Genomic diversity and transmission routes of Mycobacterium tuberculosis isolates in Morocco.

Whole-genome sequencing (WGS) provides high-resolution insight into the genetic diversity, transmission dynamics, and drug-resistance mechanisms of Mycobacterium tuberculosis (MTB), yet genomic data from Morocco remain limited. In this study, we analyzed 68 WGS datasets of Moroccan MTB isolates and contextualized them using 19 global reference genomes. Lineages and resistance-associated mutations were determined with TB-Profiler. Pairwise single-nucleotide polymorphism (SNP) distances, phylogeny and transmission clusters were defined using well-established SNP thresholds implemented in the MTBseq pipeline. In silico spoligotypes were further compared with the SITVITEXTEND database to place Moroccan isolates in a broader regional context. Results demonstrated that circulating MTB strains in Morocco were dominated by Euro-American Lineage 4 (89.7%), with a smaller contribution from the Beijing lineage (L2.2.1). Resistance to rifampicin and isoniazid was mainly driven by canonical rpoB Ser450Leu and katG Ser315Thr mutations, while resistance to other drugs involved diverse variants. Seventeen genomic clusters encompassing 73.5% of isolates were identified, including nine clusters with isolates differing by 0-5 SNPs, consistent with very recent transmission. Multidrug-resistant and pre-extensively drug-resistant isolates were disproportionately concentrated within LAM 4.3 and Beijing sub-lineages. Spoligotyping comparisons confirmed the predominance of Euro-American lineages across Morocco and neighboring Mediterranean countries. This first comprehensive WGS-based analysis provides crucial insights into the molecular epidemiology of tuberculosis in Morocco, emphasizing the importance of integrating genomic tools into national TB control programs. Understanding the genetic landscape and transmission pathways of M. tuberculosis will support more effective public health strategies aimed at reducing TB incidence and preventing the spread of drug-resistant strains.