Molecular Identification of Sarcocystis rileyi and Sarcocystis sp. (Closely Related to Sarcocystis wenzeli) in Intestines of Mustelids from Lithuania

SIMPLE SUMMARY: Protozoan parasites of the genus Sarcocystis are characterised by a two-host prey–predator life cycle. To date, more than 25 Sarcocystis species have been confirmed to form sarcocysts in muscles and CNS of birds. Avian Sarcocystis species are transmitted via predatory birds, placental mammals, and opossums. The objective of the study was to examine the role of predatory mammals of the family Mustelidae in the transmission of avian Sarcocystis spp. by means of molecular methods. In total, 115 small intestine samples of mustelids collected in Lithuania were tested for the presence of Sarcocystis spp. using anseriforms and domestic fowl (Gallus domesticus) as their intermediate hosts. Based on the DNA sequence analysis, S. rileyi known as forming macrocysts in muscles of ducks was detected in 11.3% of examined small intestine samples and Sarcocystis sp. was identified in two samples. The latter species was most closely related to Sarcocystis spp. isolates infecting chickens and causing encephalitis. This is the first report of avian Sarcocystis identified by molecular methods in the small intestines of mustelids, indicating the significance of these small predators for the spreading of Sarcocystis spp. using birds as intermediate hosts. Based on current knowledge, canids and mustelids are most likely the definitive hosts of S. rileyi in Europe. ABSTRACT: The genus Sarcocystis is a group of numerous protozoan parasites having a two-host life cycle. Based on laboratory experiments and/or phylogenetic analysis results it was shown that seven Sarcocystis spp. producing sarcocsyts in bird tissues are transmitted via predatory placental mammals. To date the role of small mammals of the family Mustelidae in the distribution of avian Sarcocystis spp. have not been studied. During the current investigation, intestinal mucosa scrapings of 115 mustelids belonging to five species were tested for S. albifronsi, S. anasi, S. rileyi, and S. wenzeli infecting anseriforms and chickens. Microscopically, free sporocysts, sporulating oocysts, and loose oocysts were found in 61 samples (53.0%). Using nested PCR targeting the ITS1 region and sequencing, S. rileyi was confirmed in eight American minks, two European polecats and single European badger. Sarcocystis sp. was identified in one American mink and one European pine marten. Based on the partial ITS1 region this parasite showed that 100% identity to pathogenic Sarcocystis sp. caused a fatal infection in backyard chickens from Brazil. Phylogenetically, the Sarcocystis sp. identified in our study was most closely related to S. wenzeli parasitising domestic fowl (Gallus domesticus).