Reactive Oxygen Detoxification Contributes to Mycobacterium abscessus Antibiotic Survival

When a population of bacteria are exposed to a bactericidal antibiotic most cells die rapidly. However, a sub-population of antibiotic-tolerant cells known as "persister cells" can survive for prolonged periods, and antibiotic tolerance can be strongly induced in a population by stresses such as nutrient deprivation. However, the pathways required to maintain viability in this setting, and how they are regulated are poorly understood. To identify genetic determinants of antibiotic tolerance in mycobacteria, we carried out transposon mutagenesis insertion sequencing (Tn-Seq) screens in Mycobacterium abscessus (Mabs) exposed to bactericidal translation-inhibiting antibiotics. This analysis identified genes essential for the survival of both spontaneous persister cells as well as for stress-induced tolerance, allowing the first genetic comparison of these states in mycobacteria. Pathway analysis identified multiple genes involved in the detoxification of reactive oxygen species (ROS), including the catalase-peroxidase katG, which contributed to survival in both stressed and un-stressed populations. In addition, we found that endogenous ROS were generated following antibiotic exposure, and that hypoxia significantly impaired bacterial killing. Thus, in Mabs, the lethality of some antibiotics is amplified by toxic ROS accumulation, and antibiotic-tolerant cells require detoxification systems to remain viable.