Genome-Wide Identification of Pseudomonas aeruginosa Genes Important for Desiccation Tolerance on Inanimate Surfaces

Pseudomonas aeruginosa is an opportunistic pathogen prevalent in the environment and in health care settings. Transmission in the health care setting occurs through human-human interactions and/or contact with contaminated surfaces. Moist surfaces such as respirators, sink and tub drains, and even disinfectants can serve as reservoirs. Dry surfaces such as plastic and stainless steel could also serve as a reservoir but would necessitate some degree of tolerance to desiccation. Using an assay to measure P. aeruginosa tolerance to desiccation on plastic and stainless-steel surfaces, we found that only 0.05 to 0.1% of the desiccated cells could be recovered 24 h postdesiccation. We took advantage of the strong selection imposed by desiccation to identify genes important for tolerance using Tn-seq. A highly saturated Tn-seq library was desiccated on plastic and stainless-steel surfaces. NexGen sequencing of the recovered cells identified 97 genes important for survival. Comparing cells desiccated under low- and high-nutrient conditions allowed for differentiation of genes important for desiccation tolerance. The 53 genes identified in the latter analysis are involved in maintenance of cell envelope integrity, purine and pyrimidine biosynthesis, tricarboxylic acid (TCA) cycle, and the hydrolysis of misfolded proteins. The Tn-seq findings were validated by competition experiments with wild-type (WT) cells and select Tn insertion mutants. Mutants lacking carB and surA demonstrated the largest fitness defects, indicating that pyrimidine biosynthesis and outer membrane integrity are essential for desiccation tolerance. Increased understanding of desiccation tolerance could provide insight into approaches to control environmental reservoirs of P. aeruginosa. IMPORTANCE Health care-associated infections (HAIs) caused by Pseudomonas aeruginosa result in significant morbidity and mortality and are a significant economic burden. Moist environments that promote biofilm formation are an important reservoir for P. aeruginosa. Dry environments may also serve as a reservoir but would require some degree of desiccation tolerance. Here, we took a genome-wide approach to identify genes important for desiccation tolerance on plastic and stainless-steel surfaces. Genes involved in assembly of outer membrane proteins and pyrimidine biosynthesis were particularly important. Strains lacking these functions were unable to tolerate surface desiccation. These findings suggest that inhibitors of these pathways could be used to prevent P. aeruginosa survival on dry surfaces.