CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES >

2025 , Vol. 43 >Issue 5: 670 - 677

DOI: https://doi.org/10.12140/j.issn.1000-7423.2025.05.011

ORIGINAL ARTICLES

Infection and genotype identification of Enterocytozoon bieneusi and Cryptosporidium in Wuzhishan pigs of Hainan

  • DONG Yatong ,
  • ZHANG Yan ,
  • LIU Guangliang ,
  • CAO Zongxi ,
  • YU Miao ,
  • CHEN Linlin ,
  • LIU Hailong ,
  • CHEN Suzhen
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  • 1 College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, Xinjiang, China
    2 Institute of Animal Science and Veterinary Medicine of Hainan Academy of Agricultural Sciences, Key Laboratory of Tropical Animal Breeding and Disease Research, XIA Xianzhu Academician Team Innovation Center, Haikou 571100, Hainan, China

Received date: 2025-06-20

  Revised date: 2025-09-12

  Online published: 2025-10-27

Supported by

National Key Research and Development Program of China(2023YFC3404302)

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Abstract

Objective To analyze the species of Cryptosporidium and the genotype of Enterocytozoon bieneusi in Wuzhishan pigs in Hainan, and to master the infection of the two protozoa. Methods From June 2024 to February 2025, fresh fecal samples of Wuzhishan pigs were collected from 4 farms in Zuntan Town, Qiongzhong Li and Miao Autonomous County, Wuzhishan City and Chengmai County of Hainan Province (marked as A, B, C and D farms, respectively). DNA was extracted from fecal samples, and the 18S ribosomal RNA (18S rRNA) of Cryptosporidium and the internal transcribed spacer (ITS) sequence of E. bieneusi were amplified by nested PCR. The positive amplification products were sequenced and compared to identify Cryptosporidium species or E. bieneusi genotypes. SPSS 26.0 software was used for statistical analysis. Pearson chi-square (χ2) test was used to compare the differences in positive rates, and the risk coefficient (OR) and 95% confidence interval (CI) were calculated. The phylogenetic tree was constructed by MEGA 11 software adjacency method. Results A total of 137 Wuzhishan pig manure samples were collected, of which 41 were amplified with 830 bp of Cryptosporidium 18S rRNA sequence and 69 were amplified with 390 bp of E. bieneusi ITS sequence. The total positive rate of Cryptosporidium in Wuzhishan pigs from four farms was 29.9% (41/137), with the highest in C farm (64.3%, 18/28) and the lowest in B farm (20.3%, 14/69). The positive rate of Cryptosporidium in post-weaning piglets was 57.1% (12/21), which was higher than 7.7% (2/26) in pre-weaning piglets (χ2 = 13.583, P < 0.05). The OR of post-weaning piglets and adult pigs was 16.00 times (95% CI = 2.98-85.99) and 5.14 times (95% CI = 1.15-23.31) higher than that of pre-weaning piglets. The total positive rate of E. bieneusi was 50.4% (69/137), with the highest in C farm (60.7%, 17/28) and the lowest in A farm (37.5%, 9/24). The positive rate of E. bieneusi in post-weaning piglets was 80.9% (17/21), which was higher than that in pre-weaning piglets (42.3%, 11/26) and adult pigs (45.5%, 41/90) (χ2 = 7.204, 8.551; both P < 0.01). The OR of post-weaning piglets and adult pigs was 5.80 times (95% CI = 1.52-22.10) and 1.14 times (95% CI = 0.47-2.76) higher than that of pre-weaning piglets. The positive rate of E. bieneusi in grazing Wuzhishan pigs was 62.8% (44/70), which was higher than that in barn feeding Wuzhishan pigs (37.3%, 25/67) (χ2 = 8.935, P < 0.01). The OR of infection in grazing Wuzhishan pigs was 2.84 times (95% CI = 1.42-5.69) higher than that in feeding Wuzhishan pigs. The results of sequence alignment showed that 41 Cryptosporidium 18S rRNA sequences were identified as C. scrofarum, and the sequence identity with C. scrofarum (GenBank accession number: PQ856498) was 93.36%-99.74%. In the phylogenetic tree, it was in the same large branch with C. scrofarum from Yunnan pig and Hubei pig (GenBank accession number: PQ856498, ON149807). 69 ITS sequences of E. bieneusi were identified as H, D, and EpbC genotypes. Among them, 3 H genotype sequences were 98.00% identical to the H genotype of German pig (GenBank accession number: AF135835.1). The identity of 25 D genotype sequences with D genotype (GenBank accession number: MK696083.1) from Australian cats was 97.51%-99.72%, and the identity of 41 EpbC genotype sequences with EpbC genotype (GenBank accession number: KY950535.1) from Chinese pandas was 98.59%-99.73%. In the phylogenetic tree, D genotype was in the same branch with Argentinean D genotype and Australian cat D genotype (GenBank accession number: OP650902, MK696083), EbpC genotype was in the same branch with Chinese panda EpbC genotype (GenBank accession number: KY950535.1), and H genotype was in the same branch with German pig H genotype (GenBank accession number: AF135835.1). Phylogenetic analysis showed that the three genotypes belonged to one subgroup. Conclusion There were E. bieneusi and Cryptosporidium infection in Wuzhishan pigs in Hainan. The Cryptosporidium infection species was C. scrofarum. The main genotypes of E. bieneusi were H, D and EbpC.

Key words： Enterocytozoon bieneusi; Cryptosporidium; Wuzhishan pig; Zoonotic diseases; Genotype

Cite this article

DONG Yatong , ZHANG Yan , LIU Guangliang , CAO Zongxi , YU Miao , CHEN Linlin , LIU Hailong , CHEN Suzhen . Infection and genotype identification of Enterocytozoon bieneusi and Cryptosporidium in Wuzhishan pigs of Hainan[J]. CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES, 2025 , 43(5) : 670 -677 . DOI: 10.12140/j.issn.1000-7423.2025.05.011

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