*Article* **Molecular Investigation of Zoonotic Intestinal Protozoa in Pet Dogs and Cats in Yunnan Province, Southwestern China**

**Yu-Gui Wang 1,2,3, Yang Zou 2,\*, Ze-Zhong Yu 4, Dan Chen 5, Bin-Ze Gui 3, Jian-Fa Yang 1, Xing-Quan Zhu 1,6, Guo-Hua Liu <sup>3</sup> and Feng-Cai Zou 1,\***


**Citation:** Wang, Y.-G.; Zou, Y.; Yu, Z.-Z.; Chen, D.; Gui, B.-Z.; Yang, J.-F.; Zhu, X.-Q.; Liu, G.-H.; Zou, F.-C. Molecular Investigation of Zoonotic Intestinal Protozoa in Pet Dogs and Cats in Yunnan Province, Southwestern China. *Pathogens* **2021**, *10*, 1107. https://doi.org/10.3390/ pathogens10091107

Academic Editors: David Carmena, David González-Barrio and Pamela Carolina Köster

Received: 4 August 2021 Accepted: 23 August 2021 Published: 30 August 2021

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**Abstract:** *Giardia duodenalis*, *Enterocytozoon bieneusi* and *Cryptosporidium* spp. are common enteric pathogens that reside in the intestines of humans and animals. These pathogens have a broad host range and worldwide distribution, but are mostly known for their ability to cause diarrhea. However, very limited information on prevalence and genotypes of *G. duodenalis*, *E. bieneusi* and *Cryptosporidium* spp. in pet dogs and cats are available in China. In the present study, a total of 433 fecal samples were collected from 262 pet dogs and 171 pet cats in Yunnan province, southwestern China, and the prevalence and the genotypes of *G. duodenalis*, *E. bieneusi* and *Cryptosporidium* spp. were investigated by nested PCR amplification and DNA sequencing. The prevalence of *G. duodenalis*, *E. bieneusi* and *Cryptosporidium* spp. was 13.7% (36/262), 8.0% (21/262), and 4.6% (12/262) in dogs, and 1.2% (2/171), 2.3% (4/171) and 0.6% (1/171) in cats, respectively. The different living conditions of dogs is a risk factor that is related with the prevalence of *G. duodenalis* and *E. bieneusi* (*p* < 0.05). However, there were no statistically significant difference in prevalence of three pathogens in cats. DNA sequencing and analyses showed that four *E. bieneusi* genotypes (PtEb IX, CD9, DgEb I and DgEb II), one *Cryptosporidium* spp. (*C. canis*) and two *G. duodenalis* assemblages (C and D) were identified in dogs; two *E. bieneusi* genotypes (Type IV and CtEb I), one *Cryptosporidium* spp. (*C. felis*) and one *G. duodenalis* assemblage (F) were identified in cats. Three novel *E. bieneusi* genotypes (DgEb I, DgEb II and CtEb I) were identified, and the human-pathogenic genotypes/species Type IV *C. canis* and *C. felis* were also observed in this study, indicating a potential zoonotic threat of pet dogs and cats. Our results revealed the prevalence and genetic diversity of *G. duodenalis*, *E. bieneusi* and *Cryptosporidium* spp. infection in pet dogs and cats in Yunnan province, southwestern China, and suggested the potential threat of pet dogs and cats to public health.

**Keywords:** *Giardia duodenalis*; *Enterocytozoon bieneusi*; *Cryptosporidium* spp.; zoonotic genotypes; pet dogs and cats; Yunnan province; China
