Throughout its lifecycle, Mycobacterium tuberculosis experiences numerous nutritional micro-environments and host-imposed stresses, demanding metabolic flexibility to survive. However, the physiological consequences of these fluctuating environments remain poorly understood. We have demonstrated carbon shifting dramatically affects mycobacterial susceptibility to stresses, including antibiotics. Whilst gluconeogenic carbon-sources generally confer protection, we show that antibiotic tolerance is “remembered”, following a shift to glycolytic conditions. This illustrates mycobacteria respond to antibiotics based on historical carbon conditions, demonstrating the phenomenon of mycobacterial memory. Combining single-cell, time-lapse microscopy, ¹³C-flux analysis, transcriptomics, and microbiological assays, we show mycobacterial memory is a metabolically driven, bet-hedging strategy to promote survival.