Adaptation to novel spatially-structured environments is driven by the capsule and alters virulence-associated traits

Abstract

The extracellular capsule is a major virulence factor, but its ubiquity in free-living bacteria with large environmental breadths suggests that it shapes adaptation to novel niches. Yet, how it does so, remains unexplored. Here, we evolve three Klebsiella strains and their capsule mutants in parallel. Their comparison reveals different phenotypic and genotypic evolutionary changes that alter virulence-associated traits. Non-capsulated populations accumulate mutations that reduce exopolysaccharide production and increase biofilm formation and yield, whereas most capsulated populations become hypermucoviscous, a signature of hypervirulence. Hence, adaptation to novel environments primarily occurs by fine-tuning expression of the capsular locus. The same evolutionary conditions selecting for mutations in the capsular gene wzc leading to hypermucoviscosity also result in increased susceptibility to antibiotics by mutations in the ramA regulon. This implies that general adaptive processes outside the host can affect capsule evolution and its role in virulence and infection outcomes may be a by-product of such adaptation.

Competition between lysogenic and sensitive bacteria is determined by the fitness costs of the different emerging phage-resistance strategies

“…A major finding of our work is that the dominant type of the resistance mechanisms changes across evolutionary time scales. At first, resistance emerged quickly to the loss of capsule production because functional inactivation can be achieved in many ways ( Chiarelli et al, 2020 ; Haudiquet et al, 2021 ; Nucci et al, 2022 ; Supplementary file 2 ). At later stages, resistance may emerge by changes in the composition or expression of the capsule, which can outcompete both lysogens and non-capsulated clones.…”

Latent evolution of biofilm formation depends on life-history and genetic background

“…We had previously observed that the presence or absence of the capsule drives the direction and magnitude of evolutionary change in endpoint populations [32], yet how it affects the evolutionary dynamics remained to be tested. Initial changes in biofilm formation were similar across both capsule genotypes (Figure 1A).…”

“…Because non-capsulated clones readily emerge in capsulated populations [23,32], we also included the proportion of capsulated clones in the generalized linear model. This revealed that the changes in biofilm formation is also associated with the frequency of capsulated clones (P< 0.0001).…”

“…This can have important consequences, for instance, in host colonization or increased tolerance to antibiotics. To study how biofilm formation in evolving populations changes through time, we took advantage of a previous evolution study in which we evolved in parallel six populations from three different strains of the K. pneumoniae species complex, and their respective non-capsulated mutants (Figure S1) [32]. We did so in five different liquid static environments, including artificial sputum (ASM) and urine (AUM), varying in carrying capacity (nutrient availability).…”

“…We tested how these two traits evolved throughout our evolution experiment (Figure S2BC, Table S1). Unfortunately, population yield in some capsulated populations could not be tested due to the emergence of hypermucoviscosity, which precludes accurate CFU assessment [32]. On average, evolving populations significantly increased yield (x̄ = +60%, One-sample Wilcoxon Rank Sum test, P< 0.001) (Figure S2B, Table S1), but as observed for biofilm formation, most changes occurred early during the evolution experiment.…”