Error-prone translesion synthesis (TLS) is important for the de novo development of antibiotic resistance in bacteria. Unfortunately, little is known regarding the mechanics of this relationship. Recent evidence suggests that mutation supply (population size x mutation rate) is the bottleneck in resistance development and there are, in principle, three main ways in which TLS polymerases could contribute to an increase in mutation supply. First, they could contribute to the genetic diversity present in the bacterial population, prior to selection. Secondly, their activity could promote DNA damage tolerance, thus increasing the size of the population prior to selection. Lastly, these error-prone polymerases could be integral to increasing the frequency of mutations once cells come in contact with antibiotics.
It has previously been impossible to tease out these mechanism subtleties using traditional agar based end-point assays; however, we hypothesize that spatially resolved experimental evolution techniques can be used to distinguish between these three mechanism possibilities. To this end, we have constructed four-channel microfluidic devices in which cells evolve up a continuous gradient of the SOS-inducing antibiotic ciprofloxacin. By competing TLS-defective and -competent cells, from sub-inhibitory through to inhibitory concentrations, we aim to determine what role each of the TLS polymerases play in the resistance evolution process. To date our observations support an adaptive mutation model by which TLS polymerases promote resistance by increasing the frequency of mutations upon contact with ciprofloxacin.