The Bos indicus cattle breeds have been naturally selected over thousands of years for disease resistance and thermo-tolerance. However, a genetic mechanism of these specific inherited characteristics needs to be discovered. Hence, in this study, the whole-genome comparative analysis of Bos indicus cattle breeds of Kangayam, Tharparkar, Sahiwal, Red Sindhi, and Hariana of the Indian subcontinent was conducted. The genetic variants identification analysis revealed a total of 15,58,51,012 SNPs and 1,00,62,805 InDels in the mapped reads across all Bos indicus cattle breeds. The functional annotation of 17,252 genes that comprised both, SNPs and InDels, of high functional impact on proteins, has been carried out. The functional annotation results revealed the pathways that were involved in the innate immune response including toll-like receptors, a retinoic acid-inducible gene I like receptors, NOD-like receptors, Jak-STAT signaling pathways, and the non-synonymous variants in the candidate immune genes. Further, we also identified several pathways involved in heat shock response, hair and skin properties, oxidative stress response, osmotic stress response, thermal sweating, feed intake, metabolism, and the non-synonymous variants in the candidate thermo-tolerant genes. These pathways and genes were directly or indirectly contributing to the disease resistance and thermo-tolerance adaptations of Bos indicus cattle breeds.