We assessed the genomic and phenotypic landscape of fitness-impaired E.coli lineages recovering from the deleterious effects of Muller’s ratchet via a combination of experimental evolution, genomics and global phenotypic screening. We found a seemingly paradoxical scenario where protein function appears to worsen in lineages as fitness increases. However, further investigation via functional perturbation analyses revealed that the deleterious effects of mutations that accumulated under the ratchet are probably being mitigated through novel molecular mechanisms – this gives the superficial impression that protein function is worsening. We argue that compensatory evolution explains this novelty, and we suspect compensatory mutations drive a substantial proportion of fitness and protein function recovery in the evolving lineages.

Additionally, based on the Biolog colour change assay, which measures substrate-dependent respiration, and rates-based measurements collected through time, we observed rapid and widespread metabolic erosion in populations subjected to the ratchet. Moreover, we found that phenotypic impairment was not mitigated following a period of fitness recovery.

The results presented here suggest that both the genomic and phenotypic landscape for lineages recovering from the effects of Muller’s ratchet are novel compared to that of ancestral lineages. We comment on implications of these results with respect to various ideas on the emergence of biological complexity, and offer suggestions for future work. To our knowledge, this is the first report of how mutational changes that accumulate in recovering populations impact on protein function, and no other study to date has assessed the capacity for recovering populations to grow in a very broad range of environments.