High-throughput profiling of the T cell receptor delta CDR3 repertoire reveals species-specific patterns in cattle (Bos taurus) and water buffalo (Bubalus bubalis)

Background γδ T cells constitute a substantial proportion of lymphocytes in ruminants, and the diversity of their immune receptors is critical for understanding species-specific immune functions. The complementarity-determining region 3 (CDR3) of the T cell receptor delta chain (TRD) is a key structural determinant of γδ T cell antigen recognition; however, systematic comparative analyses of TRD immune repertoire characteristics across different bovine species remain limited. In this study, high-throughput sequencing and comprehensive analysis of the TRD CDR3 immune repertoire were performed in 7 cattle (Bos taurus) and 5 water buffalo (Bubalus bubalis). Results The results demonstrated good consistency in sequencing depth and data quality between the two groups. Diversity analysis revealed that the cattle TRD CDR3 repertoire exhibited higher clonal evenness and overall diversity than that of buffalo. Clonotype composition analysis showed that both species were dominated by medium- to high-frequency clonotypes, whereas buffalo relied more heavily on a limited number of highly expanded clonotypes. V(D)J gene usage analysis identified pronounced species-specific preferences in TRDV gene usage, with high intra-group consistency but low inter-species correlation; in contrast, TRDJ gene usage was highly conserved between the two species. The CDR3 length distributions in both groups displayed similar bell-shaped patterns, suggesting structural constraints during evolution, while K-mer and motif analyses revealed differences in CDR3 microstructural features between species. Furthermore, shared clonotype analysis indicated a limited number of public CDR3 amino acid sequences between cattle and buffalo, with highly abundant shared clonotypes enriched only in a small subset of individuals. Conclusions Collectively, this study provides a systematic characterization of the similarities and differences in the TRD CDR3 immune repertoires of cattle and water buffalo, offering fundamental data and a comparative perspective for understanding the mechanisms shaping γδ TCR diversity and their potential immunological functions in high γδ T cell species.