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Ranking of persister genes in the same Escherichia coli genetic background demonstrates varying importance of individual persister genes in tolerance to different antibiotics.

Wu N, He L, Cui P, Wang W, Yuan Y, Liu S, Xu T, Zhang S, Wu J, Zhang W, Zhang Y - Front Microbiol (2015)

Bottom Line: We found that oxyR, dnaK, sucB, relA, rpoS, clpB, mqsR, and recA were prominent persister genes involved in persistence to multiple antibiotics.These results indicate that varying hierarchy and importance of persister genes exist and that persister genes can be divided into those involved in shallow persistence and those involved in deep persistence.These findings have implications for improved understanding of persistence phenomenon and developing new drugs targeting persisters for more effective cure of persistent infections.

View Article: PubMed Central - PubMed

Affiliation: Key Lab of Molecular Virology, Institute of Medical Microbiology, Department of Infectious Diseases, Huashan Hospital, Fudan University Shanghai, China.

ABSTRACT
Despite the identification of many genes and pathways involved in the persistence phenomenon of bacteria, the relative importance of these genes in a single organism remains unclear. Here, using Escherichia coli as a model, we generated mutants of 21 known candidate persister genes and compared the relative importance of these mutants in persistence to various antibiotics (ampicillin, gentamicin, norfloxacin, and trimethoprim) at different times. We found that oxyR, dnaK, sucB, relA, rpoS, clpB, mqsR, and recA were prominent persister genes involved in persistence to multiple antibiotics. These genes map to the following pathways: antioxidative defense pathway (oxyR), global regulators (dnaK, clpB, and rpoS), energy production (sucB), stringent response (relA), toxin-antitoxin (TA) module (mqsR), and SOS response (recA). Among the TA modules, the ranking order was mqsR, lon, relE, tisAB, hipA, and dinJ. Intriguingly, rpoS deletion caused a defect in persistence to gentamicin but increased persistence to ampicillin and norfloxacin. Mutants demonstrated dramatic differences in persistence to different antibiotics at different time points: some mutants (oxyR, dnaK, phoU, lon, recA, mqsR, and tisAB) displayed defect in persistence from early time points, while other mutants (relE, smpB, glpD, umuD, and tnaA) showed defect only at later time points. These results indicate that varying hierarchy and importance of persister genes exist and that persister genes can be divided into those involved in shallow persistence and those involved in deep persistence. Our findings suggest that the persistence phenomenon is a dynamic process with different persister genes playing roles of variable significance at different times. These findings have implications for improved understanding of persistence phenomenon and developing new drugs targeting persisters for more effective cure of persistent infections.

No MeSH data available.


Related in: MedlinePlus

Deletion mutant ranking depends on cell survival over time under conditions of trimtheprim exposure. Stationary phase cultures of W3110 and single-gene mutants were diluted 100-fold and exposed to 64 μg/ml trimethoprim for 3, 5, and 7 days. The data for each mutant was plotted and compared with that of W3110. Error bars indicate the standard deviation (n = 3). The asterisk indicates statistical significance as determined using Mann–Whitney U tests (∗P < 0.05).
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Figure 4: Deletion mutant ranking depends on cell survival over time under conditions of trimtheprim exposure. Stationary phase cultures of W3110 and single-gene mutants were diluted 100-fold and exposed to 64 μg/ml trimethoprim for 3, 5, and 7 days. The data for each mutant was plotted and compared with that of W3110. Error bars indicate the standard deviation (n = 3). The asterisk indicates statistical significance as determined using Mann–Whitney U tests (∗P < 0.05).

Mentions: Both mutants and the wild-type strain showed higher tolerance to the bacteriostatic agent trimethoprim. After exposure to trimethoprim, the strains showed a lower decrease in colony counts compared with the other three bactericidal antibiotics. Interestingly, trimethoprim becomes bactericidal to some mutants during prolonged incubation (see below). The uvrA, relA, clpB, oxyR, and sucB deletion mutants had significantly lower persister levels from day 3, when the wild-type strain still had a high percentage of persisters at 68.8% (Figure 4). At day 5, other genes, smpB, dnaK, ssrA, and mqsR mutants entered into the dramatically decreasing phase. In contrast to the clpB and relA mutants, where reduction was almost top ranked through days 3–7, the decreasing trend of the uvrA mutant slowed from the topmost position at day 3 and was replaced by other mutants at the following time points (Figure 4). The data again support the notion that the importance of persister genes is relative, and varies with time (Li and Zhang, 2007; Ma et al., 2010). Moreover, relA, clpB, dnaK, and sucB mutant strains were more susceptible than the wild-type strain to the bacteriostatic antibiotic trimethoprim such that no viable bacteria were left in these mutants after 7 days exposure, while the wild-type strain had about 3.3 × 105 CFU/ml remaining.


Ranking of persister genes in the same Escherichia coli genetic background demonstrates varying importance of individual persister genes in tolerance to different antibiotics.

Wu N, He L, Cui P, Wang W, Yuan Y, Liu S, Xu T, Zhang S, Wu J, Zhang W, Zhang Y - Front Microbiol (2015)

Deletion mutant ranking depends on cell survival over time under conditions of trimtheprim exposure. Stationary phase cultures of W3110 and single-gene mutants were diluted 100-fold and exposed to 64 μg/ml trimethoprim for 3, 5, and 7 days. The data for each mutant was plotted and compared with that of W3110. Error bars indicate the standard deviation (n = 3). The asterisk indicates statistical significance as determined using Mann–Whitney U tests (∗P < 0.05).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4588708&req=5

Figure 4: Deletion mutant ranking depends on cell survival over time under conditions of trimtheprim exposure. Stationary phase cultures of W3110 and single-gene mutants were diluted 100-fold and exposed to 64 μg/ml trimethoprim for 3, 5, and 7 days. The data for each mutant was plotted and compared with that of W3110. Error bars indicate the standard deviation (n = 3). The asterisk indicates statistical significance as determined using Mann–Whitney U tests (∗P < 0.05).
Mentions: Both mutants and the wild-type strain showed higher tolerance to the bacteriostatic agent trimethoprim. After exposure to trimethoprim, the strains showed a lower decrease in colony counts compared with the other three bactericidal antibiotics. Interestingly, trimethoprim becomes bactericidal to some mutants during prolonged incubation (see below). The uvrA, relA, clpB, oxyR, and sucB deletion mutants had significantly lower persister levels from day 3, when the wild-type strain still had a high percentage of persisters at 68.8% (Figure 4). At day 5, other genes, smpB, dnaK, ssrA, and mqsR mutants entered into the dramatically decreasing phase. In contrast to the clpB and relA mutants, where reduction was almost top ranked through days 3–7, the decreasing trend of the uvrA mutant slowed from the topmost position at day 3 and was replaced by other mutants at the following time points (Figure 4). The data again support the notion that the importance of persister genes is relative, and varies with time (Li and Zhang, 2007; Ma et al., 2010). Moreover, relA, clpB, dnaK, and sucB mutant strains were more susceptible than the wild-type strain to the bacteriostatic antibiotic trimethoprim such that no viable bacteria were left in these mutants after 7 days exposure, while the wild-type strain had about 3.3 × 105 CFU/ml remaining.

Bottom Line: We found that oxyR, dnaK, sucB, relA, rpoS, clpB, mqsR, and recA were prominent persister genes involved in persistence to multiple antibiotics.These results indicate that varying hierarchy and importance of persister genes exist and that persister genes can be divided into those involved in shallow persistence and those involved in deep persistence.These findings have implications for improved understanding of persistence phenomenon and developing new drugs targeting persisters for more effective cure of persistent infections.

View Article: PubMed Central - PubMed

Affiliation: Key Lab of Molecular Virology, Institute of Medical Microbiology, Department of Infectious Diseases, Huashan Hospital, Fudan University Shanghai, China.

ABSTRACT
Despite the identification of many genes and pathways involved in the persistence phenomenon of bacteria, the relative importance of these genes in a single organism remains unclear. Here, using Escherichia coli as a model, we generated mutants of 21 known candidate persister genes and compared the relative importance of these mutants in persistence to various antibiotics (ampicillin, gentamicin, norfloxacin, and trimethoprim) at different times. We found that oxyR, dnaK, sucB, relA, rpoS, clpB, mqsR, and recA were prominent persister genes involved in persistence to multiple antibiotics. These genes map to the following pathways: antioxidative defense pathway (oxyR), global regulators (dnaK, clpB, and rpoS), energy production (sucB), stringent response (relA), toxin-antitoxin (TA) module (mqsR), and SOS response (recA). Among the TA modules, the ranking order was mqsR, lon, relE, tisAB, hipA, and dinJ. Intriguingly, rpoS deletion caused a defect in persistence to gentamicin but increased persistence to ampicillin and norfloxacin. Mutants demonstrated dramatic differences in persistence to different antibiotics at different time points: some mutants (oxyR, dnaK, phoU, lon, recA, mqsR, and tisAB) displayed defect in persistence from early time points, while other mutants (relE, smpB, glpD, umuD, and tnaA) showed defect only at later time points. These results indicate that varying hierarchy and importance of persister genes exist and that persister genes can be divided into those involved in shallow persistence and those involved in deep persistence. Our findings suggest that the persistence phenomenon is a dynamic process with different persister genes playing roles of variable significance at different times. These findings have implications for improved understanding of persistence phenomenon and developing new drugs targeting persisters for more effective cure of persistent infections.

No MeSH data available.


Related in: MedlinePlus