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Enterocytozoon bieneusi is a zoonotic pathogen that infects a variety of hosts including humans, livestock, wildlife, companion animals, and birds, as well as being abundant in the environment. Humans and nonhuman animals could be infected with E. bieneusi via consumption of food or water that contains zoonotic and host-adapted genotypes. In this study, 288 fecal specimens were collected from farmed minks, blue foxes, and raccoon dogs, in Xinjiang, China. Enterocytozoon bieneusi was examined by PCR amplification based on sequence analysis of the internal transcribed spacer (ITS) region. The overall infection rate of E. bieneusi was 4.9% (14/288), with mink samples showing the highest infection rate (5.6%, 12/214), followed by blue foxes (2.9%, 1/35), and then raccoon dogs (2.6%, 1/39). Six E. bieneusi genotypes were identified, including D (n = 5), PigEBITS7 (n = 4), EbpA (n = 2), CAM5 (n = 1), WildBoar3 (n = 1), and a novel genotype XJMI-1 (n = 1). Phylogenetic analysis showed that all E. bieneusi genotypes belonged to group 1, which composed of over 300 genotypes and most of them have been identified in human and variety of animals, suggesting a risk of zoonotic transmission from farmed wildlife to humans., Graphical abstract Image 1, Highlights • Six genotypes were identified, including three zoonotic genotypes (D, EbpA ad PigEBIST7). • Farmed wildlife maybe a potential source of E. bieneusi infection for other animals and humans.

Enterocytozoon bieneusi, an obligate intracellular pathogen of the intestinal epithelium, is commonly identified in humans and many other animals and is ubiquitous in water sources and the environment generally. To determine the molecular prevalence of E. bieneusi in edible bullfrogs (Lithobates catesbeiana) and evaluate the possibility of its potential zoonotic transmission to humans via food or water, the intestinal contents of 295 bullfrogs were intermittently collected from two open markets in Aksu, China. The samples were screened for the internal transcribed spacer by polymerase chain reaction amplifications, revealing that 20.7% (61/295) of them were infected with E. bieneusi, with no significant differences found between the two sampling locations (p > 0.05). Twenty-two different E. bieneusi genotypes were identified, including one known genotype (EbpC) and 19 novel ones (named BLC1 to BLC19). The zoonotic genotype EbpC was identified in most of the E. bieneusi-positive samples (65.6%, 40/61). The remaining genotypes were identified in either one or three samples each. Our phylogenetic analysis showed that 20 of the E. bieneusi genotypes belonged to Group 1. As far as we are aware, this is the first report of E. bieneusi infections in edible bullfrogs. Our findings suggest that E. bieneusi can be maintained in edible bullfrogs and potentially transmitted via food or water. It is possible that these amphibians are unsuspected zoonotic reservoirs of E. bieneusi. Keywords: Enterocytozoon bieneusi, Bullfrogs, Genotype, Zoonotic

Cryptosporidium and Enterocytozoon bieneusi are important intestinal pathogens that infect humans and various animals. Few reports are available regarding the infections of the two pathogens in Père David's deer. In this study, polymerase chain reaction (PCR) confirmed Cryptosporidium infection in two (1.6%) and E. bieneusi in 45 (35.2%) of 128 fecal samples collected from Père David's deer in the National Nature Reserve of Shishou, Hubei Province, China. C. parvum (n = 1) and Cryptosporidium deer genotype (n = 1) were identified using the small subunit rRNA (SSU rRNA) gene. The C. parvum was further subtyped as IIdA20G1 by sequencing analysis of the 60-kDa glycoprotein (gp60) gene. The identity of E. bieneusi was confirmed by an internal transcribed spacer (ITS) gene; the HLJD-V (n = 42) and MWC_d1 (n = 3) genotypes were identified, with the former clustering in group 2 and the latter in group 1. These data suggest that the Père David's deer were infected with host-specific and/or zoonotic genotypes of these pathogens, implicating Père David's deer could be a potential source of human Cryptosporidium infection., Graphical abstract Image 1, Highlights • C. parvum subtype IIdA20G1 was firstly identified in Pere David's deer. • Genotypes HLJD-V and MWC_d1 were detected from 45 E. bieneusi-positive samples. • The present study implicated Père David's deer could be a potential source of human infection.

Cryptosporidium is one of the most prevalent zoonotic parasites and is responsible for the high burden of diarrheal disease across the globe. Rodents are globally overpopulated and are reservoirs for a variety of zoonotic pathogens. Bamboo rats are a common species of rodent that are bred for meat and wool in China. However, the genetic characterization of Cryptosporidium in bamboo rats in China is limited. The aim of this study was to determine the occurrence and genetic characterization of Cryptosporidium in bamboo rats from South Central China. From February2017to February 2018, 435 fecal samples were collected from bamboo rats in 13 farms located in 12 cities in South Central China. All fecal specimens were examined for Cryptosporidium by PCR, and through sequencing the partial small subunit of ribosomal DNA (SSU rRNA). C. parvum-positive samples were further subtyped through analysis of the 60-kDa glycoprotein (gp60) gene sequence. Meanwhile, all the new Cryptosporidium genotypes samples were selected for further sequence characterization at the 70-kDa heat shock protein (HSP70) gene and oocyst wall protein (COWP) gene as well as gp60 gene. Infection rates of 2.1% (9/435) were recorded for Cryptosporidium. Sequence analysis confirmed the presence of two Cryptosporidium species including C. parvum (n = 2), C. occultus (n = 1) and two new Cryptosporidium genotypes termed Cryptosporidium bamboo rat genotype I (n = 5) and Cryptosporidium bamboo rat genotype II (n = 1). Two subtypes of C. parvum were identified including IIdA15G1 and IIpA19 (one each).The discovery of zoonotic Cryptosporidium species/genotypes in bamboo rats suggests they have significant zoonotic potential and pose a threat to human health. The novel sequences discovered provide new insight into genotypic variations in Cryptosporidium in bamboo rats. Keywords: Cryptosporidium, Bamboo rat, Molecular detection, Zoonotic, Public health