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. 2020 Jan-Feb;34(1):65-71.
doi: 10.21873/invivo.11746.

The Role of Efflux Pumps and Environmental pH in Bacterial Multidrug Resistance

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The Role of Efflux Pumps and Environmental pH in Bacterial Multidrug Resistance

Márta Nové et al. In Vivo. 2020 Jan-Feb.

Abstract

Background/aim: One of the most studied bacterial resistance mechanisms is the resistance related to multidrug efflux pumps. In our study the pump activity of the Escherichia coli K-12 AG100 strain expressing the AcrAB-TolC pump system was investigated at pH 7 and pH 5 in the presence of the efflux pump inhibitor (EPI) promethazine (PMZ).

Materials and methods: The EPI activity was assessed by real-time fluorimetry. The influence of PMZ treatment on the relative expression of the pump genes acrA, acrB and their regulators marA, marB, marR, the stress genes soxS, rob, as well as the bacterial growth control genes ftsI, and sdiA were determined by RT-qPCR.

Results: The EPI activity of PMZ was more effective at neutral pH. The PMZ treatment induced a significant stress response in the bacterium at acidic pH by the up-regulation of genes.

Conclusion: The genetic system that regulates the activity of the main efflux pump is pH-dependent.

Keywords: Escherichia coli K-12 AG100; Multidrug resistance; efflux pump; efflux pump genes; pH-dependent; promethazine.

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Conflict of interest statement

None declared.

Figures

Figure 1
Figure 1. The growth curves of E. coli K-12 AG100 strain were determined at pH 5 (A, B) and pH 7 (C, D) in LB broth in the presence and absence of 25 μg/ml of PMZ. Graphs A and C show the growth curves by measuring the optical density (OD600) at pH 5 and pH 7, respectively. Graphs B and D show the growth curves by counting the CFUs at pH 5 and pH 7, respectively. The growth of PMZ treated bacterial culture was slower at pH 5 compared to pH 7. The growth of bacterial culture was more rapid at pH 7, moreover after 8 h of culturing the declination phase of the bacterial culture could be detected at pH 7 (Graph D).
Figure 1
Figure 1. The growth curves of E. coli K-12 AG100 strain were determined at pH 5 (A, B) and pH 7 (C, D) in LB broth in the presence and absence of 25 μg/ml of PMZ. Graphs A and C show the growth curves by measuring the optical density (OD600) at pH 5 and pH 7, respectively. Graphs B and D show the growth curves by counting the CFUs at pH 5 and pH 7, respectively. The growth of PMZ treated bacterial culture was slower at pH 5 compared to pH 7. The growth of bacterial culture was more rapid at pH 7, moreover after 8 h of culturing the declination phase of the bacterial culture could be detected at pH 7 (Graph D).
Figure 2
Figure 2. Accumulation of EB at pH 5 and pH 7 by E. coli K-12 AG100 in the presence and absence of glucose 0.4%, with and without 25 μg/ml of PMZ. The real-time accumulation curves demonstrated a higher intracellular EB concentration without glucose at pH 7 compared to pH 5. The intracellular concentration of EB was significantly higher in the presence of PMZ at pH 7, in addition the PMZ treated sample exhibited lower EB accumulation at pH 5. The calculation of statistical significance and p-value was based on the relative fluorescence index (RFI) of the given sample. The correlation is significant: p≤0.001 and p=0.005.
Figure 3
Figure 3. Relative expression of genes involved in stress response on E. coli K-12 AG100 strain in the presence of 25 μg/ml of PMZ at pH 5 and pH 7 at different time points (1-18 h). At the beginning of the culturing period most of the studied genes showed a decreased expression pattern. Increase in gene expression level was detected in the cases of the acrA and B, marR, soxS, sdiA genes after the 18th hour at pH 5. Significant gene expression could be observed in the expression levels of acrA, acrB, and marA genes in the 18th hour, of marB in the 1st hour and of ftsI after 4 h. Initially the efflux pump genes acrA and acrB were down-regulated, but at the end of the culturing period (18th hour) both genes were up-regulated at pH 7. The significant correlation is: p≤0.05.

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References

    1. Weston N, Sharma P, Ricci V, Piddock LJV. Regulation of the AcrAB-TolC efflux pump in Enterobacteriaceae. Res Microbiol. 2018;169(7-8):425–431. PMID: 29128373. DOI: 10.1016/j.resmic.2017.10.005. - PubMed
    1. Nikaido H, Takatsuka Y. Mechanisms of RND multidrug efflux pumps. Biochim Biophys Acta. 2009;1794(5):769–781. PMID: 19026770. DOI: 10.1016/j.bbapap.2008.10.004. - PMC - PubMed
    1. Sun J, Deng Z, Yan A. Bacterial multidrug efflux pumps: Mechanisms, physiology and pharmacological exploitations. Biochem Biophys Res Commun. 2014;453(2):254–267. PMID: 24878531. DOI: 10.1016/j.bbrc.2014.05.090. - PubMed
    1. Wang Z, Fan G, Hryc CF, Blaza JN, Serysheva II, Schmid MF, Chiu W, Luisi BF, Du D. An allosteric transport mechanism for the AcrAB-TolC multidrug efflux pump. eLife. 2017:e24905. PMID: 28355133. DOI: 10.7554/eLife.24905. - PMC - PubMed
    1. Pos KM. Drug transport mechanism of the AcrB efflux pump. Biochim Biophys Acta. 2009;1794(5):782–793. PMID: 19166984. DOI: 10.1016/j.bbapap.2008.12.015. - PubMed

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