In vitro models that utilize hypoxia to induce non-replicating persistence in Mycobacteria

The Wayne model and Rapid Anaerobic Dormancy model are widely used methods to analyze the response of Mycobacterium tuberculosis to hypoxia and anaerobiosis. In these models tubercle bacilli are grown in sealed tubes in which bacilli aerobic respiration produces a temporal oxygen gradient. The gradual depletion of oxygen results in a non-replicating persistent culture capable of extended microaerobic and anaerobic survival. Here we describe both models used to induce hypoxic non-replicating persistence in M. tuberculosis. Additional techniques such as the isolation of RNA, the detection of nitrate reductase activity and ATP levels, and the determination of the NAD(+)/NADH ratio are described.