Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 May 26;11(1):10994.
doi: 10.1038/s41598-021-90488-w.

The carbon source-dependent pattern of antimicrobial activity and gene expression in Pseudomonas donghuensis P482

Marta Matuszewska  1 , Tomasz Maciąg  1 , Magdalena Rajewska  1 , Aldona Wierzbicka  1 , Sylwia Jafra  2
Affiliations

The carbon source-dependent pattern of antimicrobial activity and gene expression in Pseudomonas donghuensis P482

Marta Matuszewska et al. Sci Rep. .

Abstract

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.

PubMed Disclaimer

Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1
Figure 1
Growth inhibition of Dickeya solani IFB0102 (a) and Pseudomonas syringae pv. syringae Pss762 (b) by Pseudomonas donghuensis P482 mutants tested on minimal M9-agar medium with 0.4% glucose or 0.4% glycerol as a sole carbon source. The bars represent the percentage of the growth inhibition zone obtained for P482 wt under given conditions. Pseudomonas vranovensis DSM16006T does not cause growth inhibition of the tested pathogens and was used as a negative control strain (see Supplementary Data Figure S1). The assay was performed in triplicates; error bars represent standard deviation.
Figure 2
Figure 2
RT-qPCR reference gene selection for Pseudomonas donghuensis P482. (a) Boxplot representing the distribution of Cq (threshold cycle) data among the tested potential reference genes. The plot was calculated from raw data consisting of technical replicates’ mean Cq for each sample. The band in the box represents the median value, the top edge of the box is the upper quartile (Q3) while the bottom edge of the box is the lower quartile (Q1). Q3 and Q1 referred to the 75th percentile and the 25th percentile, respectively, meaning that 75 or 25% of the data were at or below the point. The whiskers represent the maximum and minimal values excluding outliers. Outlier data is presented with black diamond symbol (♦). (b) RefFinder comprehensive stability value (CSV) calculated as a geometric mean of the ranks assigned to the tested RGs by algorithms comprising RefFinder tool. The lower the CSV, the more stable the expression of a given gene. (c) qbase + geNorm RG stability analysis. Average expression stability of tested RGs obtained with geNorm algorithm shown as geNorm M value. The lower the M value, the more stable the gene expression. (d) Determination of the optimal number of reference targets shown as qbase + geNorm V chart for the tested reference targets. Analysis shows no significant difference in experimental situation when comparing the use of 3 or 4 reference genes (geNorm V < 0.15 for V3/4), meaning 3 reference genes are sufficient for expression normalisation.
Figure 3
Figure 3
Comparison of relative expression (scaled to the mean CNRQ value calculated for each gene) of the chosen P. donghuensis P482 genes for bacteria cultured in the presence of glucose or glycerol as a sole carbon source in minimal medium M9. Error bars represent the 95% confidence interval (CI ± 95%). Statistically significant change in expression related to the particular carbon source used was observed for genes (the expression fold change value is given in brackets): 4709 (39.91), 4706 (16.01), 4705 (4.87), 3318 (2.6). Statistical analysis was performed using Student’s t-test, *) p < 0.05 **) p < 0.03. For clarity of the figure, loci references are represented by numbers only.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Gottschalk G. Bacterial Metabolism. Springer; 1986.
    1. Dagley S. Chemical unity and diversity in bacterial catabolism. In: Poindexter JS, Leadbetter ER, editors. Bacteria in Nature. Plenum Press; 1989. pp. 259–329.
    1. Drew SW, Demain AL. Effect of primary metabolites on secondary metabolism. Annu. Rev. Microbiol. 1977;31:343–356. doi: 10.1146/annurev.mi.31.100177.002015. - DOI - PubMed
    1. Görke B, Stülke J. Carbon catabolite repression in bacteria: Many ways to make the most out of nutrients. Nat. Rev. Microbiol. 2008;6:613–624. doi: 10.1038/nrmicro1932. - DOI - PubMed
    1. Ruiz B, et al. Production of microbial secondary metabolites: Regulation by the carbon source. Crit. Rev. Microbiol. 2010;36:146–167. doi: 10.3109/10408410903489576. - DOI - PubMed

MeSH terms

LinkOut - more resources