DescriptionMycobacterium abscessus (Mab) infections are on the rise and have increased adverse outcomes in cystic fibrosis (CF) patients worldwide over the past two decades. Mab is the most notoriously difficult to treat nontuberculous mycobacterium (NTM) due to its resistance to a wide variety of antibiotics and disinfectants. Mab clearance rates are exceptionally low even after aggressive and lengthy antibiotic regiments. Because Mab has the innate ability to evade antibiotic treatment we proposed Mab may enlist toxin-antitoxin (TA) systems to induce a persistent state enabling the evasion of antibiotic killing. Here we identified a total of 21 novel putative TA systems in 128 Mab clinical strains available on NCBI via sequence similarity to Mycobacterium tuberculosis (Mtb). The aim of this work was to identify the specific targets of three putative VapBC TA systems and characterize their physiological effects in Mab. In this study we determined the mechanism of action of Mab VapC5, VapC1, and VapC2. Each toxin cleaves multiple tRNAs at a single site within their anticodon resulting in a general reduction of translation. However, VapC5 and VapC1 drastically reduce translation capacity while specifically upregulating the translation of ribosomal proteins. For VapC5, this proteomic reprogramming results in an increase in persister cell formation after treatment with two first line Mab antibiotics, amikacin (AMK) and cefoxitin (FOX). Therefore, we theorize that VapC1 and VapC5 toxins trigger a phenotypic shift that enables Mab cells to persist upon antibiotic treatment because they 1) lead to a growth arrested state which reduces the efficacy of antibiotics that act on actively growing cells, and 2) selectively overproduce ribosomes to override the efficacy of antibiotics that inactivate the ribosome. These findings provide a foundation for the selection of shorter and more efficacious alternate treatment options for Mab infections using currently available antimicrobials whose targets are not subverted by these toxins.