Adaptive Divergence in Experimental Populations of Pseudomonas fluorescens

7 8 9 3 ABSTRACT 1 Pseudomonas fluorescens is a model for the study of adaptive radiation. When propagated in a 2 spatially structured environment the bacterium rapidly diversifies into a range of niche specialist 3 genotypes. Here we present a genetic dissection and phenotypic characterization of the " fuzzy 4 spreader " (FS) morphotype — a type that arises repeatedly during the course of the P. fluorescens 5 radiation and which appears to colonize the bottom of static broth microcosms. The causal 6 mutation is located within gene fuzY (pflu0478) — the fourth gene of the five gene fuzVWXYZ 7 operon. fuzY encodes a β-glycosyltransferase that is predicted to modify lipopolysaccharide (LPS) 8 O-antigens. The effect of the mutation is to cause cell flocculation. Analysis of 92 independent FS 9 genotypes showed each to have arisen as the result of a loss-of-function mutation in fuzY, although 10 different mutations have subtly different phenotypic and fitness effects. Mutations within fuzY were 11 previously shown to suppress the phenotype of mat-forming " wrinkly spreader " (WS) types. This 12 prompted a re-investigation of FS niche preference. Time-lapse photography showed that FS 13 colonizes the meniscus of broth microcosms forming cellular rafts that, being too flimsy to form a 14 mat, collapse to the vial bottom, and then repeatably reform only to collapse. This led to a 15 reassessment of the ecology of the P. fluorescens radiation. Finally, we show that ecological 16 interactions between the three dominant emergent types (SM, WS and FS), combined with the 17 interdependence of FS and WS on fuzY can, at least in part, underpin an evolutionary arms race with 18 bacteriophage SBW25Φ2, to which mutation in fuzY confers resistance.