Toxoplasma gondii, a zoonotic protozoan parasite, affects up to one-third of the global population. It can be transmitted through consumption of raw or undercooked meat, vertical transmission, or oocysts from contaminated water, soil, or food. However, there are no reports on the molecular prevalence of T. gondii in environmental sources like soil, water, and vegetables in India. This study aimed to detect and analyze T. gondii in samples from Chandigarh city, India. A total of 100 each soil & water and 500 vegetable samples were collected and analyzed using conventional PCR assay, real-time PCR assay targeting the glycerol-3-phosphate dehydrogenase (B1) gene and real-time LAMP assay targeting both B1 and Toxoplasma gondii outer wall protein (TgOWP) genes. Results showed that 15 of water, 9 of soil, and 6.4 of vegetable samples were positive for T. gondii. Real-time PCR assay and real-time LAMP assay detected slightly higher positivity rates in water and vegetable samples. Phylogenetic analysis revealed that the T. gondii isolates clustered with those from other regions such as Iran, India, Mexico, and those found in cats, pigs, and humans. This study is the first report of T. gondii contamination in environmental sources and fresh produce in India. The findings highlight the potential risk of human infection from contaminated water, soil, and vegetables in the region. © 2025 Elsevier B.V., All rights reserved.