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Over the past few decades, extensively drug resistant (XDR) resistant Klebsiella pneumoniae has become a notable burden to healthcare all over the world. Especially carbapenemase-producing strains are problematic due to their capability to withstand even last resort antibiotics. Some sequence types (STs) of K. pneumoniae are significantly more prevalent in hospital settings in comparison to other equally resistant strains. This provokes the question whether or not there are phenotypic characteristics that may render certain K. pneumoniae more suitable for epidemic dispersal between patients, hospitals, and different environments. In this study, we selected seven epidemic and non-epidemic carbapenem resistant K. pneumoniae isolates for extensive systematic characterization for phenotypic and genotypic qualities in order to identify potential factors that precede or emerge from epidemic successfulness. Studied characteristics include growth rates and densities in different conditions (media, temperature, pH, resource levels), tolerance to alcohol and drought, inhibition between strains, ability to compensate pH, as well as various genomic features. Overall, there are clear differences between isolates, yet, only drought tolerance was found to notably associate with non-epidemic K. pneumoniae strains. We further report a preliminary study on the potential to control K. pneumoniae ST11 with an antimicrobial component produced by a non-epidemic K. pneumoniae. This component initially restricts bacterial growth, but stable resistance develops rapidly in vitro.

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BACKGROUND: Consumer-resource interactions constitute one of the most common types of interspecific antagonistic interaction. In natural communities, complex species interactions are likely to affect the outcomes of reciprocal co-evolution between consumers and their resource species. Individuals face multiple enemies simultaneously, and consequently they need to adapt to several different types of enemy pressures. In this study, we assessed how protist predation affects the susceptibility of bacterial populations to infection by viral parasites, and whether there is an associated cost of defence on the competitive ability of the bacteria. As a study system we used Serratia marcescens and its lytic bacteriophage, along with two bacteriovorous protists with distinct feeding modes: Tetrahymena thermophila (particle feeder) and Acanthamoeba castellanii (surface feeder). The results were further confirmed with another study system with Pseudomonas and Tetrahymena thermophila. RESULTS: We found that selection by protist predators lowered the susceptibility to infections by lytic phages in Serratia and Pseudomonas. In Serratia, concurrent selection by phages and protists led to lowered susceptibility to phage infections and this effect was independent from whether the bacteria shared a co-evolutionary history with the phage population or not. Bacteria that had evolved with phages were overall more susceptible to phage infection (compared to bacteria with history with multiple enemies) but they were less vulnerable to the phages they had co-evolved with than ancestral phages. Selection by bacterial enemies was costly in general and was seen as a lowered fitness in absence of phages, measured as a biomass yield. CONCLUSIONS: Our results show the significance of multiple species interactions on pairwise consumer-resource interaction, and suggest potential overlap in defending against predatory and parasitic enemies in microbial consumer-resource communities. Ultimately, our results could have larger scale effects on eco-evolutionary community dynamics.

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Lytic bacteriophages and protozoan predators are the major causes of bacterial mortality in natural microbial communities, which also makes them potential candidates for biological control of bacterial pathogens. However, little is known about the relative impact of bacteriophages and protozoa on the dynamics of bacterial biomass in aqueous and biofilm phases. Here, we studied the temporal and spatial dynamics of bacterial biomass in a microcosm experiment where opportunistic pathogenic bacteria Serratia marcescens was exposed to particle-feeding ciliates, surface-feeding amoebas, and lytic bacteriophages for 8 weeks, ca. 1300 generations. We found that ciliates were the most efficient enemy type in reducing bacterial biomass in the open water, but least efficient in reducing the biofilm biomass. Biofilm was rather resistant against bacterivores, but amoebae had a significant long-term negative effect on bacterial biomass both in the open-water phase and biofilm. Bacteriophages had only a minor long-term effect on bacterial biomass in open-water and biofilm phases. However, separate short-term experiments with the ancestral bacteriophages and bacteria revealed that bacteriophages crash the bacterial biomass dramatically in the open-water phase within the first 24 h. Thereafter, the bacteria evolve phage-resistance that largely prevents top-down effects. The combination of all three enemy types was most effective in reducing biofilm biomass, whereas in the open-water phase the ciliates dominated the trophic effects. Our results highlight the importance of enemy feeding mode on determining the spatial distribution and abundance of bacterial biomass. Moreover, the enemy type can be crucially important predictor of whether the rapid defense evolution can significantly affect top-down regulation of bacteria.

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The coincidental virulence evolution hypothesis suggests that outside-host selection, such as predation, parasitism and resource competition can indirectly affect the virulence of environmentally-growing bacterial pathogens. While there are some examples of coincidental environmental selection for virulence, it is also possible that the resource acquisition and enemy defence is selecting against it. To test these ideas we conducted an evolutionary experiment by exposing the opportunistic pathogen bacterium Serratia marcescens to the particle-feeding ciliate Tetrahymena thermophila, the surface-feeding amoeba Acanthamoeba castellanii, and the lytic bacteriophage Semad11, in all possible combinations in a simulated pond water environment. After 8 weeks the virulence of the 384 evolved clones were quantified with fruit fly Drosophila melanogaster oral infection model, and several other life-history traits were measured. We found that in comparison to ancestor bacteria, evolutionary treatments reduced the virulence in most of the treatments, but this reduction was not clearly related to any changes in other life-history traits. This suggests that virulence traits do not evolve in close relation with these life-history traits, or that different traits might link to virulence in different selective environments, for example via resource allocation trade-offs.

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