Safety attributes of Pseudomonas sp. P26, an environmental microorganism with potential application in contaminated environments

Abstract

Currently, the selection of non-pathogenic microorganisms that lack clinically relevant antimicrobial resistance is crucial to bioaugmentation strategies. Pseudomonas sp. P26 (P26) is an environmental bacterium of interest due to its ability to remove aromatic compounds from petroleum, but its safety characteristics are still unknown. The study aimed to: a) determine P26 sensitivity to antimicrobials, b) investigate the presence of quinolone and β-lactam resistance genes, c) determine the presence of virulence factors, and d) evaluate the effect of P26 on the viability of Galleria mellonella (an invertebrate animal model). P26 antimicrobial sensitivity was determined in vitro using the Kirby-Bauer agar diffusion method and the VITEK 2 automated system (BioMerieux®). Polymerase Chain Reaction was employed for the investigation of genes associated with quinolone resistance, extended-spectrum β-lactamases, and carbapenemases. Hemolysin and protease production was determined in human blood agar and skimmed-milk agar, respectively. In the in vivo assay, different doses of P26 were injected into Galleria mellonella larvae and their survival was monitored daily. Control larvae injected with Pseudomonas putida KT2440 (a strain considered as safe) and Pseudomonas aeruginosa PA14 (a pathogenic strain) were included. Pseudomonas sp. P26 was susceptible to most evaluated antimicrobials, except for trimethoprim-sulfamethoxazole. No epidemiologically relevant genes associated with quinolone and β-lactam resistance were identified. Hemolysin and protease production was only evidenced in the virulent strain (PA14). Furthermore, the results obtained in the in vivo experiment demonstrated that inocula less than 10⁸ CFU/mL of P26 and P. putida KT2440 did not significantly affect larval survival, whereas larvae injected with the lowest dose of the pathogenic strain P. aeruginosa PA14 experienced instant mortality. The results suggest that Pseudomonas sp. P26 is a safe strain for its application in environmental bioremediation processes. Additional studies will be conducted to ensure the safety of this bacterium against other organisms.

Keywords: Antimicrobial resistance; Environmental bacteria; Galleria mellonella; Infective capacity.