Bacterial physiology, regulation and mutational adaptation in a chemostat environment.

The chemostat was devised over 50 years ago and rapidly adopted for studies of bacterial physiology and mutation. Despite the long history and earlier analyses, the complexity of events in continuous cultures is only now beginning to be resolved. The application of techniques for following regulatory and mutational changes and the identification of mutated genes in chemostat populations has provided new insights into bacterial behaviour. Inoculation of bacteria into a chemostat culture results in a population competing for a limiting amount of a particular resource. Any utilizable carbon source or ion can be a limiting nutrient and bacteria respond to limitation through a regulated nutrient-specific hunger response. In addition to transcriptional responses to nutrient limitation, a second regulatory influence in a chemostat culture is the reduced growth rate fixed by the dilution rate in individual experiments. Sub-maximal growth rates and hunger result in regulation involving sigma factors and alarmones like cAMP and ppGpp. Reduced growth rate also results in increased mutation frequencies. The combination of a strongly selective environment (where mutants able to compete for limiting nutrient have a major fitness advantage) and elevated mutation rates (both endogenous and through the secondary enrichment of mutators) results in a population that changes rapidly and persistently over many generations. Contrary to common belief, the chemostat environment is never in "steady state" with fixed bacterial characteristics usable for clean comparisons of physiological or regulatory states. Adding to the complexity, chemostat populations do not simply exhibit a succession of mutational sweeps leading to a dominant winner clone. Instead, within 100 generations large populations become heterogeneous and evolving bacteria adopt alternative, parallel fitness strategies. Transport physiology, metabolism and respiration, as well as growth yields, are highly diverse in chemostat-evolved bacteria. The rich assortment of changes in an evolving chemostat provides an excellent experimental system for understanding bacterial evolution. The adaptive radiation or divergence of populations into a collection of individuals with alternative solutions to the challenge of chemostat existence provides an ideal model system for testing evolutionary and ecological theories on adaptive radiations and the generation of bacterial diversity.