Abstract:

Objective: To induce Mycobacterium abscessus strain ATCC 19977 resistant to sudapyridine (WX-081) and to perform the functional annotation of resistance-associated single nucleotide polymorphism (SNP) along with related pathways. Methods: The standard strain Mycobacterium abscessus ATCC19977 was subjected to incrementally increasing WX-081 exposure in liquid medium to select resistant mutants. Genomic DNA was extracted from the WX-081-resistant strains for whole-genome sequencing to identify SNP. Sequence alignment was performed using BLAST against multiple databases, such as evolutionary genealogy of genes: non-supervised orthologous (eggNOG), gene ontology (GO), kyoto encyclopedia of genes and genomes (KEGG), non-redundant (Nr), protein families (Pfam), swiss-prot protein sequence (swiss-prot) for functional annotation, followed by GO, eggNOG, and KEGG pathway enrichment analyses. Results: Comparative genomic analysis identified 187 SNP in coding regions between the parental ATCC19977 and WX-081-resistant strains, of which 62 met quality thresholds. Among these, 43 were homozygous SNP (12 synonymous and 31 missense mutations). SNP in the coding regions accounted for 9.14% (187/2047), of the total, with stop-gained variants (nonsense mutations) being the least frequent (0.05%, 1/2047). In contrast, 90.86% (1860/2047) of SNP were located in non-coding regions, predominantly in downstream (51.83%, 1061/2047) and upstream (39.03%, 799/2047) regions.GO annotation identified 8 biological processes (BP), 5 cellular components (CC), and 4 molecular functions (MF). eggNOG functional prediction classified the variant genes as follows: 7 with unknown function, 3 involved in amino acid transport and metabolism, 1 in nucleotide transport and metabolism, 2 in lipid transport and metabolism, 1 in transcription, 2 in cell wall/membrane and/envelope biogenesis, 1 in post-translational modification, protein turnover, chaperones, 2 in inorganic ion transport and metabolism, 2 in secondary metabolite biosynthesis, transport and catabolism, and 1 in signal transduction mechanisms. KEGG pathway analysis identified 1 pathway each related to membrane transport, amino acid metabolism, and carbohydrate metabolism, 3 pathways in global and overview maps, and 4 pathways associated with lipid metabolism. Conclusion: The genome of WX-081-resistant Mycobacterium abscessus exhibited predominantly non-coding SNP, while 31 missense mutations in coding regions may directly contribute to drug resistance. Metabolic pathway analysis revealed significant enrichment of genes related to lipid metabolism, cell wall synthesis, and amino acid transport, suggesting that membrane/wall structural remodeling and metabolic reprogramming are key resistance mechanisms.

Key words: Mycobacterium, Sudapyridine, Drug resistance, Polymorphism, single nucleotide