Ninety isolates of root nodule bacteria from an invasive Mimosa pigra population in Australia were characterized by PCR assays and by sequencing of ribosomal genes. All isolates belonged to the same bacterial genus (Burkholderia) that predominates on M. pigra in its native geographic range in tropical America. However, the Australian Burkholderia strains represented several divergent lineages, none of which had a close relationship to currently known Burkholderia strains in American M. pigra populations. Inoculation of M. pigra with Australian strains resulted in equal or higher plant growth and nodule nitrogenase activity (measured by acetylene reduction assays) relative to outcomes with bacteria from M. pigra's native geographic region. The main difference in symbiotic phenotype for bacteria from the two regions involved responses to an alternate Mimosa host species: Central American strains failed to fix nitrogen in association with Mimosa pudica, while most Australian Burkholderia isolates tested had high nodule nitrogenase activity in association with both Mimosa species. Invasive M. pigra populations in Australia have therefore acquired a diverse assemblage of nodule bacteria that are effective nitrogen-fixing symbionts, despite having a broader host range and a distant genetic relationship to bacterial strains found in the plant's ancestral region.