Toxoplasma gondii is a major cause of reproductive losses in farm animals worldwide, with sheep being a particularly sensitive species. Yet, the diversity and biological characteristics of strains associated with ovine abortion remain incompletely understood in many regions, including South America whereby genetic diversity is known to be high. In this study, we investigated the genetic diversity of T. gondii associated with ovine abortion in Uruguay through genotyping and phylogenetic analyses of samples obtained from abortion cases. To place the observed diversity in a broader epidemiological context, ovine data were compared with previously reported genotypes circulating in humans locally. In addition, two novel strains were isolated from independent ovine abortion cases and subjected to determination of their infectivity both in vitro and in vivo, as well as to whole-genome sequencing. Integrated analyses revealed that the isolates represent previously undescribed genotypes and display marked differences in virulence-associated traits. Genomic analysis identified variation in loci previously linked to parasite virulence, including differences in ROP5 copy number. Together, our results provide new insight into the diversity of T. gondii associated with ovine abortion in Uruguay and expand current knowledge of the biological and genomic features of strains circulating in this species.