Predominance of the Blastocystis subtype ST5 among free-living sympatric rodents within pig farms in China suggests a novel transmission route from farms

Blastocystis is a parasitic protist that can infect humans and various domestic and wild animals. However, there is limited research on the prevalence of this parasite among rodents, particularly those living in pig farm settings. Therefore, to investigate the occurrence, molecular characterization, and zoonotic potential of Blastocystis among rodents within pig farm environments, we conducted an investigation of 227 rodents and shrews from 34 pig farms located in Henan, Shaanxi, and Shanxi provinces of China using nested PCR of the SSU rRNA gene of Blastocystis. The potential transmission and public health implications were also assessed from a One Health perspective. Blastocystis was detected in 86 (37.9%) fecal samples. The highest infection rate was observed among Ruttus norvegicus (73.7%, 42/58), followed by Ruttus tanezumi (30.1%, 41/136), and Mus musculus (12.0%, 3/25). However, it was not detected among individuals with Apodemus agrarius (n = 1) and Crocidura shantungensis (n = 7). Five known zoonotic Blastocystis subtypes (ST1-ST5) were identified, with ST4 (51.2%, 44/ 86) and ST5 (40.7%, 35/86) being the predominant ones, followed by ST1 (3.5%, 3/86), ST3 (3.5%, 3/86), and ST2 (1.2%, 1/86). ST4 was prevalent among R. norvegicus (83.3%, 35/42), while ST5 dominated R. tanezumi (70.7%, 29/41). Furthermore, ST5 exhibited the widest distribution at pig farm level, accounting for 65.0% (13/ 20) of Blastocystis-positive pig farms. This investigation presents the first documented Blastocystis infection in R. tanezumi and M. musculus, highlighting the predominant presence of the zoonotic ST5 subtype in rodents for the first time. The results demonstrate that sympatric rodents can serve as natural reservoirs for Blastocystis and play a role in its transmission. These findings provide information on the dynamics of rodent transmission and emphasize the potential public health threat posed by zoonotic Blastocystis subtypes spillover from pig farms.