Light/Dark and Temperature Cycling Modulate Metabolic Electron Flow in Pseudomonas aeruginosa Biofilms

Sunlight drives phototrophic metabolism, which affects redox conditions and produces substrates for nonphototrophs. These environmental parameters fluctuate daily due to Earth's rotation, and nonphototrophic organisms can therefore benefit from the ability to respond to, or even anticipate, such changes. Circadian rhythms, such as daily changes in body temperature, in host organisms can also affect local conditions for colonizing bacteria. Here, we investigated the effects of light/dark and temperature cycling on biofilms of the opportunistic pathogen Pseudomonas aeruginosa PA14. We grew biofilms in the presence of a respiratory indicator dye and found that enhanced dye reduction occurred in biofilm zones that formed during dark intervals and at lower temperatures. This pattern formation occurred with cycling of blue, red, or far-red light, and a screen of mutants representing potential sensory proteins identified two with defects in pattern formation, specifically under red light cycling. We also found that the physiological states of biofilm subzones formed under specific light and temperature conditions were retained during subsequent condition cycling. Light/dark and temperature cycling affected expression of genes involved in primary metabolic pathways and redox homeostasis, including those encoding electron transport chain components. Consistent with this, we found that cbb₃-type oxidases contribute to dye reduction under light/dark cycling conditions. Together, our results indicate that cyclic changes in light exposure and temperature have lasting effects on redox metabolism in biofilms formed by a nonphototrophic, pathogenic bacterium. IMPORTANCE Organisms that do not obtain energy from light can nevertheless be affected by daily changes in light exposure. Many aspects of animal and fungal physiology fluctuate in response to these changes, including body temperature and the activities of antioxidant and other redox enzymes that play roles in metabolism. Whether redox metabolism is affected by light/dark and temperature cycling in bacteria that colonize such circadian organisms has not been studied in detail. Here, we show that growth under light/dark and temperature cycling lead to rhythmic changes in redox metabolism in Pseudomonas aeruginosa and identify proteins involved in this response. P. aeruginosa is a major cause of health care-associated infections and is designated a serious threat by the CDC due to its recalcitrance during treatments. Our findings have the potential to inform therapeutic strategies that incorporate controlled light exposure or consider P. aeruginosa's responses to conditions in the host.