A Gram-negative, non-sporulating, rod-shaped, motile bacterium, with a single polar flagellum, designated strain PsJNT, was isolated from surface-sterilized onion roots. This isolate proved to be a highly effective plant-beneficial bacterium, and was able to establish rhizosphere and endophytic populations associated with various plants. Seven related strains were recovered from Dutch soils. Based on 16S rRNA gene sequence data, strain PsJNT and the Dutch strains were identified as representing a member of the genus Burkholderia, as they were closely related to Burkholderia fungorum (98·7 %) and Burkholderia phenazinium (98·5 %). Analysis of whole-cell protein profiles and DNA–DNA hybridization experiments confirmed that all eight strains belonged to a single species. Strain PsJNT had a DNA G+C content of 61·0 mol%. Only low levels of DNA–DNA hybridization to closely related species were found. Qualitative and quantitative differences in fatty acid composition between strain PsJNT and closely related species were identified. The predominant fatty acids in strain PsJNT were 16 : 0, 18 : 17c and summed feature 3 (comprising 16 : 17c and/or iso-15 : 0 2-OH). Isolate PsJNT showed high 1-aminocyclopropane-1-carboxylate deaminase activity and is therefore able to lower the ethylene level in a developing or stressed plant. Production of the quorum-sensing signal compound 3-hydroxy-C8-homoserine lactone was detected. Based on the results of this polyphasic taxonomic study, strain PsJNT and the seven Dutch isolates are considered to represent a single, novel species, for which the name Burkholderia phytofirmans sp. nov. is proposed. The type strain is strain PsJNT (=LMG 22146T=CCUG 49060T).