Molecular Epidemiology of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in Guizhou Angus Calves: Dominance of Angus Cattle-Adapted Genotypes and Zoonotic Potential of E. bieneusi

Abstract

Limited molecular data exist on zoonotic parasites Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in Angus calves from Guizhou, China. This study constitutes the first molecular epidemiological survey of these pathogens in this region. 817 fecal samples from Angus calves across 7 intensive beef farms (Bijie City). Nested PCR methods targeting SSU rRNA (Cryptosporidium spp.), gp60 (Cryptosporidium bovis subtyping), bg/gdh/tpi (G. duodenalis), and ITS (E. bieneusi) coupled with DNA sequencing were employed. DNA sequences were analyzed against the NCBI. database. Statistical differences were assessed via a generalized linear mixed-effects model. Cryptosporidium spp. prevalence 23.5% (192/817; 95% CI 28.1-34.6%), with C. bovis predominating 89.6% (172/192; 95% CI 84.4-93.5%) and six subtypes (XXVIa-XXVIf). Highest infection in 4-8-week-olds 29.9% (143/479; 95% CI 25.8-34.1%) (p < 0.01). G. duodenalis: 31.3% (256/817; 95% CI 28.1-34.6%) positive, overwhelmingly assemblage E 97.6% (6/256; 95% CI 0.9-5.0%), zoonotic assemblage A was marginal 0.7% (6/817; 95% CI 0.3-1.6%). Farm-level variation exceeded 10-fold (e.g., Gantang: 55.0% (55/100; 95% CI 44.7-65.0%) vs. Tieshi: 4.9% (5/102; 95% CI 1.6-11.1%). E. bieneusi: prevalence 19.7% (161/817; 95% CI 17.0-22.6%), exclusively zoonotic genotypes BEB4: 49.7% (80/161; 95% CI 41.7-57.7%); I: 40.4% (65/161; 95% CI 32.7-48.4%). Strong diarrhea association (p < 0.01) and site-specific patterns (e.g., Guanyindong: 39.2%). While Giardia exhibited the highest prevalence (31.3%) with minimal zoonotic risk, Enterocytozoon-despite lower prevalence (19.7%)-posed the greatest public health threat due to exclusive circulation of human-pathogenic genotypes (BEB4/I) and significant diarrhea association, highlighting divergent control priorities for these enteric parasites in Guizhou calves. Management/Public health impact: Dominant zoonotic E. bieneusi genotypes (BEB4/I) necessitate: 1. Targeted treatment of 4-8-week-old Angus calves. 2. Manure biofermentation (≥55 °C, 3 days), and 3. UV-disinfection (≥1 mJ/cm²) for karst water to disrupt transmission in this high-humidity region.

Keywords: Cryptosporidium spp.; Enterocytozoon bieneusi; Giardia duodenalis; molecular epidemiology; zoonotic parasites.

Figure 1

Geographic distribution of Angus cattle sampling sites in Guizhou Province, China.