Characterisation and subtype distribution of blastocystis sp. in cattle : its association with potential risk factor and protease activity

Abstract

Blastocystis sp. is an eukaryotic protozoan with 17 distinctive subtypes discovered in animals and humans worldwide. Cattle is one of important source of animal protein, which shown an increasing demand for its produce due to increased in growth of population especially in developing countries. However, the production is likely to be affected by infection and disease management of this animal. Currently, the zoonotic pathogen of Blastocystis sp. from cattle was identified with multiple concurrent infections with rates of infection as high as 80%. Unlike human, animal particularly cattle infected with Blastocystis sp. are commonly healthy carriers and serve as a main reservoir in transmitting the infection to human. Though, Blastocystis exhibited protease activity that suggest its pathogenicity, but its effect on host remain unclear. Thus, this study was aimed to determine occurrence of Blastocystis sp. isolated from cattle and subtypes variation in the protease activity for better understanding of the host-parasite relationships and effect of location and farm management on Blastocystis infection. A total of 120 faecal samples of cattle were collected from three farms in Pahang, Malaysia for in-vitro cultivation and microscopy identification. The gender and age of the cattle as well as management system of the farms were also noted during the sampling. Later, DNA extracted from the faecal were subjected to genotyping analysis before protease activity of three selected subtypes were determined using azocasein assay of colorimetric quantification method. The study found 30 out of 120 (25%) cattle infected with Blastocystis sp. with vacuolar as the dominant form observed during cultivation. While gender has no association with the occurrence of Blastocystis sp. and cattle of age below 3 months as well as Muazam Shah farm with integrated system were significantly (p<0.05) associated with the infection. Molecular genotyping revealed that Blastocystis STlO (21.3%) occurred predominantly in the cattle with STl (2.5%), ST3 (7.5%), ST4 (2.5%), ST5 (8.8%) and ST14 (1.3%). Phylogenetic analysis found that these subtypes were closely related and had shared common ancestors with high homologous sequences of genetic relation. ST3, ST5 and STl 0 exhibited inter-and intra-subtype quantitative protease activity variation, which mainly expressed cysteine protease and partially serine protease, aspartic protease and metallo-protease in the respective subtypes. In conclusion, moderate rates of Blastocystis sp. infection with six different subtypes were identified in the cattle. The farm management systems, cleaning and sanitation as well as segregation condition influence the distribution of Blastocystis sp. infection which significantly associated with age and condition of the farms. While, protease activity were commonly been reported in ST3, ST4 and ST?, this study has discovered in the ST3, ST5 and STl 0 of Blastocystis. The variant observed in the protease activity indicate that isolates of different subtypes may exhibit different pathogenic condition upon exhibition of diseases, yet suggests for more studies needed in the future. Nevertheless, this study presented updates on the occurrence of Blastocystis sp. in cattle from Pahang, Malaysia. This information is important in understanding host-parasite relationship associated with gastrointestinal diseases and identification of possible virulence factor in the future.