Although millions of plant growth–promoting bacteria (PGPBs) are currently used in agriculture and soil health restoration, their potential environmental threats have largely been ignored. Here, rhizosphere soils from Chilean temperate rainforests (CTR) were collected and studied as an attractive, novel source of potentially safer PGPBs. A total of 847 isolates was assembled, and representative PGPB phenotypes and biocontrol (BC) activities were screened. Forty isolates that exhibited most of the tested PGPB phenotypes and BC activities were selected, and their antibiotic resistance (AR) and presumptive virulence factors were assayed. Of these, the 4 rhizosphere isolates with the lowest AR and virulence values were selected, designated ‘potentially safer PGPBs’ and inoculated into tomato seeds. Our results showed that between 86 and 100% of the rhizosphere-collection isolates exhibited at least one of the tested PGPB phenotypes, while 26.9 to 40.7% showed in vitro antagonistic activity against at least one of the tested plant pathogens. Low percentages of AR (<40%) and presumptive virulence (<22%) were also observed among the 40 selected isolates exhibiting the PGPB phenotypes and BC activities. 16S rRNA gene sequencing of the selected ‘potentially safer PGPBs’ provided only a preliminary, genus–level affiliation of these isolates; application of isolates affiliated with the genera Ochrobactrum , Brucella , Microbacterium and Bacillus resulted in significantly ( p ≤ 0.05) greater tomato ( Solanum lycopersicum ) seedling growth compared with uninoculated controls. These results of our study were obtained in a short–term in vitro seedling screening assay and indicate that CTR rhizosphere soils merit further evaluation as a source of potentially safer PGPB candidates showing a low in vitro risk profile, in line with the ‘Food Security’ and ‘One Health’ approaches.

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