Effects of Cell-to-Cell Variability in MarA Expression on Escherichia coli Antibiotic Resistance

Traditionally, bacterial populations are viewed as individual cells that share both genotypic and phenotypic identity. Consequently, when individual cells within a population demonstrate elevated antibiotic resistance levels this was assumed to be the result of genetic mutation. However, recent studies have successfully demonstrated that there exist disparities between genotypic identity and phenotypic identity. For instance, studies have shown that, as a mechanism to evade the lethal effects of the bactericidal drugs, bacteria cells are capable of switching between a regular drug susceptible state and a drug tolerant dormant state (persister) by stochastic phenotypic variation. In addition, recent work by our group has suggested that transient antibiotic resistance can stem from stochastic pulsing in genes related to antibiotic resistance. In particular, we are focusing on a transcription factor, MarA from Escherichia coli. MarA is known to play an essential role for the regulation of more than 40 downstream antibiotic resistance genes, such as the AcrAB-TolC efflux pump genes and micF gene. In our study, we aim to demonstrate that MarA expression is heterogeneous for each individual cells within an isogenic bacterial population. We hypothesize that differences in MarA expression may render individual bacteria with different antibiotic resistant levels. To achieve this goal, we are exposing wild-type E. coli cells to various antibiotics. Plasmids that carry a fluorescence protein gene controlled by a marA promoter are used as reporters for MarA expression.  In an effort to attain a precisely controlled external environment, we are using a microfluidic chip to control addition of antibiotics and have coupled this with time-lapse microscopy. Our preliminary data indicate that there are differences in expression of fluorescence proteins between individual cells within a microcolony, which reflects noisy MarA expression. In addition, when antibiotics are added, cells demonstrate different antibiotic resistance levels, which correlated with the noisy MarA expression. Hence, the isogenic bacteria population could use this mechanism as a “bet-hedging” strategy, which could eventually lead to a new bacteria population with pre-adapted antibiotic resistance ability.