Experimental Adaptation of a Free-Living Bacterium to the Zebrafish Digestive Tract

Animals have coexisted with an omnipresent and diverse array of bacteria for the entirety of their evolutionary history. As a result, symbioses between animals and bacteria are ubiquitous and can range from mutualism to parasitism. In particular, countless studies have demonstrated the pivotal role that bacteria residing in animal digestive tracts can play in determining animal health and well-being. However, it is still unknown how bacteria evolve the ability colonize animals. Due to the dramatic impacts that animals and bacteria can have on one another’s fitness, it is imperative to understand how symbioses between bacteria and their animal hosts originate. Therefore, to elucidate how bacteria evolve novel associations with vertebrate hosts, I serially passaged six replicate populations of a bacterial species with no prior known host associations (Shewanella oneidensis) through the digestive tracts of a model vertebrate, zebrafish (Danio rerio). After 20 passages through the digestive tracts of groups of larval zebrafish that were derived bacteria free (amounting to approximately 200 bacterial generations), I observed that all six replicate populations evolved to outcompete their unpassaged ancestor in terms of their ability to colonize larval guts. I subsequently sequenced the genomes of four evolved S. oneidensis isolates from each replicate population and found that their competitive advantage stemmed from two distinct classes of mutations that occurred in a mannose sensitive hemagglutinin pilus operon as well as in genes with putative diguanylate cyclase and phosphodiesterase domains. Both types of mutations enhanced bacterial motility, which was associated with increased representation in the aqueous portion of my experimental system and more efficient per capita immigration into zebrafish guts relative to the ancestral S. oneidensis reference strain. These increases in motility, were consistent with the behavior of a closely-related Shewanella species (Shewanella sp. ZOR0012) that has recently been isolated from the zebrafish digestive tract implying that my evolved isolates may be pursuing a similar adaptive trajectory to the one taken by this host-associated species. My results suggest that a non-host-associated microorganism can rapidly improve its ability to colonize hosts, and this study is the first to capture the early adaptive steps necessary to facilitate this transition