An Engineered Variant of <i>E. coli</i> Nissle 1917 with Enhanced Transformation Efficiency and Robustness

The gut microbiome and its effects on human health have generated considerable scientific, veterinary, and medical interest in recent years. Several gut bacterial species have emerged as potential chassis organisms for the delivery of therapeutics in this milieu. Among these, <i>E. coli</i> Nissle 1917 (EcN), a nonpathogenic gut isolate bacterium, is quickly gaining popularity. However, a bottleneck in harnessing EcN's potential has been its poor transformation efficiency relative to other bacterial strains. In this study, we present the use of adaptive laboratory evolution to increase EcN's transformation efficiency by subjecting the strain to repeated cycles of electroporation and recovery. This new strain has been comprehensively characterized in comparison to the wild-type EcN, including assessments of growth under gut-mimicking duress conditions, permeability, motility, hydrophobicity, and plasmid replication. Since EcN is known to compete with pathogenic strains in the gut for iron, the competition dynamics and iron consumption of the strain were also significant factors to consider. Furthermore, we conducted genome sequencing and gene ontology enrichment analysis to identify affected genes and pathways to probe the potential mechanisms of the improved phenotype. Overall, the strain shows improved transformation efficiency and robustness while preserving its key biological functionality.