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A linkage between SmeIJK efflux pump, cell envelope integrity, and σE-mediated envelope stress response in Stenotrophomonas maltophilia.

Huang YW, Liou RS, Lin YT, Huang HH, Yang TC - PLoS ONE (2014)

Bottom Line: Moreover, the expression of smeIJK was further induced by sub-lethal concentrations of MDAs or surfactants in an σE-dependent manner.Because of the necessity of the physiological role of SmeIJK in protecting S. maltophilia from the envelope stress, smeIJK is constitutively expressed, which, in turn, contributes the intrinsic resistance to aminoglycoside and leucomycin.This is the first demonstration of the linkage among RND-type efflux pump, cell envelope integrity, and σE-mediated ESR in S. maltophilia.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology and Laboratory Science in Medicine, National Yang-Ming University, Taipei, Taiwan.

ABSTRACT
Resistance nodulation division (RND) efflux pumps, such as the SmeIJK pump of Stenotrophomonas maltophilia, are known to contribute to the multidrug resistance in Gram-negative bacteria. However, some RND pumps are constitutively expressed even though no antimicrobial stresses occur, implying that there should be some physical implications for these RND pumps. In this study, the role of SmeIJK in antimicrobials resistance, envelope integrity, and σE-mediated envelope stress response (ESR) of S. maltophilia was assessed. SmeIJK was involved in the intrinsic resistance of S. maltophilia KJ to aminoglycosides and leucomycin. Compared with the wild-type KJ, the smeIJK deletion mutant exhibited growth retardation in the MH medium, an increased sensitivity to membrane-damaging agents (MDAs), as well as activation of an σE-mediated ESR. Moreover, the expression of smeIJK was further induced by sub-lethal concentrations of MDAs or surfactants in an σE-dependent manner. These data collectively suggested an alternative physiological role of smeIJK in cell envelope integrity maintenance and σE-mediated ESR beyond the efflux of antibiotics. Because of the necessity of the physiological role of SmeIJK in protecting S. maltophilia from the envelope stress, smeIJK is constitutively expressed, which, in turn, contributes the intrinsic resistance to aminoglycoside and leucomycin. This is the first demonstration of the linkage among RND-type efflux pump, cell envelope integrity, and σE-mediated ESR in S. maltophilia.

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The role of SmeIJK pump in the tolerance to hypo-osomolarity and casein hydrolysate.The data are the average of the measurements made in triplicate. (A) The relative survival of KJΔIJK, KJΔJ, and KJΔK to the wild-type KJ in the LB medium containing different concentrations of NaCl. The relative survival percentage of individual mutant to the wild-type KJ, at each cultured condition, was calculated using the OD450 nm of the wild-type KJ as 100%. (B) The relative survival of KJΔIJK, KJΔJ, and KJΔK to the wild-type KJ in the MH medium containing different concentrations of NaCl. The relative survival percentage of individual mutant to the wild-type KJ, at each cultured condition, was calculated using the OD450 nm of the wild-type KJ as 100%. (C) The sensitivities of KJ to SDS in LB or LB containing casein hydrolysate, beef infusion, or starch were determined by the OD450 nm measurement. The percentage of survival was defined as the OD450 nm ratio of the SDS-additive group to the SDS-free counterpart. (D) Growth curves of KJΔIJK grown in the media of the LB, the LB without NaCl, the LB with casein hydrolysate, and the MH, respectively.
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pone-0111784-g002: The role of SmeIJK pump in the tolerance to hypo-osomolarity and casein hydrolysate.The data are the average of the measurements made in triplicate. (A) The relative survival of KJΔIJK, KJΔJ, and KJΔK to the wild-type KJ in the LB medium containing different concentrations of NaCl. The relative survival percentage of individual mutant to the wild-type KJ, at each cultured condition, was calculated using the OD450 nm of the wild-type KJ as 100%. (B) The relative survival of KJΔIJK, KJΔJ, and KJΔK to the wild-type KJ in the MH medium containing different concentrations of NaCl. The relative survival percentage of individual mutant to the wild-type KJ, at each cultured condition, was calculated using the OD450 nm of the wild-type KJ as 100%. (C) The sensitivities of KJ to SDS in LB or LB containing casein hydrolysate, beef infusion, or starch were determined by the OD450 nm measurement. The percentage of survival was defined as the OD450 nm ratio of the SDS-additive group to the SDS-free counterpart. (D) Growth curves of KJΔIJK grown in the media of the LB, the LB without NaCl, the LB with casein hydrolysate, and the MH, respectively.

Mentions: An MH medium typically contains 0.3% beef infusion, 1.75% casein hydrolysate, and 0.15% starch. The ingredients for LB media were 1% tryptone, 0.5% yeast extract, and 1% (0.17 M) NaCl. Given the compositions of the MH and LB media, we speculated that the observed growth retardation in the MH medium was due to the absence of NaCl in the medium. To test this assumption, three LB-based media with a final NaCl concentration of 0, 0.17, and 0.3 M were prepared. The osmotic challenge assay to KJ, KJΔJ, KJΔK, and KJΔIJK cells was determined. KJΔIJK displayed a reduced survival in the 0 M NaCl LB medium when compared to the wild-type KJ. Nevertheless, the survival of KJΔIJK cells restored to nearly wild-type levels in the LB supplemented with 0.17 M and 0.3 M NaCl (Fig. 2A). The phenotype correlated smeIJK to the hypo-osmolarity tolerance. Furthermore, the survivals of KJΔJ and KJΔK were similar to that of the wild-type KJ in spite of the concentration of NaCl in the cultured medium (Fig. 2A). The osmotic challenge assay of KJ, KJΔJ, KJΔK, and KJΔIJK was also performed in three MH-based media containing 0, 0.17, and 0.3 M NaCl. The results concluded from the MH counterpart (Fig. 2B) is consistent with those from the LB counterpart. It is worthily noted that the ΔsmeIJK-mediated compromise in hypo-osmolarity tolerance was more apparent in the MH counterpart than that in the LB counterpart, implying that some components, in addition to NaCl, in the MH medium may cause an envelope stress to S. maltophilia. To test this assumption, we supplemented LB with individual compounds present in MH medium to determine which component affected susceptibility to SDS. Only the addition of casein hydrolysate increased the sensitivity of KJ cells to SDS (Fig. 2C)


A linkage between SmeIJK efflux pump, cell envelope integrity, and σE-mediated envelope stress response in Stenotrophomonas maltophilia.

Huang YW, Liou RS, Lin YT, Huang HH, Yang TC - PLoS ONE (2014)

The role of SmeIJK pump in the tolerance to hypo-osomolarity and casein hydrolysate.The data are the average of the measurements made in triplicate. (A) The relative survival of KJΔIJK, KJΔJ, and KJΔK to the wild-type KJ in the LB medium containing different concentrations of NaCl. The relative survival percentage of individual mutant to the wild-type KJ, at each cultured condition, was calculated using the OD450 nm of the wild-type KJ as 100%. (B) The relative survival of KJΔIJK, KJΔJ, and KJΔK to the wild-type KJ in the MH medium containing different concentrations of NaCl. The relative survival percentage of individual mutant to the wild-type KJ, at each cultured condition, was calculated using the OD450 nm of the wild-type KJ as 100%. (C) The sensitivities of KJ to SDS in LB or LB containing casein hydrolysate, beef infusion, or starch were determined by the OD450 nm measurement. The percentage of survival was defined as the OD450 nm ratio of the SDS-additive group to the SDS-free counterpart. (D) Growth curves of KJΔIJK grown in the media of the LB, the LB without NaCl, the LB with casein hydrolysate, and the MH, respectively.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4229105&req=5

pone-0111784-g002: The role of SmeIJK pump in the tolerance to hypo-osomolarity and casein hydrolysate.The data are the average of the measurements made in triplicate. (A) The relative survival of KJΔIJK, KJΔJ, and KJΔK to the wild-type KJ in the LB medium containing different concentrations of NaCl. The relative survival percentage of individual mutant to the wild-type KJ, at each cultured condition, was calculated using the OD450 nm of the wild-type KJ as 100%. (B) The relative survival of KJΔIJK, KJΔJ, and KJΔK to the wild-type KJ in the MH medium containing different concentrations of NaCl. The relative survival percentage of individual mutant to the wild-type KJ, at each cultured condition, was calculated using the OD450 nm of the wild-type KJ as 100%. (C) The sensitivities of KJ to SDS in LB or LB containing casein hydrolysate, beef infusion, or starch were determined by the OD450 nm measurement. The percentage of survival was defined as the OD450 nm ratio of the SDS-additive group to the SDS-free counterpart. (D) Growth curves of KJΔIJK grown in the media of the LB, the LB without NaCl, the LB with casein hydrolysate, and the MH, respectively.
Mentions: An MH medium typically contains 0.3% beef infusion, 1.75% casein hydrolysate, and 0.15% starch. The ingredients for LB media were 1% tryptone, 0.5% yeast extract, and 1% (0.17 M) NaCl. Given the compositions of the MH and LB media, we speculated that the observed growth retardation in the MH medium was due to the absence of NaCl in the medium. To test this assumption, three LB-based media with a final NaCl concentration of 0, 0.17, and 0.3 M were prepared. The osmotic challenge assay to KJ, KJΔJ, KJΔK, and KJΔIJK cells was determined. KJΔIJK displayed a reduced survival in the 0 M NaCl LB medium when compared to the wild-type KJ. Nevertheless, the survival of KJΔIJK cells restored to nearly wild-type levels in the LB supplemented with 0.17 M and 0.3 M NaCl (Fig. 2A). The phenotype correlated smeIJK to the hypo-osmolarity tolerance. Furthermore, the survivals of KJΔJ and KJΔK were similar to that of the wild-type KJ in spite of the concentration of NaCl in the cultured medium (Fig. 2A). The osmotic challenge assay of KJ, KJΔJ, KJΔK, and KJΔIJK was also performed in three MH-based media containing 0, 0.17, and 0.3 M NaCl. The results concluded from the MH counterpart (Fig. 2B) is consistent with those from the LB counterpart. It is worthily noted that the ΔsmeIJK-mediated compromise in hypo-osmolarity tolerance was more apparent in the MH counterpart than that in the LB counterpart, implying that some components, in addition to NaCl, in the MH medium may cause an envelope stress to S. maltophilia. To test this assumption, we supplemented LB with individual compounds present in MH medium to determine which component affected susceptibility to SDS. Only the addition of casein hydrolysate increased the sensitivity of KJ cells to SDS (Fig. 2C)

Bottom Line: Moreover, the expression of smeIJK was further induced by sub-lethal concentrations of MDAs or surfactants in an σE-dependent manner.Because of the necessity of the physiological role of SmeIJK in protecting S. maltophilia from the envelope stress, smeIJK is constitutively expressed, which, in turn, contributes the intrinsic resistance to aminoglycoside and leucomycin.This is the first demonstration of the linkage among RND-type efflux pump, cell envelope integrity, and σE-mediated ESR in S. maltophilia.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology and Laboratory Science in Medicine, National Yang-Ming University, Taipei, Taiwan.

ABSTRACT
Resistance nodulation division (RND) efflux pumps, such as the SmeIJK pump of Stenotrophomonas maltophilia, are known to contribute to the multidrug resistance in Gram-negative bacteria. However, some RND pumps are constitutively expressed even though no antimicrobial stresses occur, implying that there should be some physical implications for these RND pumps. In this study, the role of SmeIJK in antimicrobials resistance, envelope integrity, and σE-mediated envelope stress response (ESR) of S. maltophilia was assessed. SmeIJK was involved in the intrinsic resistance of S. maltophilia KJ to aminoglycosides and leucomycin. Compared with the wild-type KJ, the smeIJK deletion mutant exhibited growth retardation in the MH medium, an increased sensitivity to membrane-damaging agents (MDAs), as well as activation of an σE-mediated ESR. Moreover, the expression of smeIJK was further induced by sub-lethal concentrations of MDAs or surfactants in an σE-dependent manner. These data collectively suggested an alternative physiological role of smeIJK in cell envelope integrity maintenance and σE-mediated ESR beyond the efflux of antibiotics. Because of the necessity of the physiological role of SmeIJK in protecting S. maltophilia from the envelope stress, smeIJK is constitutively expressed, which, in turn, contributes the intrinsic resistance to aminoglycoside and leucomycin. This is the first demonstration of the linkage among RND-type efflux pump, cell envelope integrity, and σE-mediated ESR in S. maltophilia.

Show MeSH
Related in: MedlinePlus