Intra-sequences (query) similarity matrix.

Theileria orientalis, an obligatory intracellular blood protozoon, which causes the substantial economic losses to the cattle industry and is widely prevalent throughout Bangladesh. The study was aimed to assess the nationwide prevalence, genetic diversity and evolutionary divergence of T. orientalis parasites in Bangladesh by analyzing bovine blood samples. The genetic characterization, haplotype network analysis and Codon-based evolutionary divergence was conducted by targeting the major piroplasm surface protein (MPSP) gene. The overall prevalence of T. orientalis was 63.75% (n = 800), as determined by MPSP-PCR across the eight divisions of Bangladesh with significant regional variation. Subsequent phylogenetic analysis showed that the query MPSP sequences were clustered into two genotypes namely 5 and 7. Genetic diversity indicated high haplotype and moderate nucleotide diversity but consistent with contrasting demographic and selective pressures across regions. The evolutionary divergence analysis revealed the complex genetic landscape of T. orientalis population which shaped by historical expansion, ongoing gene flow and localized diversification. The haplotype network analysis identified total 29 distinct haplotypes, highlighting substantial diversity within the two genotypes. These findings demonstrate the occurrence and genetic richness of T. orientalis in cattle of Bangladesh. Although the pathogenic impact could not be assessed in this study, the presence of diverse genotypes suggests a need for continued surveillance and future research to determine the clinical and economic relevance of this parasite.