Two natural isolates of Agrobacterium tumefaciens contain a linear dicentric chromosome

Multipartite bacterial genomes offer advantages including coordinated gene regulation and faster genome replication but are challenging to maintain. Agrobacterium tumefaciens lineages often contain a circular chromosome (Ch1), a linear chromosome (Ch2), and multiple plasmids. We previously observed that in some stocks of the lab model strain C58, Ch1 and Ch2 were fused into a linear dicentric chromosome. Here we analyzed Agrobacterium natural isolates from the French soil collection and identified two strains with fused chromosomes. Chromosome conformation capture identified integration junctions that were different from the C58 fusion strain. Genome-wide DNA replication profiling showed both replication origins remained active. Transposon sequencing revealed that partitioning systems of both chromosome centromeres were essential. Importantly, the site-specific recombinases XerCD are required for the survival of the strains containing the fusion chromosome. Our findings show that replicon fusion happens in natural environments and balanced replication arm sizes and proper resolution systems enable the survival of such strains.