, harbouring cysts or tachyzoites, or water or raw fresh produce contaminated with oocysts shed in feline feces. The South Punjab, Pakistan hosts intensive caprine population for meat and milk production, where goats are bred in very poor conditions, with unknown abortion history and milk is consumed raw. This study aimed to investigate the seroprevalence, risk factors, molecular detection, and genotyping of T. gondii in goats from an unexplored study area (Layyah District). Material and methods The prevalence of T. gondii infection was estimated using a two-stage sample design. All caprine farms in the study area were stratified by size, and from these 110 were randomly selected. Twelve goats (>1-year-old) were selected from each farm and a total of 1320 serum samples were collected and tested by ELISA. A questionnaire on the conditions and management practices of each farm was administered to 110 farmers. For molecular study, from three farms, previously identified highly seropositive, milk samples were collected from 40 goats at three different lactation intervals: 15-30, 90, and >120 days after parturition (360 samples in total). Foetal brain and placenta tissue samples were also collected from three aborted goats. A 162 bp fragment of the 529-bp repeat element was used to detect Toxoplasma DNA by real-time PCR. Positive samples were further genotyped using nested-PCR targeting four genetic markers (GRA6, SAG1, 5’- SAG2, CS3). Results Four hundred and sixteen out of 1320 sera samples (31.5%) were found positive and 89% of the flock had at least one seropositive goat. The proportion of seropositive goats tested within each flock ranged from 8.3% to 83.3%. with several factors contributing to this heterogeneity. Goat age played a significant role in the presence of cats. Significant interactions were related to goat farms having floor of dirt and kitten presence. Moreover, age class, abortion history and water source supply were modulated by owner education levels. T. gondii DNA was detected in 14 milk samples (3.9%), with higher detection in samples >120 days lactation stage (6.7%). Farm-wise detection was 5.8%, 2.5%, and 3.3%. All aborted tissues were positive, except one foetal brain tissue (83.3%). DNA isolates revealed type II in all samples, except one milk sample, and four aborted tissues, with either type I or III. Conclusions This is the first and most extensive study in Pakistan to report both serological and molecular prevalence of T. gondii in goats, including genotype analysis from milk. The findings highlight a significant public health risk and reproductive impact on goat herds, underscoring the need for improved farm hygiene, water management, and education-based control strategies.