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

Limited molecular data exist on zoonotic parasites <i>Cryptosporidium</i> spp., <i>Giardia duodenalis</i>, and <i>Enterocytozoon bieneusi</i> 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 (<i>Cryptosporidium</i> spp.), <i>gp60</i> (<i>Cryptosporidium bovis</i> subtyping), <i>bg</i>/<i>gdh</i>/<i>tpi</i> (<i>G. duodenalis</i>), and ITS (<i>E. bieneusi</i>) 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. <i>Cryptosporidium</i> spp. prevalence 23.5% (192/817; 95% CI 28.1-34.6%), with <i>C. bovis</i> 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%) (<i>p</i> < 0.01). <i>G. duodenalis</i>: 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%). <i>E. bieneusi</i>: 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 (<i>p</i> < 0.01) and site-specific patterns (e.g., Guanyindong: 39.2%). While <i>Giardia</i> exhibited the highest prevalence (31.3%) with minimal zoonotic risk, <i>Enterocytozoon</i>-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 <i>E. bieneusi</i> 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.