Polymicrobial Lysis-Hi-C experiments of three bacteria and plasmids derived from HiC-Pro

Microbes interact in biofilms and polymicrobial infections in a spatially structured manner, yet next-generation sequencing approaches generally fail to recover in situ spatial proximity between distinct genotypes in the interactome or eDNA matrix. This study explored a modified Hi-C approach involving an initial lysis phase prior to DNA cross-linking, to test whether adjacent cell chromatin can be cross-linked, anticipating that this could provide a new avenue for the study of spatial-mutational dynamics in structured microbial communities. An artificial polymicrobial mixture of Pseudomonas aeruginosa, Staphylococcus aureus, and Escherichia coli was lysed for 1 to 18 hours, then prepared for Hi-C. A murine biofilm infection model was treated with sonication, mechanical lysis, or chemical lysis before Hi-C. Bioinformatic analyses of resulting Hi-C interspecies chromatin links showed that while microbial species differed from one another, generally lysis significantly increased links bet...