The evolutionary pattern of Pto DC3000 conducted in this study was the long-term daily transfer for 700 generations in KB medium. We have hypothesized that Pto DC3000 in our study may remove their flagella under evolution in the shaking environment, resulting from nutrition acquisition independent of its motility. Further, the mystery we want to reveal is what other adaptive mutations have been accumulated in this evolutionary trajectory, and the corresponding genotype to phenotype map. Whole-genome resequencing and proteomics, combined with biochemistry assays were performed in this study. The results showed that the flagella of Pto DC3000 indeed disappeared under our evolved conditions, and this phenotype conferred a fitness advantage to evolved populations. Correspondingly, mutations were detected in genes of the flagellar pathway. However, the other two genes, dksA and glnE, emerged as beneficial genetic changes. These two mutants contributed to the flagellar loss phenotype. Additionally, the evolved non-motile populations act as the hitchhiker to exploit flagellated WT bacteria, leading them to extend and direct motion to new environments. Our results not only expand the understanding of evolutionary tradeoffs for Pto DC3000 but also provide a more theoretical basis to improve the value of Pto DC3000 as a model microorganism.
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