ABSTRACT: Bovine tuberculosis (bTB), caused by Mycobacterium bovis (M. bovis), poses a signi!cant global health and economic burden. Despite extensive research, a comprehensive understanding of M. bovis pathogenesis, particularly its transcriptional adaptation across different growth phases and within the host environment, remains incomplete. Here, we performed a comprehensive transcriptomic analysis of virulent M. bovis and the attenuated M. bovis BCG strain (BCG) across early-log, mid-log, and stationary growth phases to elucidate the molecular underpinnings of their phenotypic distinctions. RNA sequencing revealed distinct gene expression pro!les, highlighting signi!cant differences in pathways related to cell wall biosynthesis, lipid metabolism, transcriptional regulation, protein secretion, and the PE/PPE protein family. Notably, the virulent M. bovis exhibited an upregulation of virulence-associated genes, especially those involved in lipid metabolism and secretion systems, during the transition from early- and mid-log phases to the stationary phase. In contrast, BCG demonstrated an enrichment in stress response and metabolic adaptation pathways during the similar transition. Gene network analysis of differentially expressed genes (DEGs) provided a systems-level view of the transcriptional programs governing M. bovis and M. bovis BCG physiology, identifying !ve key modules of co-expressed genes that regulate small molecules transport, amino acid biosynthesis and immune evasion in M. bovis. Furthermore, we analyzed M. bovis transcriptional responses during murine lung infection, identifying a core set of DEGs linked to host-pathogen interactions and mechanisms of persistence. These !ndings offer novel insights into M. bovis adaptation strategies during survival under de!ned, in vitro growth conditions and highlight potential targets for improved vaccine design and therapeutic interventions following further analyses during host infection.
