Global responses to oxytetracycline treatment in tetracycline-resistant Escherichia coli

Thea S B Møller,Kristian Thamsborg,John E Olsen,Morten O A Sommer,Mark G Poolman,Sisse Mortensen,Hassan B Hartman,Luca Guardabassi,Andreas E Johansen,Martin H Rau,Gang Liu

doi:10.1038/s41598-020-64995-1

Abstract

We characterized the global transcriptome of Escherichia coli MG1655:: tetA grown in the presence of ½ MIC (14 mg/L) of OTC, and for comparison WT MG1655 strain grown with 1//2 MIC of OTC (0.25 mg/L OTC). 1646 genes changed expression significantly (FDR > 0.05) in the resistant strain, the majority of which (1246) were also regulated in WT strain. Genes involved in purine synthesis and ribosome structure and function were top-enriched among up-regulated genes, and anaerobic respiration, nitrate metabolism and aromatic amino acid biosynthesis genes among down-regulated genes. Blocking of the purine-synthesis- did not affect resistance phenotypes (MIC and growth rate with OTC), while blocking of protein synthesis using low concentrations of chloramphenicol or gentamicin, lowered MIC towards OTC. Metabolic-modeling, using a novel model for MG1655 and continuous weighing factor that reflected the degree of up or down regulation of genes encoding a reaction, identified 102 metabolic reactions with significant change in flux in MG1655:: tetA when grown in the presence of OTC compared to growth without OTC. These pathways could not have been predicted by simply analyzing functions of the up and down regulated genes, and thus this work has provided a novel method for identification of reactions which are essential in the adaptation to growth in the presence of antimicrobials.

Highlights

Tetracycline drugs are bacteriostatic antimicrobials which bind reversibly to the bacterial ribosome and interfere with protein translation[1]
We have shown that a TetA-producing E. coli strain shows prolonged lag phase proportionally to the concentration of tetracycline in the growth medium, and that this concurs with increased expression of tetA10
Similar observations of cellular adaptation in antimicrobial resistant bacteria grown in the presence of the drug to which they are resistant have been described for Extended Spectrum Beta-Lactamase producing (ESBL) E. coli[11,12], methicillin resistant Staphylococcus aureus[12] and colistin-resistant Klebsiella pneumoniae[13], and interfering with these adaptive responses in certain cases re-sensitize the bacteria to the drugs[11,13]

Summary

Introduction

Tetracycline drugs are bacteriostatic antimicrobials which bind reversibly to the bacterial ribosome and interfere with protein translation[1]. The majority of the more than 40 genes encoding tetracycline resistance encode membrane-associated efflux proteins[7], which selectively pumps tetracycline from the cytosol to the periplasm in exchange for a proton[8]. We have shown that a TetA-producing E. coli strain shows prolonged lag phase proportionally to the concentration of tetracycline in the growth medium, and that this concurs with increased expression of tetA10. This suggests that expression of the resistance mechanism or incorporation of the efflux pump into the cell-membrane constitute a burden to the bacterium. Genes down trpE trpD mtr tdcB trpC dcuC nrfC nrfB nrfD trpB trpA gudX cadB nrfA tdcC garD narH apaG garL

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Conclusion