Disentangling the impacts of abiotic and biotic environmental factors and dispersal dynamics on the pangenome fluidity of bacterial pathogens

ABSTRACT Understanding how pangenomes originate and evolve is crucial for predicting evolutionary trajectories and uncovering ecological interactions of bacterial pathogens. Pangenome fluidity has been attributed to adaptive evolution, yet the underlying ecological drivers for bacterial pathogens persisting in natural reservoirs remain poorly understood. Listeria monocytogenes (Lm), a foodborne pathogen causing fatal listeriosis, serves as an ideal model for investigating the ecological mechanisms underlying pangenome fluidity in bacterial pathogens due to its high evolutionary divergence, broad ecological versatility, and significant public health concern. Through pangenome analysis of 177 Lm isolates representing three evolutionary lineages (I, II, and III) that we isolated from soils across the United States, we found that substantial genome variation was strongly associated with climatic factors (e.g. precipitation and temperature), soil properties (e.g. aluminum, pH, and molybdenum), and bacterial community composition, particularly Nitrospirae, Planctomycetes, Acidobacteria, and Cyanobacteria. These factors exerted selective pressure across many gene functions, with pronounced effects on genes involved in cell envelope synthesis, defense mechanisms, and replication, recombination, and repair. Among Lm lineages occupying varied habitats, distinct pangenome properties were observed. Lineage III exhibited a highly fluid pangenome, which was attributed to local adaptation to nutrient-limited conditions and strong dispersal limitation. In contrast, lineage I maintained a conserved pangenome, likely due to frequent homogenizing dispersal. Consistent with these dispersal patterns, we identified an elevated risk of soil-to-human transmission in lineage I, evidenced by epidemiological links between three soil-derived and 17 clinical isolates. Collectively, this study reveals the pivotal role of environmental selection imposed by both abiotic factors and bacterial communities in governing the adaptive pangenome evolution in bacterial pathogens. It also highlights significant differences in pangenome flexibility, ecological niches, and transmission dynamics across lineages of the same pathogen species, underscoring the need for tailored source tracking strategies. AUTHOR SUMMARY Studying the full set of genes found in different strains of a bacterium (i.e. pangenome) helps us understand how bacterial pathogens develop and adapt to changes in the environment. Here, we focused on Listeria monocytogenes (Lm), a pathogen capable of spreading through food and surviving in diverse environments, to understand how environmental factors and the way that bacteria move across locations can influence the pangenome content in this important bacterium. By analyzing the genomes of 177 Lm strains representing three evolutionary lineages (I, II, and III) collected from soils across the United States, we found that variation in climate, soil chemistry, and surrounding bacteria (e.g., Nitrospirae) was closely linked to genetic differences among strains. These environmental conditions seemed to affect genes that help build the cell envelop, protect the bacteria from harm, and fix damaged DNA. We also observed different levels of genome flexibility across Lm lineages which were found to be related to how they move across different locations. Lineage III showed evidence of barriers to spreading, which may enhance genetic differentiation across populations, leading to a more flexible pangenome. In contrast, lineage I appeared to spread more readily and was epidemiologically linked to human clinical cases, which may facilitate genetic exchange that reduce pangenome diversity. This study shows that both non-living environmental conditions—like precipitation and pH—and nearby groups of bacteria play a big role in shaping how bacterial pathogens change their genes to survive. It also highlights that different subtypes of the same pathogen can have different gene flexibility and spread in different ways, calling for specific biocontrol measures. Competing Interest Statement The authors have declared no competing interest. Footnotes The title, abstract, and discussion have been updated.