Genomic and proteomic analyses of the maize root isolate <i>Rhodococcus erythropolis</i> NI86/21 reveal extensive genome plasticity and parallel evolution of herbicide degradation

<i>Rhodococcus erythropolis</i> NI86/21, isolated from maize rhizosphere in Hungary, possesses one of the largest genomes (8.046 Mb) within the species. The genome comprises a 6.83 Mb chromosome and 1.22 Mb of extrachromosomal elements, including three circular and two fragmented linear plasmids. Comparative analysis identified five horizontally acquired genomic islands (HGTi), totaling 0.64 Mb with mosaic-like architecture derived from plasmids, phages, and chromosomal segments of other <i>Nocardiaceae</i>. Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic analysis revealed a lower expression of genes located in HGT elements (53%) compared to core chromosomal genes (73%), indicating regulatory silencing of foreign DNA. Nevertheless, an inducible cytochrome P450 monooxygenase (CYP116) responsible for thiocarbamate and atrazine degradation is encoded on HGTi_V and actively expressed upon herbicide exposure. Strikingly, an identical CYP450 locus is present on a conjugative plasmid in <i>Rhodococcus</i> sp. TE1 isolated from thiocarbamate-treated soil in Canada, demonstrating independent acquisition of the same catabolic module from a high GC% content <i>Rhodococcus</i>, under similar selective pressure. Frequent recombination between chromosomal and mobile elements generates the observed mosaic-like HGT structures, which we found common for <i>R. erythropolis</i> strains. These results highlight extraordinary genomic plasticity and rapid adaptive evolution in <i>Rhodococci</i>, enabling efficient colonization of herbicide-contaminated agro-ecosystems.IMPORTANCE<i>Rhodococcus erythropolis</i> NI86/21 exemplifies how bacterial genomes evolve through horizontal gene transfer and mobile elements. Its unusually large, plastic genome contains extensive HGT islands and a high load of active transposons, which shape mosaic genomic architectures and hinder complete genome assembly. These horizontally acquired regions, although partially silenced, encode key adaptive functions such as an inducible CYP116 monooxygenase enabling thiocarbamate and atrazine degradation. Remarkably, an identical CYP116 module is present in <i>Rhodococcus</i> sp. TE1 from thiocarbamate-treated Canadian soil, demonstrating that similar environmental pressures can drive independent acquisition of the same biodegradation trait. Together, the dynamic transposon activity, mosaic HGT structure, and geographically convergent gene recruitment highlight the extraordinary genomic plasticity of <i>R. erythropolis</i> and underscore its rapid adaptive potential in agro-ecosystems, with implications for microbial evolution and bioremediation strategies.