Physiologic recovery of Mycobacterium tuberculosis from drug injury: A molecular study of post antibiotic effect in vitro and in vivo

Post-antibiotic effect (PAE) describes the delay in bacterial growth that continues after antibiotics are cleared. The physiologic basis of PAE in Mycobacterium tuberculosis (Mtb) remains poorly understood. Here, we evaluated the long-standing hypothesis that PAE reflects the time required for bacteria to recover from drug-induced physiologic damage by comparing Mtb after varying durations of treatment with the four-drug isoniazid, rifampin, pyrazinamide, ethambutol combination in vitro and in BALB/c mice using two novel molecular readouts of bacterial health. In aerobic axenic culture and in the high-dose aerosol mouse model, quantification of Mtb rRNA synthesis via the RS ratio and Mtb transcriptional profiling via SEARCH-TB revealed that longer drug exposure was assocated with greater injury and adaptation during treatment, as well as slower recovery after treatment, i.e., longer PAE. Recovery followed a conserved sequence, from resumption of rRNA synthesis, to broad transcriptional reprogramming, to eventual CFU change. In mice, however, physiologic recovery was markedly slower and less complete than in vitro, indicating longer PAE in the context of immunity. Our observation that PAE in Mtb depends on the duration of drug exposure and correlates with the degree of bacterial injury support the hypothesis that nonlethal physiologic damage contributes to PAE. Our observation that PAE of the standard TB regimen is longer in mice than in vitro indicates that immunity augments PAE for Mtb. Molecular evaluation of bacterial physiology provides a new basis for probing recovery from drug exposure and understanding PAE.