Genomic Profiling and Mutation Analysis of Mycobacterium bovis BCG Strains Causing Clinical Disease

Tuberculosis remains one of the most prevalent infectious diseases, and the only currently available vaccine is the Mycobacterium bovis bacillus Calmette-Guèrin (BCG) vaccine. The uncontrolled passaging of the BCG strain led to genetically diverse BCG strains. Seven samples from clinical BCG-associated disease were obtained from the National Tuberculosis Reference Laboratory. Whole-genome sequencing and bioinformatics analysis were performed using tools such as fastqc, Trimmomatic, and CLC Genomics Workbench 24.0.3 to obtain consensus sequences and analyse deletions between M. bovis AF2122/97, BCG Danish, and clinical samples. Snippy was used to generate the phylogenomic tree, Prokka for annotation, and an in-house script to detect potential drug resistance. Four deletions were identified between M. bovis wildtype and M. bovis BCG. The phylogenomic tree showed that of the seven strains analysed, one was phylogenetically close to M. tuberculosis H37Rv, and another to the Danish BCG vaccine. Other samples were distantly related to each other and to reference strains. Two of the samples showed possible resistance to ethambutol. This would imply original misdiagnosis of the disease and subsequent ineffective treatment. This study emphasises the importance of genomic testing for accurate diagnosis of BCG disease and underscores the need for phylogenomic surveillance of M. bovis BCG strains circulating in South Africa.