527.5 - Klebsiella pneumoniae utilizes intestinal mucus to increase fitness in the gastrointestinal tract

Taylor Ticer (Medical University of South Carolina, Medical University of South Carolina), Terri Ellis (University of North Florida), Melinda Engevik (Medical University of South Carolina, Medical University of South Carolina)

Background: Klebsiella pneumoniae is a growing concern in the healthcare setting, as carbapenem-resistant Enterobacteriaceae are considered an urgent health threat by the Center of Disease Control (CDC). K. pneumoniae is found in the mucosa of the human gastrointestinal tract, but upon intestinal disruption K. pneumoniae can migrate into the blood stream and cause infection. One of the barriers against bacterial translocation is the intestinal mucus layer. Mucus proteins are covered with branching glycans which can serve as a fuel source for microbes that harbor mucus-specific glycosyl hydrolases. Hypothesis: We hypothesized that K. pneumoniae could degrade and utilize mucus as an energy source to increase its fitness in the GI tract. Methods amp;

Results: Using the Carbohydrate-Active enZYme Database (CAZY), we identified that 274 K. pneumoniae genomes harbored at least 3 mucin-degrading glycosyl hydrolase families. To confirm the ability of K. pneumoniae to degrade mucus, we grew five commercially available K. pneumoniae strains and six clinical isolates of K. pneumoniae in a chemically defined minimal media (CDMM) lacking glucose supplemented with mucin-associated oligosaccharides or dialyzed porcine intestinal MUC2 mucus. All K. pneumoniae strains grew well is sialic acid, GalNAc, fucose and galactose. Interestingly, K. pneumoniae strains exhibited robust growth during log phase with glucose, but were unable to sustain long term growth. Surprisingly, some clinical isolates showed similar growth with GluNAC as with glucose, while other strains could not utilize GluNAC as an energy source. All but one strain of K. pneumoniae could grow with dialyzed mucus as the sole carbon source, indicating that K. pneumoniae can enzymatically degrade mucus and utilize the liberated sugars for growth. To determine how mucus influenced the pathogenesis of K. pneumoniae, we grew one commercially available and one clinical isolate of K. pneumoniae in CDMM with fucose or mucus, isolated the RNA and examined the expression of genes associated with K. pneumoniae virulence by qPCR. We examined virulence factors related to iron uptake (iroD), nucleic acid metabolism (allS), endotoxin production (clbA), fatty acid biosynthesis (clbB), type 1- fimbriae (fimH), and flagella (fliC). Both K. pneumoniae strains had increased iroD expression in the presence of mucus. Interestingly, we observed that the clinical isolate, but not the commercially available strain of K. pneumoniae had increased expression of virulence genes clbB, allS and fliC when grown with mucus when compared to being grown with fucose. K. pneumoniae is traditionally considered a nonmotile bacterium, however, we confirmed the expression of flagella in our clinical isolate by performing a chemotaxis assay using mucus-associated sugars and mucus. We indeed saw that our commercially available strain was unable to migrate towards the mucus or mucus-associated sugars. However, our clinical isolate was able to swim towards mucus-associated sugars.

Conclusions: These results suggest that K. pneumoniae utilizes intestinal mucus as an energy source in the GI tract, which in turn influences the pathogenicity of clinical isolates of K. pneumoniae.

T32:1T32DK124191-01A