Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are significant parasitic gastrointestinal pathogens with global distribution in humans and domestic animals, including calves. The main symptoms of calf infection are severe diarrhea, dehydration, growth retardation, and sometimes even death. To date, there has been limited information on the prevalence of Cryptosporidium spp., G. duodenalis, and E. bieneusi infections in calves in Ningxia, China, especially between diarrheic and non-diarrheic calves. A total of 438 fecal samples were collected from diarrheic (201) and non-diarrheic (237) calves in Ningxia. PCR and DNA sequencing were used to find the prevalence of Cryptosporidium spp. at 46.8% (205/438), G. duodenalis at 16.9% (74/438), and E. bieneusi at 10.0% (44/438). The prevalence of Cryptosporidium spp. infection in diarrheic and non-diarrheic calves was 54.0% (128/237) and 38.3% (77/201), respectively, and statistical analysis showed a positive correlation between the prevalence of Cryptosporidium spp. infection and calf diarrhea (p < 0.01). However, in this study, there was no statistical correlation between the prevalence of G. duodenalis infection as well as E. bieneusi infection and calf diarrhea (p > 0.05). Furthermore, four known Cryptosporidium species were successfully identified by comparing them with SSU rRNA gene sequences, including C. parvum, C. bovis, C. ryanae, and C. andersoni. In addition, all 74 G. duodenalis-positive samples were identified as assemblage E by comparative analysis of bg gene sequences. Among the 44 E. bieneusi-positive samples sequenced in the present study, 4 distinct E. bieneusi genotypes were successfully identified by comparative analysis of ITS sequences, including 3 known genotypes (J, BEB4, and N) and 1 novel genotype, the latter of which was identified and designated as NX1. These findings indicated that the high genetic diversity and complex population structures of Cryptosporidium spp., G. duodenalis, and E. bieneusi in Ningxia diarrhea calves and non-diarrhea calves, which provide new data for understanding the epidemiological status of Cryptosporidium spp., G. duodenalis, and E. bieneusi in Ningxia calves.
IntroductionCalf diarrhea is a complex disease that has long been an unsolved problem in the cattle industry. Ningxia is at the forefront of China in the scale of cattle breeding, and calf diarrhea gravely restricts the development of Ningxia's cattle industry.MethodsFrom July 2021 to May 2022, we collected diarrhea stool samples from calves aged 1–103 days from 23 farms in five cities in Ningxia, and performed PCR using specific primers for 15 major reported pathogens of calf diarrhea, including bacteria, viruses, and parasites. The effect of different seasons on the occurrence of diarrhea in calves was explored, the respective epidemic pathogens in different seasons were screened, and more detailed epidemiological investigations were carried out in Yinchuan and Wuzhong. In addition, we analyzed the relationship between different ages, river distributions and pathogen prevalence.ResultsEventually, 10 pathogens were detected, of which 9 pathogens were pathogenic and 1 pathogen was non-pathogenic. The pathogens with the highest detection rate were Cryptosporidium (50.46%), Bovine rotavirus (BRV) (23.18%), Escherichia coli (E. coli) K99 (20.00%), and Bovine coronavirus (BCoV) (11.82%). The remaining pathogens such as Coccidia (6.90%), Bovine Astrovirus (BoAstV) (5.46%), Bovine Torovirus (BToV) (4.09%), and Bovine Kobuvirus (BKoV) (3.18%) primarily existed in the form of mixed infection.DiscussionThe analysis showed that different cities in Ningxia have different pathogens responsible for diarrhea, with Cryptosporidium and BRV being the most important pathogens responsible for diarrhea in calves in all cities. Control measures against those pathogens should be enforced to effectively prevent diarrhea in calves in China.
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