Cultivation in Long-term Simulated Microgravity is Detrimental to Biofilm Formation Ability and Pyocyanin Production of Pseudomonas aeruginosa PA14 Wild Type, ΔflgK, and ΔpelA

Pseudomonas aeruginosa can attach to surfaces and form aggregates known as biofilms. It has been previously found that P. aeruginosa cultivated in space form thicker and structurally different biofilms than those grown in Earth gravity. The purpose of our study was to investigate how microgravity, simulated in a laboratory setting, would influence the biofilm formation abilities of P. aeruginosa PA14 wild type strain as well as mutants ΔflgK and ΔpelA. While ΔflgK is defective in the initialization of biofilm formation, ΔpelA is hindered in biofilm growth and maintenance. The bacteria were cultivated in a High Aspect Ratio Vessel (HARV) on a Rotary Cell Culture System (RCCS) that was used to simulate microgravity. For the Earth gravity control cultures, the RCCS was oriented horizontally and cultures were rotated in HARVS around a vertical axis. Incubation time was six days, and, in contrast to studies done by others, the bacteria were allowed to grow into stationary phase without replenishment of culture medium. At the end of the incubation time, the bacteria were extracted and cultured in a 24-well plate under identical conditions in Earth gravity. After 24 hours, the robustness of biofilm formation was compared by removing the soluble culture from the wells, staining with crystal violet, solubilizing the remaining biofilm, and quantifying spectrophotometrically. Additionally, the concentration of pyocyanin produced during cultivation was determined through extraction and spectrophotometry. Cultures grown under simulated microgravity had a lower biofilm formation ability as well as lower pyocyanin production compared to those grown under Earth gravity. This raises the possibility that P. aeruginosa experiencing nutritional starvation under long-term simulated microgravity may become less virulent