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Sublethal Concentrations of Antibiotics Cause Shift to Anaerobic Metabolism in Listeria monocytogenes and Induce Phenotypes Linked to Antibiotic Tolerance.

Knudsen GM, Fromberg A, Ng Y, Gram L - Front Microbiol (2016)

Bottom Line: A mutant in the bifunctional acetaldehyde-CoA/alcohol dehydrogenase encoded by lmo1634 did not have altered antibiotic tolerance.However, a mutant in lmo1179 (eutE) encoding an aldehyde oxidoreductase where rerouting caused increased ethanol production was tolerant to three of four antibiotics tested.Thus, sublethal concentrations of antibiotics caused metabolic and physiological changes indicating that the organism is preparing to withstand lethal antibiotic concentrations.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, Technical University of Denmark Kongens Lyngby, Denmark.

ABSTRACT
The human pathogenic bacterium Listeria monocytogenes is exposed to antibiotics both during clinical treatment and in its saprophytic lifestyle. As one of the keys to successful treatment is continued antibiotic sensitivity, the purpose of this study was to determine if exposure to sublethal antibiotic concentrations would affect the bacterial physiology and induce antibiotic tolerance. Transcriptomic analyses demonstrated that each of the four antibiotics tested caused an antibiotic-specific gene expression pattern related to mode-of-action of the particular antibiotic. All four antibiotics caused the same changes in expression of several metabolic genes indicating a shift from aerobic to anaerobic metabolism and higher ethanol production. A mutant in the bifunctional acetaldehyde-CoA/alcohol dehydrogenase encoded by lmo1634 did not have altered antibiotic tolerance. However, a mutant in lmo1179 (eutE) encoding an aldehyde oxidoreductase where rerouting caused increased ethanol production was tolerant to three of four antibiotics tested. This shift in metabolism could be a survival strategy in response to antibiotics to avoid generation of ROS production from respiration by oxidation of NADH through ethanol production. The monocin locus encoding a cryptic prophage was induced by co-trimoxazole and repressed by ampicillin and gentamicin, and this correlated with an observed antibiotic-dependent biofilm formation. A monocin mutant (ΔlmaDCBA) had increased biofilm formation when exposed to increasing concentration of co-trimoxazole similar to the wild type, but was more tolerant to killing by co-trimoxazole and ampicillin. Thus, sublethal concentrations of antibiotics caused metabolic and physiological changes indicating that the organism is preparing to withstand lethal antibiotic concentrations.

No MeSH data available.


Related in: MedlinePlus

Killing of L. monocytogenes EGD by (A) gentamicin, (B) co-trimoxazole, (C) tetracycline, and (D) ampicillin. An early stationary phase culture (16 h at 37°C) was diluted to OD600 = 0.1 and exposed to either MilliQ or different concentrations of the four antibiotics. The concentrations are given relative to the concentration of antibiotic used for the transcriptomic analysis (1X), i.e., 50X (for gentamicin and co-trimoxacole), 100X (for all four antibiotics), and 1000X (for ampicillin and tetracycline). The experiment was performed with three biological replicates and error bar are standard deviation.
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Figure 2: Killing of L. monocytogenes EGD by (A) gentamicin, (B) co-trimoxazole, (C) tetracycline, and (D) ampicillin. An early stationary phase culture (16 h at 37°C) was diluted to OD600 = 0.1 and exposed to either MilliQ or different concentrations of the four antibiotics. The concentrations are given relative to the concentration of antibiotic used for the transcriptomic analysis (1X), i.e., 50X (for gentamicin and co-trimoxacole), 100X (for all four antibiotics), and 1000X (for ampicillin and tetracycline). The experiment was performed with three biological replicates and error bar are standard deviation.

Mentions: We assessed the quantitative differences in bactericidal/bacteriostatic action of the four antibiotics to investigate if there was a relationship between the degree of shift in metabolic gene expression and the level of bactericidal action (Supplementary Figure S3). We tested 100-fold higher concentrations than used in the transcriptomic analyses. Gentamicin and co-trimoxazole caused a 5 and 3.4-log reduction in bacterial count after 12 h (Figures 2A,B), respectively, whereas, ampicillin and tetracycline caused less than 1-log reduction after 12 h (Figures 2C,D). After 72 h the reduction in bacterial count was 2.3-log for tetracycline and more than 4-log for ampicillin, indicating a bactericidal action of ampicillin after prolonged treatment. Thus the level of killing by the four antibiotics after 12 h reflected the degree of lmo1634-expression at 3 h exposure to sublethal concentration of antibiotics.


Sublethal Concentrations of Antibiotics Cause Shift to Anaerobic Metabolism in Listeria monocytogenes and Induce Phenotypes Linked to Antibiotic Tolerance.

Knudsen GM, Fromberg A, Ng Y, Gram L - Front Microbiol (2016)

Killing of L. monocytogenes EGD by (A) gentamicin, (B) co-trimoxazole, (C) tetracycline, and (D) ampicillin. An early stationary phase culture (16 h at 37°C) was diluted to OD600 = 0.1 and exposed to either MilliQ or different concentrations of the four antibiotics. The concentrations are given relative to the concentration of antibiotic used for the transcriptomic analysis (1X), i.e., 50X (for gentamicin and co-trimoxacole), 100X (for all four antibiotics), and 1000X (for ampicillin and tetracycline). The experiment was performed with three biological replicates and error bar are standard deviation.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4940397&req=5

Figure 2: Killing of L. monocytogenes EGD by (A) gentamicin, (B) co-trimoxazole, (C) tetracycline, and (D) ampicillin. An early stationary phase culture (16 h at 37°C) was diluted to OD600 = 0.1 and exposed to either MilliQ or different concentrations of the four antibiotics. The concentrations are given relative to the concentration of antibiotic used for the transcriptomic analysis (1X), i.e., 50X (for gentamicin and co-trimoxacole), 100X (for all four antibiotics), and 1000X (for ampicillin and tetracycline). The experiment was performed with three biological replicates and error bar are standard deviation.
Mentions: We assessed the quantitative differences in bactericidal/bacteriostatic action of the four antibiotics to investigate if there was a relationship between the degree of shift in metabolic gene expression and the level of bactericidal action (Supplementary Figure S3). We tested 100-fold higher concentrations than used in the transcriptomic analyses. Gentamicin and co-trimoxazole caused a 5 and 3.4-log reduction in bacterial count after 12 h (Figures 2A,B), respectively, whereas, ampicillin and tetracycline caused less than 1-log reduction after 12 h (Figures 2C,D). After 72 h the reduction in bacterial count was 2.3-log for tetracycline and more than 4-log for ampicillin, indicating a bactericidal action of ampicillin after prolonged treatment. Thus the level of killing by the four antibiotics after 12 h reflected the degree of lmo1634-expression at 3 h exposure to sublethal concentration of antibiotics.

Bottom Line: A mutant in the bifunctional acetaldehyde-CoA/alcohol dehydrogenase encoded by lmo1634 did not have altered antibiotic tolerance.However, a mutant in lmo1179 (eutE) encoding an aldehyde oxidoreductase where rerouting caused increased ethanol production was tolerant to three of four antibiotics tested.Thus, sublethal concentrations of antibiotics caused metabolic and physiological changes indicating that the organism is preparing to withstand lethal antibiotic concentrations.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, Technical University of Denmark Kongens Lyngby, Denmark.

ABSTRACT
The human pathogenic bacterium Listeria monocytogenes is exposed to antibiotics both during clinical treatment and in its saprophytic lifestyle. As one of the keys to successful treatment is continued antibiotic sensitivity, the purpose of this study was to determine if exposure to sublethal antibiotic concentrations would affect the bacterial physiology and induce antibiotic tolerance. Transcriptomic analyses demonstrated that each of the four antibiotics tested caused an antibiotic-specific gene expression pattern related to mode-of-action of the particular antibiotic. All four antibiotics caused the same changes in expression of several metabolic genes indicating a shift from aerobic to anaerobic metabolism and higher ethanol production. A mutant in the bifunctional acetaldehyde-CoA/alcohol dehydrogenase encoded by lmo1634 did not have altered antibiotic tolerance. However, a mutant in lmo1179 (eutE) encoding an aldehyde oxidoreductase where rerouting caused increased ethanol production was tolerant to three of four antibiotics tested. This shift in metabolism could be a survival strategy in response to antibiotics to avoid generation of ROS production from respiration by oxidation of NADH through ethanol production. The monocin locus encoding a cryptic prophage was induced by co-trimoxazole and repressed by ampicillin and gentamicin, and this correlated with an observed antibiotic-dependent biofilm formation. A monocin mutant (ΔlmaDCBA) had increased biofilm formation when exposed to increasing concentration of co-trimoxazole similar to the wild type, but was more tolerant to killing by co-trimoxazole and ampicillin. Thus, sublethal concentrations of antibiotics caused metabolic and physiological changes indicating that the organism is preparing to withstand lethal antibiotic concentrations.

No MeSH data available.


Related in: MedlinePlus