Date Published: April 22, 2019
Publisher: Public Library of Science
Author(s): Chelsie E. Armbruster, Valerie S. Forsyth, Alexandra O. Johnson, Sara N. Smith, Ashley N. White, Aimee L. Brauer, Brian S. Learman, Lili Zhao, Weisheng Wu, Mark T. Anderson, Michael A. Bachman, Harry L. T. Mobley, Andreas J. Baumler.
The Gram-negative bacterium Proteus mirabilis is a common cause of catheter-associated urinary tract infections (CAUTI), which can progress to secondary bacteremia. While numerous studies have investigated experimental infection with P. mirabilis in the urinary tract, little is known about pathogenesis in the bloodstream. This study identifies the genes that are important for survival in the bloodstream using a whole-genome transposon insertion-site sequencing (Tn-Seq) approach. A library of 50,000 transposon mutants was utilized to assess the relative contribution of each non-essential gene in the P. mirabilis HI4320 genome to fitness in the livers and spleens of mice at 24 hours following tail vein inoculation compared to growth in RPMI, heat-inactivated (HI) naïve serum, and HI acute phase serum. 138 genes were identified as ex vivo fitness factors in serum, which were primarily involved in amino acid transport and metabolism, and 143 genes were identified as infection-specific in vivo fitness factors for both spleen and liver colonization. Infection-specific fitness factors included genes involved in twin arginine translocation, ammonia incorporation, and polyamine biosynthesis. Mutants in sixteen genes were constructed to validate both the ex vivo and in vivo results of the transposon screen, and 12/16 (75%) exhibited the predicted phenotype. Our studies indicate a role for the twin arginine translocation (tatAC) system in motility, translocation of potential virulence factors, and fitness within the bloodstream. We also demonstrate the interplay between two nitrogen assimilation pathways in the bloodstream, providing evidence that the GS-GOGAT system may be preferentially utilized. Furthermore, we show that a dual-function arginine decarboxylase (speA) is important for fitness within the bloodstream due to its role in putrescine biosynthesis rather than its contribution to maintenance of membrane potential. This study therefore provides insight into pathways needed for fitness within the bloodstream, which may guide strategies to reduce bacteremia-associated mortality.
The Gram-negative bacterium Proteus mirabilis commonly causes catheter-associated urinary tract infection (CAUTI), particularly in the elderly and in healthcare facilities [1–5]. Consequences of P. mirabilis CAUTI can include infection of the kidneys, urinary stone formation due to bacterial urease (urolithiasis), permanent renal damage, dissemination of bacteria to the bloodstream (bacteremia and/or sepsis), and possibly death [5–9]. In healthcare facilities including nursing homes, CAUTI is the most common cause of secondary bacteremia, which is associated with a one-year mortality rate of 10–13% in most settings, but can be as high as 66% [6, 7, 10, 11].
Proteus mirabilis HI4320 has been used as a model strain for decades to explore virulence determinants of this unusual bacterial species, particularly for urinary tract infection [2, 5]. With the availability of the complete genome sequence in 2008 , in vivo transcriptome assessment , and signature-tagged mutagenesis studies [34, 46, 47], much has been learned concerning how this organism adapts to the urinary tract and the virulence factors that contribute to ascending UTI [see [5, 48, 49] for review]. Our prior study extended this work by detailing the use of Tn-Seq for identification of fitness factors in a murine model of catheter-associated urinary tract infection (CAUTI), which uncovered genes essential for growth in rich medium and numerous previously unrecognized P. mirabilis fitness determinants, while also highlighting key differences in fitness requirements during ascending UTI versus CAUTI . Several P. mirabilis fitness factors identified through these studies have been tested for contribution to spleen colonization during secondary bacteremia, yet global fitness of P. mirabilis within the bloodstream has never been directly assessed. In the present study, we utilized Tn-Seq to elucidate fitness determinants for bloodstream infection versus factors that contribute to survival in serum in vitro. Through these efforts, we identified 143 candidate infection-specific fitness factors for survival of P. mirabilis within the bloodstream.