BACKGROUND: Tuberculosis disease is a major global health concern. In Mexico, information regarding the genomic variants of Mycobacterium tuberculosis (MTB) prevailing in the country and the existence of specific biogeographical patterns remains extremely scarce. OBJECTIVE: This study aimed to identify the genotypic patterns of MTB isolates in Mexico and determine the genes and specific single nucleotide polymorphisms involved in the evolution of these populations. METHODS: Phylogenomic and pan-genomic analyses were performed using publicly available Mexican MTB genomes along with 33 newly sequenced genomes from Jalisco, considering a global context. RESULTS: The L4 sublineages of MTB, such as L4.1.1 (X), L4.1.2 (H), and L4.3 (LAM), were the most prevalent in Mexico. We found exclusive mutations and gene clusters in a virulent sublineage L4.1.1.3 (X3), which is endemic to Mexico. These genes encoded three PE/PPE family proteins: a multidrug transporter, thioredoxin domain-containing protein, quinone-dependent l-lactate dehydrogenase, DUF1725 domain-containing protein, amidase, poly (A) polymerase, and six hypothetical/uncharacterised proteins. Additionally, the genes encode an ESX-1 secretion-associated protein and a deazaflavin-dependent nitroreductase (ddn). CONCLUSION: X3 was distinguished from the rest of the sublineages by containing genes related to pathogenicity and virulence, as well as a gene linked to delamanid, an antibiotic for active multidrug-resistant tuberculosis. These findings provide valuable insight into the circulation and spread of MTB in Mexico.