The stringent response enables () to shut down its replication and metabolism under various stresses. Here we show that lacking the stringent response enzyme Rel was unable to slow its replication rate during nutrient starvation. Metabolomics analysis revealed that the nutrient-starved -deficient strain had increased metabolism similar to that of exponentially growing wild-type bacteria in nutrient-rich broth， consistent with an inability to enter quiescence. Deficiency of increased the susceptibility of mutant bacteria to killing by isoniazid during nutrient starvation and in the lungs of chronically infected mice. We screened a pharmaceutical library of over 2 million compounds for inhibitors of Rel and showed that the lead compound X9 was able to directly kill nutrient-starved and enhanced the killing activity of isoniazid. Inhibition of Rel is a promising approach to target persisters， with the potential to shorten the duration of TB treatment.