Antibiotic Efficacy in Escherichia coli and Klebsiella pneumoniae Under Nutrient Limitation and Effectiveness of Colistin-Based Antibiotic Combinations to Eradicate Persister Cells

Persister cells are responsible for recurrent or chronic infections resulting in antibiotic treatment failure. We aimed to investigate antibiotic efficacy in Escherichia coli and Klebsiella pneumoniae strains with limited metabolic activity. Bacterial cells cultured in nutrient-limited media showed characteristic persister phenotypes, including low intracellular ATP concentration, maintenance of antibiotic susceptibility, and an increase of (p)ppGpp levels. Amikacin showed no bactericidal activity under nutrient limitation conditions; however, metabolism-dependent ciprofloxacin exhibited metabolism-independent activity. The activity of colistin was metabolism-dependent, but it was retained under limited nutrient conditions. Nutrient limitation and antibiotic stress were related to the SOS response through recA expression in all four strains of E. coli and K. pneumoniae. However, the mRNA expression patterns of relA and spoT (associated with (p)ppGpp synthesis) and hpf and rpoS (downstream target genes of (p)ppGpp signaling) varied according to bacterial species, strain, and antibiotics, indicating diverse responses to nutrient stress in various persister cells. We also investigated the efficacy of antibiotic combinations to eradicate persister cells. As a result, colistin-based combinations were effective in the eradication of both E. coli and K. pneumoniae persister cells. In this study, persister cells were shown to be induced by metabolic stress, reducing antibiotic efficacy. We identified that combinations of colistin with amikacin or ciprofloxacin were effective to eliminate E. coli and K. pneumoniae persister cells.