Spread of biological species from primary into novel habitats leads to within-species adaptive niche differentiation and is commonly driven by acquisition of point mutations in individual genes that increase fitness in the alternative environment. However, finding footprints of adaptive niche differentiation in specific genes remains a challenge. Here we describe a novel method to analyze the footprint of pathogenicity-adaptive, or pathoadaptive, mutations in the Escherichia coli gene encoding FimH-the major, mannose-sensitive adhesin. Analysis of distribution of mutations across the nodes and branches of the FimH phylogenetic network shows (1) zonal separation of evolutionary primary structural variants of FimH and recently derived ones, (2) dramatic differences in the ratio of synonymous and nonsynonymous changes between nodes from different zones, (3) evidence for replacement hot-spots in the FimH protein, (4) differential zonal distribution of FimH variants from commensal and uropathogenic E. coli, and (5) pathoadaptive functional changes in FimH brought by the mutations. The selective footprint in fimH indicates that the pathoadaptive niche differentiation of E. coli is either in its initial stages or undergoing an evolutionary "source/sink" dynamic.