Species and condition dependent mutational spectrum in experimentally evolved biofilms of Bacilli

Laboratory experimental evolution provides a powerful tool for studying microbial adaptation to different environments. To understand the differences and similarities of the dynamic evolutionary landscapes of two model species from the Bacillus genus as they adapt to abiotic and biotic surfaces, we revived the archived population samples from our four previous experimental evolution studies and performed longitudinal whole-population genome sequencing. Surprisingly, higher number of mutations, higher genotypic diversity, and higher evolvability were detected in the biotic conditions with smaller population size. Different adaptation strategies were observed in different environments within each species, with more diversified mutational spectrum detected in biotic conditions. The insertion sequences of Bacillus thuringiensis are critical for its adaptation to the plastic bead-attached biofilm environment, but insertion sequence mobility was a general phenomenon in this species independent of the selection condition. Additionally, certain parallel evolution has been observed across species and environments, particularly when two species adapt to the same environment at the same time. Further, the higher degree of genetic diversification observed in biotic selective environments indicates an increased spatial niche heterogeneity and a nutritional source difference created by the plant host that provided a strong strength of selection during the adaptation process. Together, these results provide the first comprehensive mutational landscape of two bacterial species’ biofilms that is adapted to an abiotic and biotic surface.