Physiologically distinct subpopulations formed in Escherichia coli cultures in response to heat shock.

Bacteria can form heterogeneous populations containing phenotypic variants of genetically identical cells. The heterogeneity of populations can be considered a bet-hedging strategy allowing adaptation to unknown environmental changes - at least some individual subpopulations or cells might be able to withstand future adverse conditions. Using Percoll gradient centrifugation, we demonstrated that in an Escherichia coli culture exposed to heat shock at 50 °C, two physiologically distinct subpopulations were formed. A high-density subpopulation (HD50) demonstrated continued growth immediately after its transfer to LB medium, whereas the growth of a low-density subpopulation (LD50) was considerably postponed. The LD50 subpopulation contained mainly viable but non-culturable bacteria and exhibited higher tolerance to sublethal concentrations of antibiotics or H2O2 than HD50 cells. The levels of aggregated proteins and main molecular chaperones were comparable in both subpopulations; however, a decreased number of ribosomes and a significant increase in protein oxidation were observed in the LD50 subpopulation as compared with the HD50 subpopulation. Interestingly, under anaerobic heat stress, the formation of the HD50 subpopulation was decreased and culturability of the LD50 subpopulation was significantly increased. In both subpopulations the level of protein aggregates formed under anaerobic and aerobic heat stress was comparable. We concluded that the formation of protein aggregates was independent of oxidative damage induced by heat stress, and that oxidative stress and not protein aggregation limited growth and caused loss of LD50 culturability. Our results indicate that heat stress induces the formation of distinct subpopulations differing in their ability to grow under standard and stress conditions.

Antibiotics promoting oxidative stress inhibit formation of Escherichia coli biofilm via indole signalling.

Recent studies have revealed that antibiotics can promote the formation of reactive oxygen species which contribute to cell death. In this study, we report that five different antibiotics known to stimulate production of reactive oxygen species inhibited growth of Escherichia coli biofilm. We demonstrated that supression of biofilm formation was mainly a consequence of the increase in the extracellular concentration of indole, a signal molecule which suppresses growth of bacterial biofilm. Indole production was enhanced under antibiotic-mediated oxidative stress due to overexpression of tryptophanase (TnaA), which catalyzes synthesis of indole. We found that DMSO (dimethyl sulfoxide), a hydrogen peroxide scavenger, or the lack of trypthophanase, which catalyzes production of indole, partly restored formation of E. coli biofilm in the presence of antibiotics. In conclusion, these findings confirmed that antibiotics which promote formation of ROS (reactive oxygen species) can inhibit development of E. coli biofilm in an indole-dependent process.