Simple SummarySarcocystidae is a family of protozoa whose life cycles usually include two hosts: an intermediate or paratenic host, where protozoa form tissular cysts, and a definitive host, whose intestines usually become infected after consuming tissular cysts. In Chile, studies of sarcocystids have been performed mostly on domestic animals, and scarce data are available from wildlife. A total of 207 rodents, encompassing six native species, from 13 localities in Central and Southern Chile, were studied to assess the presence of Sarcocystidae. DNA was isolated from several tissues and phylogenetic analyses were performed. Blood smear examinations and histological studies of several organs were performed. Overall, three individuals of rodents were PCR-positive and three different genotypes were retrieved belonging toSarcocystis,Besnoitia, and Toxoplasmatinae protozoa. No protozoa were found during microscopic examinations. Although it was not possible to confirm whether the findings corresponded to parasitism or accidental encounter, the phylogenetic positions of the genotypes support the hypothesis that they are parasites of rodents. All three genotypes are suggested as potential new taxa in the Sarcocystidae family.AbstractIn Chile, studies of parasites from the family Sarcocystidae (Apicomplexa) have mostly been related to domestic animals. We aimed to assess the presence of Sarcocystidae taxa in cricetid rodents from Central and Southern Chile. We studied 207 rodents, encompassing six species, from 13 localities. We isolated DNA from tissue samples, amplified the Sarcocystidae 18S rRNA gene with polymerase chain reaction, and performed phylogenetic analyses using maximum likelihood and Bayesian inferences. In addition, we examined blood smears and performed histological studies in organs from Sarcocystidae DNA-positive animals. Three specimens were DNA-positive and three genotypes were retrieved and named:Sarcocystissp. P61, related toSarcocystis strixi, was detected in twoAbrothrix olivacea. Toxoplasmatinae gen. sp. P99 was retrieved from those same two specimens, and was related toToxoplasmaand other genera, although it branched independently.Besnoitiasp. R34 was detected in oneAbrothrix hirta, and was clustered with congeneric species associated with rodents. No protozoa were found during microscopic studies; thus, it was not possible to confirm parasitic interactions rather than accidental encounters. However, the close relatedness of the retrieved genotypes to parasites of rodents supports the hypothesis of host–parasite associations. All three genotypes are suggested as potential new taxa, including a putative new genus.