Genome-wide analysis of chromosomal import patterns after natural transformation of Helicobacter pylori. H. pylori genomes of endonuclease mutants generated by multiplex genome editing technology

Recombination plays a dominant role in the evolution of Helicobacter pylori, but its dynamics remain incompletely understood. We use an in vitro transformation system combined with genome sequencing to study chromosomal integration patterns after natural transformation. A single transformation cycle results in up to 21 imports and repeated transformations generate a maximum of 92 imports (8% sequence replacement). Import lengths show a bimodal distribution with averages of 28 and 1645 bp (micro/macro-imports). Reanalysis of paired genomes from chronically infected individuals demonstrates the same bimodal import pattern in vivo. Restriction endonucleases of the recipient fail to inhibit integration of homeologous DNA, independently of methylation. In contrast, restriction endonucleases limit the import of heterologous DNA. We conclude that restriction-modification systems inhibit the genomic integration of novel sequences, while they pose no barrier to homeologous recombination, which reconciles the observed stability of the H. pylori gene content and its highly recombinational population structure.