We previously reported that Escherichia coli strains carrying a firefly luciferase reporter gene (luc+) showed a posttranslationally-generated bioluminescence burst upon entry into the stationary phase. In this paper, we studied the mechanism underpinning this burst by using a series of "Keio" gene deletion strains. When luc+ driven by the lac gene promoter (lacp::luc+) was introduced into a group of Keio strains, the resulting reporter strains showed significantly altered timing and/or sizes of the burst. Remarkably, a reporter strain that lacked phosphoglucose isomerase (PGI), which catalyzes the second step of glycolysis, showed no burst, while the onset of the stationary phase of this strain was the same as that of the wild-type (WT) reporter strain. Consistently, the WT reporter strain showed no burst, when grown on arabinose or xylose instead of glucose as the carbon source. These results suggest that a process in carbohydrate metabolism is involved in the mechanism of generation of the burst. We measured temporal changes in intracellular NADPH concentrations but could not detect a significant increase or decrease relative to the occurrence of the burst. Functional implications and possible applications of the burst are discussed. (C) 2020 Elsevier Inc. All rights reserved.