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Metabolic regulation of mycobacterial growth and antibiotic sensitivity.

Baek SH, Li AH, Sassetti CM - PLoS Biol. (2011)

Bottom Line: This pathway plays a causal role in reducing growth and antibiotic efficacy by redirecting cellular carbon fluxes away from the tricarboxylic acid cycle.Mutants in which this metabolic switch is disrupted are unable to arrest their growth in response to stress and remain sensitive to antibiotics during infection.Thus, this regulatory pathway contributes to antibiotic tolerance in vivo, and its modulation may represent a novel strategy for accelerating TB treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.

ABSTRACT
Treatment of chronic bacterial infections, such as tuberculosis (TB), requires a remarkably long course of therapy, despite the availability of drugs that are rapidly bacteriocidal in vitro. This observation has long been attributed to the presence of bacterial populations in the host that are "drug-tolerant" because of their slow replication and low rate of metabolism. However, both the physiologic state of these hypothetical drug-tolerant populations and the bacterial pathways that regulate growth and metabolism in vivo remain obscure. Here we demonstrate that diverse growth-limiting stresses trigger a common signal transduction pathway in Mycobacterium tuberculosis that leads to the induction of triglyceride synthesis. This pathway plays a causal role in reducing growth and antibiotic efficacy by redirecting cellular carbon fluxes away from the tricarboxylic acid cycle. Mutants in which this metabolic switch is disrupted are unable to arrest their growth in response to stress and remain sensitive to antibiotics during infection. Thus, this regulatory pathway contributes to antibiotic tolerance in vivo, and its modulation may represent a novel strategy for accelerating TB treatment.

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Metabolic modulation reverses the antibiotic tolerance induced by low                            iron and hypoxic conditions.Bacterial survival in the presence of the indicated antibiotics under                            hypoxic conditions (A, C, E, G) and in low iron media (B, D, F, H).                            Isoniazid (“INH”, 2 and 0.25 µg ml−1,                            A and B), streptomycin (“SMP”, 2 and 1 µg                                ml−1, C and D), ciprofloxacin (“CIP”, 4                            and 1 µg ml−1, E and F), and ethambutol                            (“EMB”, 5 and 3 µg ml−1, G and H)                            were introduced into each culture. Antibiotics were added to the hypoxic                            vials after 14 d of culture. Means ± SD of two independent                            experiments each performed in duplicate are shown.
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pbio-1001065-g003: Metabolic modulation reverses the antibiotic tolerance induced by low iron and hypoxic conditions.Bacterial survival in the presence of the indicated antibiotics under hypoxic conditions (A, C, E, G) and in low iron media (B, D, F, H). Isoniazid (“INH”, 2 and 0.25 µg ml−1, A and B), streptomycin (“SMP”, 2 and 1 µg ml−1, C and D), ciprofloxacin (“CIP”, 4 and 1 µg ml−1, E and F), and ethambutol (“EMB”, 5 and 3 µg ml−1, G and H) were introduced into each culture. Antibiotics were added to the hypoxic vials after 14 d of culture. Means ± SD of two independent experiments each performed in duplicate are shown.

Mentions: Since decreased metabolic activity generally correlates with lower antibiotic efficacy, we speculated that TAG synthesis might contribute to the drug-tolerant phenotype induced by stress. We tested this hypothesis using in vitro conditions that trigger TAG accumulation. Indeed, we found that the Δtgs1 mutant remained significantly more sensitive to a variety of antibiotics under tolerance-inducing conditions such as hypoxia and iron limitation (Figure 3). The antibiotics used were chemically distinct and targeted diverse cellular pathways, suggesting that the general hypersensitivity of the Δtgs1 bacteria was due to a fundamental alteration in cellular metabolism. As expected, the increased multidrug-susceptibility of the Δtgs1 mutant was much less pronounced under favorable growth conditions in which this gene is not induced (Figure S7). Under these conditions, the mutant was no more susceptible than wild type to any of the drugs tested, except the fatty acid synthesis inhibitor, isoniazid (INH). We conclude that while TAG synthesis may influence INH sensitivity through multiple mechanisms, the multidrug susceptibility of the Δtgs1 mutant is due to a general increase in growth rate and/or metabolic activity. This conclusion was supported by the remarkable antibiotic sensitivity of the citA* strain that we observed in tolerance-inducing cultures (Figure 3). This strain was killed even more rapidly than the Δtgs1 mutant, verifying that metabolic rate is a major determinant of antibiotic susceptibility under these conditions.


Metabolic regulation of mycobacterial growth and antibiotic sensitivity.

Baek SH, Li AH, Sassetti CM - PLoS Biol. (2011)

Metabolic modulation reverses the antibiotic tolerance induced by low                            iron and hypoxic conditions.Bacterial survival in the presence of the indicated antibiotics under                            hypoxic conditions (A, C, E, G) and in low iron media (B, D, F, H).                            Isoniazid (“INH”, 2 and 0.25 µg ml−1,                            A and B), streptomycin (“SMP”, 2 and 1 µg                                ml−1, C and D), ciprofloxacin (“CIP”, 4                            and 1 µg ml−1, E and F), and ethambutol                            (“EMB”, 5 and 3 µg ml−1, G and H)                            were introduced into each culture. Antibiotics were added to the hypoxic                            vials after 14 d of culture. Means ± SD of two independent                            experiments each performed in duplicate are shown.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3101192&req=5

pbio-1001065-g003: Metabolic modulation reverses the antibiotic tolerance induced by low iron and hypoxic conditions.Bacterial survival in the presence of the indicated antibiotics under hypoxic conditions (A, C, E, G) and in low iron media (B, D, F, H). Isoniazid (“INH”, 2 and 0.25 µg ml−1, A and B), streptomycin (“SMP”, 2 and 1 µg ml−1, C and D), ciprofloxacin (“CIP”, 4 and 1 µg ml−1, E and F), and ethambutol (“EMB”, 5 and 3 µg ml−1, G and H) were introduced into each culture. Antibiotics were added to the hypoxic vials after 14 d of culture. Means ± SD of two independent experiments each performed in duplicate are shown.
Mentions: Since decreased metabolic activity generally correlates with lower antibiotic efficacy, we speculated that TAG synthesis might contribute to the drug-tolerant phenotype induced by stress. We tested this hypothesis using in vitro conditions that trigger TAG accumulation. Indeed, we found that the Δtgs1 mutant remained significantly more sensitive to a variety of antibiotics under tolerance-inducing conditions such as hypoxia and iron limitation (Figure 3). The antibiotics used were chemically distinct and targeted diverse cellular pathways, suggesting that the general hypersensitivity of the Δtgs1 bacteria was due to a fundamental alteration in cellular metabolism. As expected, the increased multidrug-susceptibility of the Δtgs1 mutant was much less pronounced under favorable growth conditions in which this gene is not induced (Figure S7). Under these conditions, the mutant was no more susceptible than wild type to any of the drugs tested, except the fatty acid synthesis inhibitor, isoniazid (INH). We conclude that while TAG synthesis may influence INH sensitivity through multiple mechanisms, the multidrug susceptibility of the Δtgs1 mutant is due to a general increase in growth rate and/or metabolic activity. This conclusion was supported by the remarkable antibiotic sensitivity of the citA* strain that we observed in tolerance-inducing cultures (Figure 3). This strain was killed even more rapidly than the Δtgs1 mutant, verifying that metabolic rate is a major determinant of antibiotic susceptibility under these conditions.

Bottom Line: This pathway plays a causal role in reducing growth and antibiotic efficacy by redirecting cellular carbon fluxes away from the tricarboxylic acid cycle.Mutants in which this metabolic switch is disrupted are unable to arrest their growth in response to stress and remain sensitive to antibiotics during infection.Thus, this regulatory pathway contributes to antibiotic tolerance in vivo, and its modulation may represent a novel strategy for accelerating TB treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.

ABSTRACT
Treatment of chronic bacterial infections, such as tuberculosis (TB), requires a remarkably long course of therapy, despite the availability of drugs that are rapidly bacteriocidal in vitro. This observation has long been attributed to the presence of bacterial populations in the host that are "drug-tolerant" because of their slow replication and low rate of metabolism. However, both the physiologic state of these hypothetical drug-tolerant populations and the bacterial pathways that regulate growth and metabolism in vivo remain obscure. Here we demonstrate that diverse growth-limiting stresses trigger a common signal transduction pathway in Mycobacterium tuberculosis that leads to the induction of triglyceride synthesis. This pathway plays a causal role in reducing growth and antibiotic efficacy by redirecting cellular carbon fluxes away from the tricarboxylic acid cycle. Mutants in which this metabolic switch is disrupted are unable to arrest their growth in response to stress and remain sensitive to antibiotics during infection. Thus, this regulatory pathway contributes to antibiotic tolerance in vivo, and its modulation may represent a novel strategy for accelerating TB treatment.

Show MeSH
Related in: MedlinePlus