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Genetic diversity of Enterocytozoon bieneusi in 1099 wild animals and 273 imported pastured donkeys in northern China

Parasites & Vectors volume 18, Article number: 105 (2025) Cite this article

Abstract

Background

Enterocytozoon bieneusi is the most frequently detected microsporidian species in humans, wildlife and domestic animals. In northern China, to the best of our knowledge, no information on E. bieneusi infection has been reported in wild animals. The aim of the present study was to survey the occurrence of and genetically characterize E. bieneusi from a broad spectrum of vertebrate species in this region.

Methods

A total of 1372 small intestine or fecal specimens were collected from 1019 mammals, 121 reptiles and 232 birds in Xinjiang Uygur Autonomous Region (XUAR) and Inner Mongolia Autonomous Region (IMAR), northern China. Each animal species was identified according to morphological characteristics and amplification of mitochondrial genes. Genotype analysis of E. bieneusi was performed by amplifying the internal transcribed spacer (ITS) region.

Results

A total of 68 wild animal species were identified, including 34 mammal species, six reptile species and 28 bird species. The average rate of infection with E. bieneusi was 9.7% (133/1372 specimens). Twelve genotypes of E. bieneusi, including BEB6, CHG7, D, E, EbpD, horse1, MWC_d1, NCF2, NCF6, PL14, SN45 and XJHT4, were identified in specimens from XUAR, IMAR and Kyrgyzstan. Phylogenetically, these genotypes belonged to four groups, namely Group 1, Group 2, Group 12 and Group 14.

Conclusions

To our knowledge, this study reports for the first time E. bieneusi genotype NCF2 in marbled polecats (Vormela peregusna), genotype NCF6 in red foxes (Vulpes vulpes), genotype D in grey wolf (Canis lupus), genotypes CHG7, horse1 and PL14 in rodents and genotypes MWC_d1, PL14 and SN45 in wild birds. The results also indicate that genotypes horse1, NCF2 and NCF6 were acquired either by the fecal–oral transmission route or during predator–prey interaction.

Graphical Abstract

Background

Enterocytozoon bieneusi, a fungus-like protist parasite, is the most frequently detected microsporidian species causing microsporidiosis, accounting for > 90.0% of human microsporidiosis cases. The main clinical manifestation of E. bieneusi infection is diarrhea [1]. The life-cycle stages of E. bieneusi include schizonts, sporonts and spores. Infectious spores can usually be acquired from the environment by multiple host species, but some genotypes of E. bieneusi are host specific [2]. It has been isolated from a variety of host taxa, including humans, companion animals, livestock and wildlife. Since mature spores are shed in the host’s feces, the transmission routes of this pathogen may involve person-to-person contact as well as water-borne or food-borne infection, especially in developing countries [3]. Due to the variety of transmission routes, the sylvatic epidemiological cycle, which involves wild animal hosts such as carnivores and rodents, can pose a significant risk factor to the health of humans and domestic animals living nearby.

Enterocytozoon bieneusi genotypes are usually identified and classified by PCR and sequence analysis of the internal transcribed spacer region (ITS) of nuclear ribosomal DNA (rRNA) [4]. To date, approximately 600 distinct genotypes of E. bieneusi have been recorded in humans and approximately 170 distinct genotypes animal species. Among these, 49 genotypes have been found both in humans and animals [5].

In the southwestern and central regions of China, 361 E. bieneusi genotypes have been confirmed [6]. These genotypes were assigned to 14 major genetic groups, denoted as Group 1 to 14. Interestingly, genotypes in Groups 1 and 2 were found to be zoonotic, while genotypes in Groups 3 to 14 were restricted to specific hosts, as exemplified by genotype CD5 that is only detected in dogs [7, 8].

Xinjiang Uygur Autonomous Region (XUAR) and Inner Mongolia Autonomous Region (IMAR), located in northern China, cover 1,6749 and 1,1830 million km2, respectively [9]. The multiple natural landscapes (e.g. deserts, alpine meadows, forests and wetlands) and vast area of these regions provide suitable habitats for an abundance of terrestrial wildlife, including mammals and birds. These regions also have several endemic species, such as the Yarkand hare (Lepus yarkandensis). To evaluate the zoonotic potential of the isolates at the genotype level in XUAR and IMAR, the aim of this study was to explore the genotypes and phylogenetic groups of E. bieneusi detected in samples from 1084 wild animals, 15 zoo inhabitants and 273 pastured donkeys (Equus asinus) imported from Kyrgyzstan.

Methods

Animal fecal sample collection

During 2015–2024, intestinal samples from 273 pastured donkeys imported from Kyrgyzstan and part of the intestine or fecal samples from 1099 wild animals were collected in XUAR and IMAR, including 610 wild rodents, 81 Mongolian pikas (Ochotona pallasi), one Yarkand hare (Lepus yarkandensis), two common shrews (Sorex araneus), three Przewalski’s gazelles (Procapra przewalskii), 16 red foxes (Vulpes vulpes), eight marbled polecats (Vormela peregusna), six Asian badgers (Meles leucurus), three Eurasian lynxes (Lynx lynx), five long-eared hedgehogs (Hemiechinus auritus), 121 lizards, 228 birds, three sika deer (Cervus nippon), one Bactrian camel (Camelus bactrianus), one wolf (Canis lupus), two Serengeti lions (Panthera leo), one tiger (Panthera tigris), one brown bear (Ursus arctos), one Asiatic black bear (Ursus thibetanus), one rhesus monkeys (Macaca mulatta), three emus (Dromaius novaehollandiae) and one peafowl (Pavo cristatus)(Additional file 1: Table S1; Additional file 2: Table S2; Fig. 1). The species of sampled hosts were identified by key morphological characteristics [10, 11] and by genetic markers including the 910-bp 16S rRNA fragment for birds [12], the 924-bp displacement loop (D-loop) mitochondrial DNA (mtDNA) for bird feces [13], the 650-bp cytochrome c oxidase subunit I (COX1) for reptiles [14] and a 1242-bp cytochrome B (cytb) sequence for rodents (Additional file 3) [15].

Fig. 1
figure 1

Locations where samples were collected for this study

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DNA extraction

Genomic DNA from each intestinal sample was extracted using the TIANamp Genomic DNA Kit (TIANGEN, Beijing, China) following the manufacturer’s instructions. Genomic DNA from each fecal sample was extracted using the EasyPure Stool Genomic DNA Kit (TRANS, Beijing, China). The DNA quantity was assessed on a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA). Samples with a DNA concentration of at least 30 ng/μl could be used to detect pathogens.

Molecular detection of Enterocytozoon bieneusi

Enterocytozoon bieneusi was identified and genotyped using a nested PCR protocol with both forward and reverse primers for the sequence, targeting an approximately 390-bp nucleotide fragment in the ITS region [16]. The PCR products were purified using the TIANgel Midi Purification Kit (TIANGEN) and sequenced.

Nucleotide sequencing and analysis

The obtained sequences were edited and compared to sequences from GenBank using the BLASTN program (http://www.ncbi.nlm.nih.gov/BLAST/). Fifty sequences that served to identify host species were deposited in the GenBank database (16S rRNA: PQ394778-PQ394785, PQ451723-PQ451727 and PQ459365; COX1: PQ451734-PQ451736; D-loop mtDNA: PQ373942-PQ373943, PQ393132-PQ393138 and PQ474771; cytb: PQ450150-PQ450162, PQ450164-PQ450166, PQ474772-PQ474773, OR548119 and PQ581939). In addition, 23 sequences of E. bieneusi from this study were also deposited in GenBank, under accession numbers PP489440-PP489449, PP601260, PP959050-PP959059 and PQ489475-PQ489476.

Phylogenetic analyses

A neighboring-joining (NJ) phylogenetic tree was constructed for E. bieneusi using the maximum composite likelihood model with MEGA 7.0 software. Bootstrap values were obtained with 1000 replicates.

Statistical analyses

Prevalence data were compared by Fisher’s exact test, and differences were regarded significant when p < 0.05.

Results

Identification of wild animal species

In this study, 1372 samples were collected from 68 species of animals, including 34 mammalian, six reptilian and 28 avian species (Table 1).

Table 1 Prevalence and distribution of Enterocytozoon bieneusi genotypes in wild animals in northwest China
Full size table

Prevalence of E. bieneusi

We detected E. bieneusi in 9.7% (133/1372) of samples, including 11.9% (121/1019) of samples from mammals, 4.3% (10/232) of samples from birds and none of the samples (0/121) from reptiles (Table 1). Among the samples from mammals, the highest rate of infection was detected in carnivores (23.1%) and rodents (13.6%). Within the same genus of rodents, the prevalence was significantly higher in Meriones tamariscinus than in Meriones libycus, and in Spermophilus erythrogenys compared to Spermophilus undulatus (p < 0.0001) (Additional file 1: Table 1).

Genotypes of E. bieneusi

Based on the ITS region, a total of 12 genotypes of E. bieneusi were identified, including BEB6 (synonyms: CHS4, CHC9, CHHLJS1, CHS18, JSS1 and SH5), CHG7, D (synonyms: WL8, Peru9, PigEBITS9, PtEb VI and CEbC), E (synonyms: EbpC, WL13, Peru4 and WL17), EbpD, horse1, MWC_d1 (synonym: BJED-V), NCF2, NCF6, PL14, SN45 and XJHT4 (Fig. 2). All ITS sequences showed 98.0–100.0% similarity to the following GenBank reference sequences: MW429409 for genotype BEB6; KP262358 for genotype CHG7; MT895457 for genotype D; KJ700426 for genotype E; KJ728797 for genotype EbpD; MW429428 for genotype horse1; MK121776 for genotype MWC_d1; MG976814 for genotype NCF2; PQ165135 for genotype NCF6; MZ400637 for genotype PL14; MN378369 for genotype SN45; and ON165751 for genotype XJHT4.

Fig. 2
figure 2

Phylogenetic relationships of Enterocytozoon bieneusi genotypes identified in the present study and other known genotypes deposited in GenBank, inferred from ITS sequences analyzed with the neighboring-joining method using the maximum composite likelihood model, with 1000 replicates. Each sequence is shown according to its accession number, host origin and genotype designation. Rodents (black triangles), Ochotona pallasi (Mongolian pika; black inverted triangles), Equus asinus (wild asses; black squares), carnivores (black circles) and birds (black diamonds) are indicated

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Among these E. bieneusi genotypes, PL14 (31.6%, 42/133) was the most prevalent, followed by the CHG7 (24.8%, 33/133), horse1 (16.5%, 22/133), EbpD (9.8%, 13/133), XJHT4 (4.5%, 6/133), D (3.8%, 5/133), NCF2 (3.0%, 4/133), NCF6 (2.3%, 3/133), MWC_d1 (1.5%, 2/133), BEB6 (0.8%, 1/133), E (0.8%, 1/133) and SN45 genotypes (0.8%, 1/133). The distribution of these among different host species is shown in Table 1.

We observed a varied distribution pattern of the E. bieneusi genotypes among different animal species. Genotypes EbpD, PL14 and XJHT4 were only found in rodent species that were positive for the pathogen. Genotypes E, MWC_d1 and SN45 were only found in bird species, and genotypes NCF2 and NCF6 were only found in carnivores.

Phylogenetic relationships of E. bieneusi genotypes

Genotypes identified in this study represented four major phylogenetic groups, namely Groups 1, 2, 12 and 14. Genotypes CHG7, D, E, EbpD, horse1, MWC_d1, NCF2, NCF6 and SN45 clustered in Group 1. At the same time, genotypes BEB6, PL14 and XJHT4 belonged to Groups 2, 12 and 14, respectively (Fig. 2).

Discussion

In this study, most of our genotypes of E. bieneusi fall into Group 1, including genotypes CHG7, D, E, EbpD, horse1, MWC_d1, NCF2, NCF6 and SN45. Group 1 is the largest genotype group, comprising 314 genotypes among which genotypes D, E and Type IV are widely spread across host species and geographic ranges and, importantly, they are also frequently found in human beings. According to a previous multi-locus sequence typing (MLST) analysis, Group 1 is further divided into eight subgroups, from subgroup 1a to 1h [17].

Wildlife is a potential source of human infections of E. bieneusi in central and southern China. In this study, we detected E. bieneusi in 9.4% (103/1099) of the samples from wild animals. This prevalence is lower than reported than that reported other studies; for example, E. bieneusi prevalence in animals from Zhengzhou Zoo and Chengdu Zoo was reported to be the same, namely 15.8% (32/203) [18] and 15.8% (43/272) [19], respectively, while the prevalence of E. bieneusi from Shanghai Zoo and Zhejiang Zoo was 11.5% (21/182) [20]. The difference in E. bieneusi prevalence between our results and those of previous studies may be due to differences in geographical location, in habitats and/or in host species.

Rodents, especially wild-living species, can act as reservoirs for numerous pathogens, and some of these pathogens (as also exemplified by E. bieneusi) are zoonotic [6, 21]. There are over 2000 species of rodents worldwide. Previously, 60 genotypes of E. bieneusi, among them 18 zoonotic ones, have been confirmed in rodents in different parts of Eurasia, including China [2, 8, 17, 22,23,24,25,26]. In addition, genotype PL14, representing phylogenetic group 12, has been occasionally identified in farmed masked palm civet (Paguma larvata) in Yunnan Province, southwestern China [27]. In the present study, we identified genotype PL14 for the first time in rodents, including the red-cheeked ground squirrel (Spermophilus erythrogenys), Tamarisk gerbils (Meriones tamariscinus) and the great gerbil (Rhombomys opimus). On the other hand, genotype XJHT4 was previously reported in samples from gray marmots (Marmota baibacina) [28]. Our results indicate broadening of its host spectrum, with genotype XJHT4 also detected in the long-tailed ground squirrel (Spermophilus undulatus) and pygmy wood mouse (Apodemus uralensis). Genotype D was the most frequently detected variant in rodents in Poland and the border region of Czech Republic-Germany [21]. In contrast, PL14 was the most dominant genotype in rodents in the present study, with other pathogenic genotypes, including CHG7, D, EbpD, horse1 and XJHT4, also shown to be present in local rodents in northern China.

Pikas (Ochotonidae) are mainly distributed in Central Asia, but their range also includes northeastern Asia, western North America and Europe [29]. There are only 30 species of pikas, of which 24 species occur in China [30]. Enterocytozoon bieneusi genotypes CHN14 and CHS17 have been observed previously in plateau pikas (Ochotona curzoniae) [23]. In the present study, we found genotype BEB6, a member of phylogenetic Group 2, for the first time in Mongolian pikas (Ochotona pallasi) collected in Beitashan Mountain, XUAR. Interestingly, BEB6 was previously identified in Tibetan sheep and yaks [31]. Beitashan Mountain, covering 3848 km2 along the China-Mongolia border, has a fauna rich in wild animal species that inhabit alpine meadows, such as pikas, yaks and sand gazelles (Gazella subgutturosa). Therefore, in the future, the epidemiological significance of transmission routes involving these mammals should be further evaluated.

Genotype D of E. bieneusi is widely distributed in animal and human patients [1, 5, 22], and genotypes NCF2 and NCF6 may infect a broad range of carnivores, such as raccoons (Procyon lotor), red foxes (Vulpes vulpes), European badgers (Meles meles) [32] and arctic foxes (Vulpes lagopus) [32, 33]. In the present study study, several new host associations were revealed, including genotype NCF2 in marbled polecats (Vormela peregusna), genotypes NCF6 in red foxes (Vulpes vulpes) and genotype D in the gray wolf (Canis lupus). These findings extend the host ranges of genotypes D, NCF2 and NCF6. Interestingly, genotype NCF2 was detected in both marbled polecats and red foxes, and genotype D in rodents, red fox and gray wolf. These data suggest that red foxes play a pivotal role in pathogen spillover via the predator–prey relationship, and that E. bieneusi can be transmitted via the “rodent-red fox-gray wolf” or “marbled polecat-red fox” food chain (Additional file 4).

Enterocytozoon bieneusi genotype horse1 is widely found in equines, but genotype CHG7 was previously detected only in pigs and sheep [34, 35]. In the present study, genotypes horse1 and CHG7 were both detected in donkeys imported from Kyrgyzstan. Similarly, genotype SN45 was previously found in wild Himalayan marmots (Marmota himalayana) and Alashan ground squirrels (Spermophilus alashanicus) [37], while genotype MWC_d1 was only shown to be present in wild ungulate species [37, 38]. We report here for the first time the detection of genotypes MWC_d1 and SN45 in migratory birds [such as the greylag goose (Anser anser) and northern wheatear (Oenanthe oenanthe)]. There are three international flyways in XUAR: (i) the West Asia-Middle East-East Africa flyway; (ii) the Siberia-Central Asia-South Asia flyway; and (iii) the Arctic tundra-Asia-Australia flyway. Multiple genotypes of E. bieneusi were detected in imported donkeys and migratory birds in our study, and some of these were shared between these two host types, indicating that the host range of E. bieneusi genotypes is extended due to pathogen spillover, and that the risks of transmission of E. bieneusi across the borders are high in XUAR.

There is an earlier report of a pathogen spillover-related outbreak between migratory birds and resident birds, as in the case of genotype Peru6 transmission between red-crowned crane (Grus japonensis) and pigeons (Columba livia) [40]. In the present study, genotype CHG7 of E. bieneusi was detected in both wild-living resident birds (e.g. red-naped ibis) and migratory birds [e.g., Pallas’s gull (Larus ichthyaetus) off the coast of Ganjam and lesser black-backed gull (Larus fuscus)], indicating that some E. bieneusi genotypes circulate between resident birds and migratory birds in northwestern China.

No published reports are available on E. bieneusi infection in lizards. In this study, E. bieneusi was not detected in lizard samples, possibly due to pathogen specificity or rare infection in lizards. At the same time, E. bieneusi genotype horse1 was widely distributed among rodents (e.g. Himalayan marmots) and resident birds [e.g. long-eared owls (Asio otus), jungle nightjars (Caprimulgus indicus) and Hume’s groundpeckers (Pseudopodoces humilis)] in their habitat-overlap region, suggesting that genotype horse1 is transmitted via the fecal–oral route (Additional file 5).

Despite the significance of the findings of this study, there are some limitations because the species diversity in regional wildlife is only partly represented. Therefore, it is probably that numerous examples of predator–prey relationships that are important in the epidemiology of E. bieneusi remain to be explored.

Conclusions

A total of 1372 samples, collected from 34 mammalian, six reptilian and 28 avian species, were examined in this study. Twelve genotypes of E. bieneusi, including BEB6, CHG7, D, E, EbpD, horse1, MWC_d1, NCF2, NCF6, PL14, SN45 and XJHT4, were identified in the samples collected from XUAR, IMAR and Kyrgyzstan. These genotypes represent four phylogenetic groups, namely Group 1, Group 2, Group 12 and Group 14. These findings indicate that red foxes play a vital role in pathogen spillover via the predator–prey relationship; that the risk of cross-border transmission of E. bieneusi is high in XUAR owing to imported livestock and migratory birds; and that the genotype horse1 circulates between marmots and resident birds in habitat-overlap regions due to fecal–oral transmission.

Availability of data and materials

No datasets were generated or analysed during the current study.

Abbreviations

16S rRNA :

16S Ribosomal RNA

COX1 :

Cytochrome c oxidase subunit I

Cyt b :

Cytochrome B

D-loop:

Displacement loop region

References

  1. Santín M, Fayer R. Microsporidiosis: Enterocytozoon bieneusi in domesticated and wild animals. Res Vet Sci. 2011;90:363–71.

    Article  PubMed  Google Scholar 

  2. Wang S-S, Wang R-J, Fan X-C, Liu T-L, Zhang L-X, Zhao G-H. Prevalence and genotypes of Enterocytozoon bieneusi in China. Acta Trop. 2018;183:142–52.

    Article  CAS  PubMed  Google Scholar 

  3. Zhang K, Fu Y, Li J, Zhang L. Public health and ecological significance of rodents in Cryptosporidium infections. One Health. 2021;14:100364.

    Article  PubMed  PubMed Central  Google Scholar 

  4. Feng Y, Li N, Dearen T, Lobo ML, Matos O, Cama V, et al. Development of a multilocus sequence typing tool for high-resolution genotyping of Enterocytozoon bieneusi. Appl Environ Microbiol. 2011;77:4822–8.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Zhang Y, Koehler AV, Wang T, Gasser RB. Enterocytozoon bieneusi of animals—with an ‘Australian twist.’ Adv Parasitol. 2021;111:1–73.

    Article  PubMed  Google Scholar 

  6. Zhao W, Zhou H, Yang L, Ma T, Zhou J, Liu H, et al. Prevalence, genetic diversity and implications for public health of Enterocytozoon bieneusi in various rodents from Hainan Province China. Parasit Vectors. 2020;13:438.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Zhou H-H, Zheng X-L, Ma T-M, Qi M, Zhou J-G, Liu H-J, et al. Molecular detection of Enterocytozoon bieneusi in farm-raised pigs in Hainan Province, China: infection rates, genotype distributions, and zoonotic potential. Parasite. 2020;27:12.

    Article  PubMed  PubMed Central  Google Scholar 

  8. Li J, Jiang Y, Wang W, Chao L, Jia Y, Yuan Y, et al. Molecular identification and genotyping of Enterocytozoon bieneusi in experimental rats in China. Exp Parasitol. 2020;210:107850.

    Article  CAS  PubMed  Google Scholar 

  9. Liu G, Zhao S, Hornok S, Chen X, Wang S, Tan W, et al. Taenia laticollis and a potentially novel Taenia species from the Eurasian lynx (Lynx) in Northwestern China. Int J Parasitol Parasites Wildl. 2021;16:183–6.

    Article  PubMed  PubMed Central  Google Scholar 

  10. Prompiram P, Mongkolphan C, Poltep K, Chunchob S, Sontigun N, Chareonviriyaphap T. Baseline study of the morphological and genetic characteristics of Haemoproteus parasites in wild pigeons (Columba livia) from paddy fields in Thailand. Int J Parasitol Parasites Wildl. 2023;21:153–9.

    Article  PubMed  PubMed Central  Google Scholar 

  11. Pawar SD, Pande SA, Tare DS, Keng SS, Kode SS, Singh DK, et al. Morphological and biochemical characteristics of avian faecal droppings and their impact on survival of avian influenza virus. Food Environ Virol. 2018;10:99–106.

    Article  PubMed  Google Scholar 

  12. Liu D. Identification of a bird remain ina case destroying wildlife resources based on the COI and 16SrRNA Genes. Chin J Wildl. 2022;43:715–24.

    Google Scholar 

  13. Crochet P-A, Desmarais E. Slow rate of evolution in the mitochondrial control region of Gulls (Aves: Laridae). Mol Biol Evol. 2000;17:1797–806.

    Article  CAS  PubMed  Google Scholar 

  14. Che J, Chen H, Yang J, Jin J, Jiang K, Yuan Z, et al. Universal COI primers for DNA barcoding amphibians. Mol Ecol Resour. 2012;12:247–58.

    Article  CAS  PubMed  Google Scholar 

  15. Yu Z-M, Chen J-T, Qin J, Guo J-J, Li K, Xu Q-Y, et al. Identification and characterization of Jingmen tick virus in rodents from Xinjiang China. Infect Genet Evol. 2020;84:104411.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Buckholt MA, Lee JH, Tzipori S. Prevalence of Enterocytozoon bieneusi in swine: an 18-month survey at a slaughterhouse in Massachusetts. Appl Environ Microbiol. 2002;68:2595–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Thellier M, Breton J. Enterocytozoon bieneusi in human and animals, focus on laboratory identification and molecular epidemiology. Parasite. 2008;15:349–58.

    Article  CAS  PubMed  Google Scholar 

  18. Li J, Qi M, Chang Y, Wang R, Li T, Dong H, et al. Molecular characterization of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in captive Wildlife at Zhengzhou Zoo China. J Eukaryot Microbiol. 2015;62:833–9.

    Article  CAS  PubMed  Google Scholar 

  19. Li W, Deng L, Yu X, Zhong Z, Wang Q, Liu X, et al. Multilocus genotypes and broad host-range of Enterocytozoon bieneusi in captive wildlife at zoological gardens in China. Parasit Vectors. 2016;9:395.

    Article  PubMed  PubMed Central  Google Scholar 

  20. Zhang Y, Mi R, Yang L, Gong H, Xu C, Feng Y, et al. Wildlife Is a Potential Source of Human Infections of Enterocytozoon bieneusi and Giardia duodenalis in Southeastern China. Front Microbiol. 2021;12:692837.

    Article  PubMed  PubMed Central  Google Scholar 

  21. Perec-Matysiak A, Buńkowska-Gawlik K, Kváč M, Sak B, Hildebrand J, Leśniańska K. Diversity of Enterocytozoon bieneusi genotypes among small rodents in southwestern Poland. Vet Parasitol. 2015;214:242–6.

    Article  PubMed  Google Scholar 

  22. Wang H, Liu Q, Jiang X, Zhang Y, Zhao A, Cui Z, et al. Dominance of zoonotic genotype D of Enterocytozoon bieneusi in bamboo rats (Rhizomys sinensis). Infect Genet Evol. 2019;73:113–8.

    Article  PubMed  Google Scholar 

  23. Liu X, Du S, Yang X, Xia X, An Z, Qi M. First genotyping of Enterocytozoon bieneusi in plateau pikas (Ochotona curzoniae) from the Qinghai Plateau Northwest China. Vet Res Commun. 2021;45:453–7.

    Article  PubMed  Google Scholar 

  24. Sak B, Kváč M, Květoňová D, Albrecht T, Piálek J. The first report on natural Enterocytozoon bieneusi and Encephalitozoon spp. infections in wild East-European House Mice (Mus musculus musculus) and West-European House Mice (M. m. domesticus) in a hybrid zone across the Czech Republic-Germany border. Vet Parasitol. 2011;178:246–50.

    Article  PubMed  Google Scholar 

  25. Lv C, Li C, Wang J, Qian W. Prevalence and molecular characterization of Enterocytozoon bieneusi and a new Enterocytozoon sp. in pet hairless guinea pigs (Cavia porcellus) from China. Parasite. 2023;30:37.

    Article  PubMed  PubMed Central  Google Scholar 

  26. Lv C, Wang J, Li C, Zhang M, Qian W. First detection and genotyping of Enterocytozoon bieneusi in pet golden hamsters (Mesocricetus auratus) and Siberian hamsters (Phodopus sungorus) in China. Parasite. 2022;29:15.

    Article  PubMed  PubMed Central  Google Scholar 

  27. Lin X, Xin L, Cao Y, Hou M, Qiao F, Li J, et al. Common occurrence of Enterocytozoon bieneusi genotypes SHR1 and PL2 in farmed masked palm civet (Paguma larvata) in China. Int J Parasitol Parasites Wildl. 2021;16:99–102.

    Article  PubMed  PubMed Central  Google Scholar 

  28. Tuo H, Zhang B, He Y, Zhao A, Zhang Z, Qi M, et al. Molecular characterization of Enterocytozoon bieneusi genotypes in wild Altai marmot (Marmota baibacina) in Xinjiang, China: host specificity and adaptation. Parasitol Res. 2024;123:7.

    Article  Google Scholar 

  29. Tang R-X, Wang J, Li Y-F, Zhou C-R, Meng G-L, Li F-J, et al. Genomics and morphometrics reveal the adaptive evolution of pikas. Zool Res. 2022;43:813–26.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Lambert JP, Zhang X, Shi K, Riordan P. The pikas of China: a review of current research priorities and challenges for conservation. Integr Zool. 2023;18:110–28.

    Article  CAS  PubMed  Google Scholar 

  31. Li W, Feng Y, Zhang L, Xiao L. Potential impacts of host specificity on zoonotic or interspecies transmission of Enterocytozoon bieneusi. Infect Genet Evol. 2019;75:104033.

    Article  CAS  PubMed  Google Scholar 

  32. Perec-Matysiak A, Leśniańska K, Buńkowska-Gawlik K, Merta D, Popiołek M, Hildebrand J. Zoonotic genotypes of Enterocytozoon bieneusi in wild living invasive and native carnivores in Poland. Pathogens. 2021;10:1478.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Zhang X-X, Cong W, Lou Z-L, Ma J-G, Zheng W-B, Yao Q-X, et al. Prevalence, risk factors and multilocus genotyping of Enterocytozoon bieneusi in farmed foxes (Vulpes lagopus) northern China. Parasit Vectors. 2016;9:72.

    Article  PubMed  PubMed Central  Google Scholar 

  34. Ma Y-Y, Zou Y, Ma Y-T, Nie L-B, Xie S-C, Cong W, et al. Molecular detection and genotype distribution of Enterocytozoon bieneusi in farmed silver foxes (Vulpes vulpes) and arctic foxes (Vulpes lagopus) in Shandong Province, eastern China. Parasitol Res. 2020;119:321–6.

    Article  PubMed  Google Scholar 

  35. Shi K, Li M, Wang X, Li J, Karim MR, Wang R, et al. Molecular survey of Enterocytozoon bieneusi in sheep and goats in China. Parasit Vectors. 2016;9:23.

    Article  PubMed  PubMed Central  Google Scholar 

  36. Zhang N, Wu R, Ji T, Cui L, Cao H, Li D, et al. Molecular detection, multilocus genotyping, and population genetics of Enterocytozoon bieneusi in pigs in southeastern China. J Eukaryot Microbiol. 2020;67:107–14.

    Article  CAS  PubMed  Google Scholar 

  37. Xu J, Wang X, Jing H, Cao S, Zhang X, Jiang Y, et al. Identification and genotyping of Enterocytozoon bieneusi in wild Himalayan marmots (Marmota himalayana) and Alashan ground squirrels (Spermophilus alashanicus) in the Qinghai-Tibetan Plateau area (QTPA) of Gansu Province China. Parasit Vectors. 2020;13:367.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Zhang Q, Zhong Z, Xia Z, Meng Q, Shan Y, Guo Q, et al. Molecular epidemiology and genetic diversity of Enterocytozoon bieneusi in cervids from Milu Park in Beijing China. Animals (Basel). 2022;12:1539.

    Article  PubMed  Google Scholar 

  39. Zhang Y, Koehler AV, Wang T, Haydon SR, Gasser RB. First detection and genetic characterisation of Enterocytozoon bieneusi in wild deer in Melbourne’s water catchments in Australia. Parasit Vectors. 2018;11:2.

    Article  PubMed  PubMed Central  Google Scholar 

  40. Zhao W, Yu S, Yang Z, Zhang Y, Zhang L, Wang R, et al. Genotyping of Enterocytozoon bieneusi (Microsporidia) isolated from various birds in China. Infect Genet Evol. 2016;40:151–4.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

The author would like to thank all of the veterinarians who participated in the study as well as all of their colleagues who contributed to sample collection and sample preparation.

Funding

This work was supported in part by the Natural Science Foundation of China (82260399, 82260414 and 32202833), the Natural Science Key Project of Xinjiang Uygur Autonomous Region (2022B03014), which independently supports the establishment of science and technology Plans of Shihezi University (ZZZC202123) and Tianshan Young Talent Scientific and Technological Innovation Team: Innovative Team for Research on Prevention and Treatment of High-incidence Diseases in Central Asia (2023TSYCTD0020).

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Author notes

  1. Ziqi Wang, Nannan Cui, Jia Zhang and Zhixian Jiang contributed equally to this work.

Authors and Affiliations

  1. Key Laboratory for Prevention and Control of Emerging Infectious Diseases and Public Health Security of the XPCC, School of Medicine, Shihezi University, Shihezi, 832002, Xinjiang Uygur Autonomous Region, People’s Republic of China

    Ziqi Wang, Nannan Cui, Jia Zhang, Ruiqi Song, Wenbo Tan & Yuanzhi Wang

  2. NHC Key Laboratory of Prevention and Treatment of Central Asia High Incidence Diseases, Shihezi University, Shihezi, 832003, Xinjiang Uygur Autonomous Region, People’s Republic of China

    Ziqi Wang, Nannan Cui, Jia Zhang, Ruiqi Song, Wenbo Tan & Yuanzhi Wang

  3. Department of Forest, College of Agriculture, Shihezi University, Shihezi, 832002, Xinjiang Uygur Autonomous Region, People’s Republic of China

    Zhixian Jiang & Meihua Yang

  4. Department of Parasitology and Zoology, University of Veterinary Medicine, Budapest, Hungary

    Sándor Hornok

Authors
  1. Ziqi Wang
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Contributions

ZW, NC, JZ and YW conceived and designed the study, and wrote the manuscript. ZW, NC, WT, RS and MY performed the experiments and analyzed the data. ZW, HS and YW contributed to study design and edited the manuscript. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Yuanzhi Wang.

Ethics declarations

Ethics approval and consent to participate

This study was reviewed and approved by the ethics committee of School of Medicine, Shihezi University in accordance with the medical regulations of China (Approval numbers shown in Additional file 2: Table S2).

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Not applicable.

Competing interests

The authors declare no competing interests.

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Supplementary Information

13071_2025_6739_MOESM1_ESM.pdf

Additional file 1: Table S1. Prevalence and distribution of Enterocytozoon bieneusi genotypes in wild animals in Northwest China.

Additional file 2: Table S2. Animal species used as sources of samples in the study.

Additional file 3:

Characteristics of amplified fragments and corresponding primer sequences.

Additional file 4:

In this study, transmission routes involving the predator-prey relationship between hosts of genotype D and genotype NCF2 were investigated.

Additional file 5:

Genotype horse1 was detected in Himalayan marmots, owls, jungle nightjars and Hume’s groundpeckers, revealing habitat overlap in these animals.

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Wang, Z., Cui, N., Zhang, J. et al. Genetic diversity of Enterocytozoon bieneusi in 1099 wild animals and 273 imported pastured donkeys in northern China. Parasites Vectors 18, 105 (2025). https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s13071-025-06739-6

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Keywords

Parasites & Vectors

ISSN: 1756-3305

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