Molecular prevalence and associated risk factors of Cryptosporidium spp. infection in dairy cattle in Khon Kaen, Thailand

Abstract

Background and aim: Cryptosporidium spp. are important parasites in the small intestines of humans and animals, particularly cattle. The aim of this study was to estimate the molecular prevalence and associated risk factors of Cryptosporidium infection in dairy cattle in five districts of Khon Kaen province, Thailand, and to identify Cryptosporidium spp.

Materials and methods: From July 2020 to October 2021, 296 fecal samples were collected from three groups of dairy cattle: Calves aged <3 months, calves aged 3 months-1 year, and calves aged >1 year. Cryptosporidium spp. were detected by polymerase chain reaction (PCR) amplifying the 18s RNA gene. Both genus-specific and species-specific primers were used to identify Cryptosporidium confirmed by DNA sequencing. Age, house floor type, and water trough type were evaluated as risk factors. We analyzed all associated risk factor information using the logistic regression test in the Statistical Package for the Social Sciences.

Results: PCR results showed that 40 (13.51%) out of 296 samples were positive for Cryptosporidium spp., including Cryptosporidium bovis (57.50%) and Cryptosporidium ryanae (2.50%). There was a significant association between Cryptosporidium incidence, cattle age, and house floor type (p < 0.05). National Center for Biotechnology Information Basic Local Alignment Search Tool displayed 99.48%-100% nucleotide similarity of each Cryptosporidium spp. isolate with references recorded on GenBank.

Conclusion: C. bovis and C. ryanae are commonly found in dairy cattle, especially calves, in Khon Kaen, Thailand, and the incidence was associated with age and house floor type. A molecular technique may be influential for species identification. The results of the present study would provide useful information for veterinarians and animal owners to understand better Cryptosporidium spp. and how to manage farms properly.

Keywords: Cryptosporidium; bovine cryptosporidiosis; dairy cattle; molecular prevalence.

Figure-1

Location of research area (source: https://en.wikipedia.org/wiki/Khon_Kaen_province).

Figure-2

Gel electrophoresis of PCR products. (a) Lane 1–7=Cryptosporidium spp. (840 bp) (b) Lane 8–11=Cryptosporidium bovis (241 bp) (c) Lane 12–14=Cryptosporidium ryanae (415 bp) M=Marker (2000–100 bp). PCR=Polymerase chain reaction.

Figure-3

Alignment of the local sequences obtained from 18S rRNA gene of each Cryptosporidium bovis and Cryptosporidium ryanae from this study and some reference sequences from GenBank. Dots (.) denote the identical nucleotides.

Figure-4

Phylogenetic tree of local Cryptosporidium ryanae (red) and Cryptosporidium bovis (purple) Khon Kaen isolates, and the references recorded on GenBank. The phylogenetic tree was constructed using the maximum likelihood method in MEGA 11 software. Numbers indicate the bootstrap percentages from 1000 replicates.