K1 Capsule-dependent phage-driven evolution in Escherichia coli leading to phage resistance and biofilm production.

Abstract

Understanding bacterial phage resistance mechanisms has implications for developing phage-based therapies. This study aimed to explore the development of phage resistance in Escherichia coli K1 isolates' to K1-ULINTec4, a K1-dependent bacteriophage. Resistant colonies were isolated from two different strains (APEC 45 and C5), both previously exposed to K1-ULINTec4. Genome analysis and several parameters were assessed, including growth capacity, phage adsorption, phenotypic impact at capsular level, biofilm production and virulence in the in-vivo Galleria mellonella larvae model. One out of the 6 resistant isolates exhibited a significantly slower growth rate suggesting the presence of a resistance mechanism altering its fitness. Comparative genomic analysis revealed insertion sequences in the region 2 of the kps gene cluster involved in the capsule biosynthesis. In addition, an immunoassay targeting the K1 capsule showed a very low positive reaction compared to the control. Nevertheless, microscopic images of resistant strains revealed the presence of capsules with a clustered organization of bacterial cells and biofilm assessment showed an increased biofilm production compared to the sensitive strains. In the G. mellonella model, larvae infected with phage-resistant isolates showed better survival rates than larvae infected with phage-sensitive strains. A phage resistance mechanism was identified at the genomic level and had a negative impact on the K1 capsule production. The resistant isolates showed an increased biofilm production, and a decreased virulence in vivo.