Multilocus genotyping of Giardia duodenalis in pre-weaned calves with diarrhea in the Republic of Korea

Giardia duodenalis is a protozoan parasite that infects humans, companion animals, livestock, and wildlife. Infections in cattle caused by this parasite are often asymptomatic, but such infections can cause diarrhea, reduced weight gain, and ill-thrift in young calves. Although G. duodenalis causes diarrhea in calves, only a few studies have been conducted on calves in the Republic of Korea (ROK). Here, we aimed to determine the prevalence and distribution of G. duodenalis assemblages in pre-weaned calves with diarrhea in the ROK, identify the association between the occurrence of G. duodenalis and the age of calf, and perform molecular characterization of G. duodenalis. We collected 455 fecal samples from pre-weaned native Korean calves (<= 60 days old) with diarrhea in four different regions. G. duodenalis was detected using nested PCR targeting the beta-giardin (bg) gene, and positive samples were further genotyped for the glutamate dehydrogenase (gdh) and triosephosphate isomerase (tpi) genes. The overall prevalence of G. duodenalis in calves with diarrhea was 4.4% (20/455) based on the analysis of bg. The highest prevalence was observed in calves aged 11-30 days (7.5%; 95% confidence interval: 3.7%-11.3%), whereas the lowest prevalence was observed in neonatal calves. From the 20 samples that were positive for bg, 16, 5, and 6 sequences were obtained following genotyping of bg, gdh, and tpi, respectively. Sequencing analysis of the bg gene revealed the presence of assemblage E (n = 15) and sub-assemblage AI (n = 1) in the samples. Moreover, we detected mixed infections with assemblages E and A in two calves for the first time. Among the sequences obtained herein, two new subtypes of assemblage E were detected in gdh and tpi sequences each. The results suggest that G. duodenalis is an infectious agent causing diarrhea in calves, and pre-weaned calves are at a higher risk of infection than neonatal calves. Multilocus genotyping should be performed to confirm the presence of potentially zoonotic genotypes. These results highlight the importance of cattle as a source of zoonotic transmission of G. duodenalis to humans.