Pseudomonas donghuensis P482 is a tomato rhizosphere isolate with the ability to inhibit growth of bacterial and fungal plant pathogens. Herein, we analysed the impact of the carbon source on the antibacterial activity of P482 and expression of the selected genes of three genomic regions in the P482 genome. These regions are involved in the synthesis of pyoverdine, 7-hydroxytropolone (7-HT) and an unknown compound (“cluster 17”) and are responsible for the antimicrobial activity of P482. We showed that the P482 mutants, defective in these regions, show variations and contrasting patterns of growth inhibition of the target pathogen under given nutritional conditions (with glucose or glycerol as a carbon source). We also selected and validated the reference genes for gene expression studies in P. donghuensis P482. Amongst ten candidate genes, we found gyrB, rpoD and mrdA the most stably expressed. Using selected reference genes in RT-qPCR, we assessed the expression of the genes of interest under minimal medium conditions with glucose or glycerol as carbon sources. Glycerol was shown to negatively affect the expression of genes necessary for 7-HT synthesis. The significance of this finding in the light of the role of nutrient (carbon) availability in biological plant protection is discussed.

Reverse transcription quantitative PCR (RT-qPCR), a method of choice for quantification of gene expression changes, requires stably expressed reference genes for normalization of data. So far, no reference genes were established for the Alphaproteobacteria of the genus Ochrobactrum. Here, we determined reference genes for gene expression studies in O. quorumnocens A44. Strain A44 was cultured under 10 different conditions and the stability of expression of 11 candidate genes was evaluated using geNorm, NormFinder and BestKeeper. Most stably expressed genes were found to be rho, gyrB and rpoD. Our results can facilitate the choice of reference genes in the related Ochrobactrum strains. O. quorumnocens A44 is able to inactivate a broad spectrum of N-acyl homoserine lactones (AHLs) – the quorum sensing molecules of many Gram-negative bacteria. This activity is attributed to AiiO hydrolase, yet it remains unclear whether AHLs are the primary substrate of this enzyme. Using the established RT-qPCR setup, we found that the expression of the aiiO gene upon exposure to two AHLs, C6-HLS and 3OC12-HSL, does not change above the 1-fold significance threshold. The implications of this finding are discussed in the light of the role of quorum sensing-interfering enzymes in the host strains.

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A fast, economical, and reproducible method for nanoparticle synthesis has been developed in our laboratory. The reaction is performed in an aqueous environment and utilizes light emitted by commercially available 1 W light-emitting diodes (λ =420 nm) as the catalyst. This method does not require nanoparticle seeds or toxic chemicals. The irradiation process is carried out for a period of up to 10 minutes, significantly reducing the time required for synthesis as well as environmental impact. By modulating various reaction parameters silver nanoparticles were obtained, which were predominantly either spherical or cubic. The produced nanoparticles demonstrated strong antimicrobial activity toward the examined bacterial strains. Additionally, testing the effect of silver nanoparticles on the human keratinocyte cell line and human peripheral blood mononuclear cells revealed that their cytotoxicity may be limited by modulating the employed concentrations of nanoparticles.

Pseudomonas donghuensis P482 is a tomato rhizosphere isolate with the ability to inhibit growth of bacterial and fungal plant pathogens. Herein, we analysed the impact of the carbon source on the antibacterial activity of P482 and expression of the selected genes of three genomic regions in the P482 genome. These regions are involved in the synthesis of pyoverdine, 7‑hydroxytropolone (7-HT) and an unknown compound (“cluster 17”) and are responsible for antimicrobial activity of P482. We showed that the P482 mutants, defective in these regions, show variations and contrasting patterns of growth inhibition of the target pathogen under given nutritional conditions (with glucose or glycerol as a carbon source). We also selected and validated the reference genes for gene expression studies in P. donghuensis P482. Amongst ten candidate genes, we found gyrB, rpoD and mrdA the most stably expressed. Using selected reference genes in RT-qPCR, we assessed the expression of the genes of interest under minimal medium conditions with glucose or glycerol as carbon sources. Glycerol was shown to negatively affect the expression of genes necessary for 7-HT synthesis. The significance of this finding in the light of the role of nutrient (carbon) availability in biological plant protection is discussed.