Extensive spontaneous genome reduction in Paraburkholderia sabiae.

Paraburkholderia sabiae LMG24235T is a beta-proteobacterium originally isolated from Mimosa caesalpiniifolia nodules that efficiently outcompetes phytopathogens both in vitro and in planta. Here we show that P. sabiae LMG24235T can spontaneously lose a 2.3-Mbp replicon, resulting in a reduction of genome size by approximately one quarter and representing the first reported case of a bacterium losing 23% of its genome in a single event. Phylogenetic analysis of the replication initiation protein A (RepA) revealed that this replicon is closely related to megaplasmids found in other Paraburkholderia strains and a randomly barcoded transposon mutagenesis (RB-TnSeq) analysis showed that it contains very few predicted likely-essential genes, all of which are dispensable for growth. In contrast to the wild-type strain, P. sabiae lacking the second replicon (Δ2) exhibited a growth defect on D-trehalose and L-leucine, reduced motility and decreased survival under low-oxygen conditions. While growth in rich medium was otherwise unaffected, P. sabiae Δ2 was outcompeted by the wild type in vitro, and M. caesalpiniifolia nodules occupied by Δ2 fixed 40% less nitrogen compared to wild-type nodules. These results suggest that the second replicon is not an essential chromosomal element but rather a dispensable large megaplasmid that confers a competitive advantage to P. sabiae LMG24235T in its natural environment. The online version contains supplementary material available at 10.1038/s41598-026-49026-9.