Carotenoid-Related Alteration of Cell Membrane Fluidity Impacts Staphylococcus aureus Susceptibility to Host Defense Peptides
ABSTRACT
Carotenoid pigments of Staphylococcus aureus provide integrity to its cell membrane (CM) and limit oxidative host defense mechanisms. However, the role of carotenoids in staphylococcal resistance to nonoxidative host defenses has not been characterized. The current study examined the relationship among CM carotenoid content, membrane order, and in vitro susceptibility to daptomycin or to prototypic neutrophil-derived, platelet-derived, or bacterium-derived cationic antimicrobial peptides (human neutrophil defensin-1 [hNP-1], platelet microbicidal proteins [PMPs], or polymyxin B, respectively). A previously characterized methicillin-susceptible Staphylococcus aureus (MSSA) isogenic clinical strain set was used, including a parental isolate with an intact carotenoid biosynthetic operon (crtOPQMN) containing the crtM gene encoding early steps in staphyloxanthin biosynthesis, a crtM deletion mutant, and a crtMN multicopy plasmid-complemented variant. Compared to the parental and crtM knockout strains, the crtMN-complemented strain exhibited (i) increased carotenoid production, (ii) increased CM rigidity (P < 0.001), and (iii) uniformly reduced susceptibility to killing by the above-mentioned range of cationic peptides (statistically significant for hNP-1 [20 μg/ml]; P = 0.0037). There were no significant differences in phospholipid composition and asymmetry, fatty acid profiles, surface charge, or cell wall thickness among the strain set. Collectively, these data support the concept that carotenoid biosynthesis can contribute to the ability of S. aureus to subvert nonoxidative host defenses mediated by cationic peptides, potentially by increasing target membrane rigidity.
Over 90% of the Staphylococcus aureus strains isolated from human infections are pigmented (18). Staphyloxanthin (STX) is an orange-red triterpenoid, membrane-bound carotenoid which plays a role in the environmental fitness of S. aureus (4, 25). Membrane pigments have also been hypothesized to be virulence factors in S. aureus, potentially by detoxifying reactive oxygen species produced by phagocytes (19). Carotenoids may also stabilize the S. aureus membrane during infection and pathogenesis. For example, Rohmer et al. (26) postulated that polar carotenoids regulate membrane properties of prokaryotes in a manner similar to that observed for cholesterol in eukaryotes. In this regard, Liu et al. (19, 20) recently observed that STX synthesis in staphylococcal cells is associated with resistance to phagocyte-mediated killing in vitro and staphylococcal persistence in target organs in relevant in vivo animal models.
In contrast, the potential role of S. aureus pigments in resistance to killing by nonoxidative host defenses has not been extensively studied. It is known that polar carotenoids modulate the fluidity properties of natural and model lipid membranes (7, 8, 9, 27, 30), and such fluidity characteristics are critical to the interaction of membrane-targeting host defense cationic antimicrobial peptides (CAPs) with S. aureus. Thus, the current study was designed to assess the relationships among STX production, cell membrane (CM) biophysics, and susceptibility to host defense and other cationic peptides.
The biosynthetic pathway for STX is depicted in Fig. 1. This figure includes those crt operon genes pivotally involved at the various STX synthesis steps, as well as showing how the presence or absence of carotenoids might theoretically impact staphylococcal membrane interactions with cationic peptides.
(This work was presented in part at the 49th ICAAC, San Francisco, CA, 12 to 15 September 2009 [23a].)
MATERIALS AND METHODS
Bacterial strains.
The strain set used in this study is listed in Table 1, including the methicillin-susceptible S. aureus (MSSA) parental strain (SA144, with an intact crt operon), its crtM deletion mutant (SA145), and a crtMN plasmid-complemented variant of SA145 (SA147). The last strain contains a multicopy plasmid (pDCerm::crtMN) (11) resulting in enhanced STX production compared to the level for the parental strain. In addition, for selected assays, we employed an “empty plasmid” variant of SA145 containing the above-mentioned plasmid vector alone (i.e., without the crtMN genes). The genetic constructions of the crtM mutant and the plasmid-complemented variant have previously been described in detail (20).
DAP susceptibilities.
The MICs of the strain set for a conventional positively charged (calcium-decorated), membrane-targeting antibiotic, daptomycin (DAP), were determined by Etest (AB Biodisk, Dalvagen, Sweden) on Mueller-Hinton agar (MHA) plates (Difco Laboratories, Detroit, MI). For DAP Etest, plates were calcium supplemented per the manufacturer's recommendations (50 μg/ml CaCl2).
CAP susceptibilities.
Standard MIC assays for many CAPs are not routinely performed, as conventional nutrient medium can cause underestimation of activities of these peptides. Thus, bactericidal assays were carried out with liquid medium by kill curve techniques for the following cationic peptides: (i) two kinds of prototypic mammalian-derived host defense CAPs, thrombin-induced platelet microbicidal proteins (tPMPs) and human neutrophil-derived defensin 1 (hNP-1); (ii) a synthetic platelet peptide congener (RP-1) modeled after the α-helical microbicidal domain of such molecules (35); (iii) a bacterium-derived cyclic CAP, polymyxin B (PMB); and (iv) the cyclic lipopeptide calcium-decorated daptomycin. The bioactive concentrations of the peptides used in these killing assays were 0.125 to 0.25 for tPMP-1, 10 to 20 μg/ml for hNP-1, 2.5 to 5 μg/ml for RP-1, 40 to 80 μg/ml for PMB, and 0.625 to 1.25 μg/ml for DAP. These concentrations were selected based on the ability to reduce the survival of the parental strain (SA144) by >50% in preliminary studies and included concentrations used in prior investigations of CAP-S. aureus interactions (24, 33). The hNP-1 was purchased from Peptide International (Louisville, KY), PMB was purchased from Sigma-Aldrich (St. Louis, MO), and DAP was obtained from Cubist Pharmaceuticals (Lexington, MA). The tPMP preparation was recovered from thrombin-stimulated rabbit platelets as previously described (36). RP-1 was synthesized as previously detailed (35). Stationary-phase cells were utilized in all assays (24-h growth) to parallel the time of maximal carotenoid production in vitro.
All CAPs mentioned above were reconstituted in appropriate diluents as described previously (33, 35). DAP was reconstituted according to the manufacturer's recommendations. S. aureus cells were diluted into the peptide solutions to achieve the desired final inoculum (103 CFU/ml) (32, 33) and peptide concentrations and then incubated at 37°C. At 120 min of exposure, samples were removed and processed for quantitative culture to assess the extent of killing by each CAP. Final data were expressed as mean (± standard deviation [SD]) log10 numbers of surviving CFU/ml. A minimum of two experimental runs on separate days was performed.
In addition, we quantified the killing of our study strains by one of the CAPs mentioned above (hNP-1) over a 6-h time period. The same methods as those described above for the standard 2-h killing assay were employed for the 6-h experiments.
Carotenoid extraction.
Extraction and quantification of CM carotenoids were carried out as previously described (20). Stationary-phase (24-h) cultures of S. aureus were equilibrated for growth yield and then subjected to methanol extraction (22). Carotenoid content was then quantified by the absorbance profile of the extracts as determined spectrophotometrically at an optical density at a wavelength of 450 nm (OD450) (20).
CM fluidity.
The CM order appears to influence the interactions of certain host defense CAPs with S. aureus (1, 24, 33). We utilized polarizing spectrofluorometry to analyze the membrane fluidity properties of the study S. aureus cells grown at 37°C for 24 h. Fluidity measurements were carried out using the fluorescent probe 1,6-diphenyl-1,3,5-hexatriene (DPH). The protocol for DPH incorporation into target CMs, the measurement of fluorescence polarization, and the calculation of the degree of fluorescence polarization (polarization index [PI]) are detailed elsewhere (Biotek model SFM 25 spectrofluorimeter; excitation and emission wavelengths of 360 and 426 nm, respectively, for DPH) (1). Prior studies have established that an inverse relationship exists between PI values and CM fluidity (i.e., a lower PI equates to a greater extent of CM fluidity) (1). These assays were performed a minimum of three times for each strain on separate days.
CM fatty acid composition.
In addition to carotenoid content, fatty acid composition can substantially influence the biophysical properties of the CM (1). Fatty acids of total lipid extracts from S. aureus CMs were analyzed using gas-liquid chromatography after derivatization to their methyl ester form as previously confirmed (courtesy of Microbial ID, Inc., Newark, DE) (1).
Surface charge.
In addition to CM order, cell envelope charge can influence the capacity of some host defense CAPs to kill S. aureus (12). Although STX is a charge-neutral molecule, displacement of negatively charged lipids by this carotenoid could conceivably alter surface charge. To rule out this possibility, cell surface charges were compared among the study strains. We utilized poly-l-lysine (PLL; a polycationic molecule) to estimate net surface charge (10, 16). A fluorescein isothiocyanate-labeled PLL binding assay was performed using flow cytometry as previously described (FACSCalibur; Beckman Instruments, Alameda, CA) (24). Data were expressed as mean (± SD) relative fluorescence units. In this assay, a lower-level residual cell-associated label is indicative of a greater positive charge of the S. aureus cell envelope (24). At least two independent runs were performed on separate days.
CW thickness.
The carotenoid content of the CM has the potential to secondarily alter overall cell wall (CW) architecture (5, 23). To screen for compensatory changes in the staphylococcal CW in response to modulations of CM carotenoid content, CW thickness was assessed by transmission electron microscopy (TEM) (21). The mean (± SD) thickness of 100 cells was determined for our strain set at a magnification of ×190,000 (model 100CX; JEOL, Tokyo, Japan), using digital image capture and morphometric measurement (v54; Advanced Microscopy Techniques, Danvers, MA). Cells were prepared for TEM by previously published techniques (17).
CM PL content and PL asymmetry.
The phospholipid (PL) composition of the S. aureus CM, as well as the PL translocation from the inner to the outer leaflet of the CM, can impact the net surface charge of the organism. This phenomenon is particularly relevant to host defense CAPs, where translocation of the positively charged PL lysyl-phosphotidylglycerol (LPG) to the outer CM leaflet can alter peptide association with the CM (12). Thus, the three major S. aureus PLs (phosphotidylglycerol [PG], LPG, and cardiolipin [CL]) were separated using two-dimensional thin-layer chromatography (2D-TLC). These PLs were then isolated and quantified spectrophotometrically at OD660. Outer LPG was identified by UV detection of fluorescamine labeling as previously detailed (23, 24).
Statistical analysis.
Means and standard deviations were calculated for all variables. Differences between strains for killing and CM order assays were analyzed with one-way analysis of variance (ANOVA), using the Sidak method for multiple comparison adjustments. P values of ≤0.05 were considered “significant.”
RESULTS
MIC.
Comparative genotypes and DAP MIC values for the study strains are summarized in Table 1. Notably, the DAP MICs for strains SA144, SA145, and SA146 were 0.75 μg/ml (daptomycin-susceptible). In comparison, a 2-fold increase in DAP MIC was seen in the carotenoid-overproducing strain, SA147 (1.5 μg/ml; daptomycin nonsusceptible).
Carotenoid analysis.
As expected, the carotenoid knockout mutant (ΔcrtM; SA145) had minimal detectable carotenoid compared to the parental strain, SA144. Also, as anticipated, the quantity of carotenoid pigment was highest in the complemented, crtMN-overexpressing strain, SA147 (data not shown).
CM fluidity.
As predicted, the carotenoid-overproducing strain, SA147, exhibited the most-rigid CMs, while the crtM knockout mutant, SA145, had the most-fluid CMs (compared to the level for the parental strain) (Fig. 2). These results achieved statistical significance in comparison of strains SA144 and SA145 (P = 0.008), SA144 and SA147 (P < 0.001), and SA145 and SA147 (P < 0.001).
CAP susceptibilities.
Of interest, the ΔcrtM knockout mutant (SA145) did not exhibit any change in CAP susceptibility compared to the level for its isogenic parental strain (SA144). In contrast, the carotenoid-overproducing strain, SA147, exhibited an increased resistance (∼2-fold less killing) to most of the study CAPs, compared to the level for the parental strain, SA144 (Table 2). These results achieved statistical significance for hNP-1 (20 μg/ml; P = 0.0037 [SA147 versus SA144]). There was a trend toward increased hNP-1 resistance for SA147 versus that for SA145, although this did not reach statistical significance (P = 0.087). In addition, the survival profile of SA147 was increased over a 6-h exposure to hNP-1 (20 μg/ml), compared to the levels for both SA144 and SA145 (Fig. 3). To eliminate the possibility of a nonspecific plasmid vector effect, we tested CAP-induced killing of the “empty plasmid” construct (SA146). The extent of 2 h of killing of SA146 by tPMP (0.25 μg/ml) and hNP-1 (10 μg/ml) was very similar to that observed for the parental and ΔcrtM knockout mutant strains for these same CAPs (data not shown).
(It should be mentioned that upon post hoc analysis, we identified technical errors in a minority of the peptide assays done for tPMP, RP-1, DAP, and PMB [but not hNP-1]. These data were excluded from the above-described analysis.)
Cell surface charge.
Net surface charge did not significantly differ among strains SA144, SA145, and SA147 (data not shown).
CW thickness.
No significant differences were observed in CW architecture or thickness as a function of carotenoid expression or quantity in the study strain set (cell wall thicknesses of 31.1 ± 3.8 nm for SA144, 33.9 ± 3.8 nm for SA145, and 31.2 ± 3.5 nm for SA147).
CM PL and fatty acid composition.
Detailed fatty acid analysis, including chain length profiling, branched-chain (iso and anteiso species) and straight-chain profiles, and unsaturation indices, were virtually identical in the strain set (Table 3). In addition, CM phospholipid (PL) composition and inner-to-outer-CM translocation profiles were not significantly different among the strain set (Table 3).
DISCUSSION
Prior studies have indicated that carotenoids may afford S. aureus protection against oxidative host defense mechanisms (4, 19, 20, 25). Such antioxidant potential of carotenoids may be related to their specific orientation, localization, and organization within the membrane (29). Nonpolar and polar carotenoids exert distinct effects on lipid membrane structure and physiology (29). On the basis of several recent seminal studies (19, 20), it appears that STX production plays a role in the ability of S. aureus to resist clearance by the oxidative limb of the host immune system (i.e., the NADPH oxidase or “respiratory burst” pathway).
The STX pigment consists of a C30-polyene backbone with alternating single and double bonds, capable of quenching oxidation by reactive oxygen species (6, 21). Deletion of the gene encoding the upstream STX biosynthesis enzyme, crtM, renders the bacterium more susceptible to killing by human and mouse polymorphonuclear leukocytes (PMNs) or whole blood (4, 20). In addition, knockout mice with homozygotic deletions of the gp91phox gene (encoding a component of the NADPH oxidase enzyme complex involved in oxidative immune defense) exhibited in vitro defects in killing carotenoid-producing S. aureus strains and decreased the ability to clear such organisms from systemic infections (20). In a complementary manner, compared with its wild-type parental strain, a S. aureus crtM mutant strain was much more susceptible to killing by hydrogen peroxide, superoxide radicals, hydroxyl radicals, hypochloride, and singlet oxygen species (4, 20). Finally, loss of STX pigmentation translated to a significant reduction in S. aureus virulence in murine skin abscess as well as systemic infection models. Despite these compelling investigations corroborating the protective role of carotenoid pigments in S. aureus resistance to host oxidative defenses, the potential function of such pigments in nonoxidative innate host defense pathways is less well studied.
As carotenoids may significantly impact the CM biophysical properties of S. aureus, and most mammalian host defense peptides initially target the microbial electronegative CM, examining the correlations among pigment production, CM biophysics, and peptide susceptibilities is important. In this regard, Liu et al. (20) showed that carotenoid-deficient and wild-type S. aureus strains had equivalent susceptibilities to murine CRAMP, an 18-amino-acid cathelicidin host defense peptide (CAP) that carries a charge of +5. Of interest, Katzif et al. have shown that pigment production in S. aureus is dependent on the function of at least two stress response gene-dependent mechanisms (sigma factor B and cold shock protein A [sigB-cspA]) (15). Moreover, they observed that cspA is involved in the susceptibility of S. aureus to at least one human CAP (cathepsin G) (14). To investigate and clarify CAP-carotenoid interactions further, the current studies compared a panel of host defense peptides of different charges, structures, and sources regarding carotenoid influence on antistaphylococcal efficacy. A well-characterized isogenic MSSA strain set was studied, including a clinically derived parental strain (with an intact crtOPQMN operon), its crtM mutant, and a crtMN-complemented and carotenoid-overproducing variant (20).
Several interesting observations emerged from our studies. Fluidity characteristics of the CM are essential for bacterial viability. For example, CM-bound carotenoids stabilize intra- and extracellular leaflets of the lipid bilayer and increase CM rigidity by ordering the alkyl chains of CM lipids (26, 28, 29, 31). In the present study, the amount of pigment production was directly correlated with the relative fluidity-rigidity CM profiles in our strain set. Thus, the CMs of the carotenoid-overproducing strain exhibited substantially more-rigid CMs than the parental and carotenoid-deficient strains. Of interest, we and others have shown a prominent correlation between the relative state of CM order in S. aureus and the organism's profiles of susceptibility to selected host defense CAPs. Therefore, irrespective of the pathway or mechanisms involved, extremes of CM fluidity-rigidity can be associated with an altered susceptibility to certain CAPs (2). For example, the presence of a CM-spanning quaternary ammonium transporter (qacA) both alters CM fluidity and reduces killing by tPMPs in an efflux pump-independent manner (2). Similarly, clinical S. aureus strains which acquired DAP resistance in vivo have been documented to coevolve both enhanced CM fluidity and reduced killing by mammalian host defense CAPs (12, 34). In contrast, S. aureus strains emerging as DAP resistant by serial in vitro DAP passage became concomitantly more resistant to killing by certain host defense CAPs, while exhibiting more-rigid CMs (23). Collectively, such findings have led to the hypothesis that CM fluidity differentially influences distinct binding of peptide to or subsequent perturbations of the target CM. A corollary to this paradigm is that extremes of CM order (i.e., very fluid or very rigid CMs) can be associated with reduced killing by selected CAPs, and there likely exists a CM order optimum for each bacterial CM-peptide interaction (the so-called “sweet spot”) (1, 23, 28). This notion is underscored in our investigation by the finding that tipping CM order toward increased fluidity (e.g., in the crtM knockout) did not impact CAP susceptibility profiles but that tipping CM order toward enhanced rigidity (i.e., in the carotenoid overproducer) exerted a major impact in this context. The current findings are also consistent with our recent nuclear magnetic resonance (NMR)-defined interaction studies between RP-1 and model prokaryotic lipid membrane systems (3).
In the present study, the carotenoid-overproducing strain was less susceptible to in vitro killing by many of the study peptides than the parental and/or crtM knockout strain. It was important to confirm that the differences we observed in these susceptibility profiles were specifically related to extremes in carotenoid content and not an epiphenomena to the influences of other factors impacting CM or surface envelope characteristics (e.g., CM lipid composition, net surface charge, etc.). We were particularly interested in CM fatty acid profiles, since carotenogenesis modulation can lead to a buildup of C30 precursor species that can then be shuttled into menaquinone-fatty acid oxidation pathways (13). However, fatty acid profiling showed no differences in our strain set in any parameter known to impact CM order, i.e., chain lengths, unsaturation indices, or branched-chain (iso and anteiso species) and straight-chain profiles (12).
The above-mentioned findings underscore the hypothesis that a net carotenoid homeostasis influences S. aureus survival in the face of nonoxidative host defenses. Thus, although the ΔcrtM knockout strain produced negligible amounts of STX, this strain did not substantially differ in CAP profiling from the parental strain. In contrast, excess STX production was sufficient to substantially impact such susceptibility profiles (albeit only statistically significant for the PMN CAP, hNP-1). A parallel theme emphasizes the idea that the ability of STX overexpression (and its attendant enhancement of CM rigidity) to impair CAP-mediated killing of S. aureus likely represents a relatively “CAP-nonspecific” mechanism. For example, these trends for STX overexpression to be associated with reduced CAP susceptibility were observed for an entire range of peptide secondary structures and cationicity, including RP-1 (α-helix), hNP-1 (β-sheet), PMB (cyclic peptide), and DAP (cyclic lipopeptide).
We recognize that our study has certain limitations: (i) only a single S. aureus strain set was evaluated; (ii) the parental strain was clinically derived, raising the possibility that host exposures and/or genetic pathways outside crtOPQMN might have influenced these results; (iii) a relatively narrow range of host defense CAPs was investigated, leaving open the potential that CAPs of different sources, structures, or mechanisms of action might exhibit distinct effects on their activity by carotenoid-related CM perturbations; and (iv) host defense CAPs were tested individually, in austere buffer systems, and at sublethal concentrations in vitro. These facts limit the potential for in vivo translatability of our findings. Nonetheless, the results from these investigations support the hypothesis that S. aureus employs adaptive mechanisms by which it may utilize carotenoids to modify their CMs in order to subvert innate host defenses. Current studies are ongoing to further explore these concepts.
FIG. 1.
FIG. 2.
FIG. 3.
TABLE 1.
| Strain | Description | DAP MIC (μg/ml) |
|---|---|---|
| SA144 | crtM parent | 0.75 |
| SA145 | ΔcrtM-null mutant | 0.75 |
| SA146 | ΔcrtM-pDCerm (empty plasmid vector) | 0.75 |
| SA147 | ΔcrtM-pDCerm::crtMN complemented/overexpressing | 1.5 |
TABLE 2.
| Strain | % survival (mean ± SD)a after 2-h exposure to indicated concn (μg/ml) of: | ||||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| tPMP | RP-1 | hNP-1 | PMB | DAP | |||||||||||||||
| 0.25 | 0.125 | 5 | 2.5 | 20 | 10 | 80 | 40 | 1.25 | 0.625 | ||||||||||
| SA144 | 4 ± 4 | 7 ± 1 | 11 ± 1 | 23 ± 9 | 20 ± 7* | 60 ± 2 | 13 ± 11 | 29 ± 4 | 12 ± 6 | 18 ± 14 | |||||||||
| SA145 | 0 ± 0 | 2 ± 2 | 13 ± 2 | 18 ± 4 | 27 ± 13 | 60 ± 2 | 14 ± 7 | 23 ± 9 | 21 ± 15 | 24 ± 9 | |||||||||
| SA147 | 8 ± 2 | 14 ± 1 | 23 ± 9 | 55 ± 12 | 52 ± 13** | 66 ± 2 | 26 ± 13 | 25 ± 6 | 49 ± 15 | 55 ± 7 | |||||||||
a
*, P = 0.0037 (SA144 versus SA147); **, P = 0.087 (SA145 versus SA 147).
TABLE 3.
| Strain | % of total phospholipid content and fatty acid (mean ± SD)a | ||||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Inner* LPG | Outer* LPG | Total LPG | PG | CL | SCFA | BCFA | Total iso | Total anteiso | UI | ||||||||||
| SA144 | 15.38 ± 0.65 | 2.08 ± 0.05 | 17.45 ± 0.70 | 74.28 ± 0.54 | 8.27 ± 0.15 | 15.91 ± 0.11 | 74.07 ± 0.63 | 14.19 ± 0.17 | 60.3 ± 1.07 | 0.044 ± 0.00 | |||||||||
| SA145 | 15.24 ± 2.29 | 2.63 ± 1.46 | 17.86 ± 0.83 | 69.92 ± 5.30 | 12.22 ± 6.12 | 15.61 ± 0.13 | 73.35 ± 2.16 | 14.05 ± 0.31 | 59.3 ± 1.85 | 0.046 ± 0.00 | |||||||||
| SA147 | 15.04 ± 1.51 | 3.83 ± 1.37 | 18.86 ± 0.15 | 69.42 ± 8.94 | 11.72 ± 9.08 | 15.97 ± 0.36 | 75.42 ± 0.70 | 15.31 ± 0.57 | 60.5 ± 0.66 | 0.044 ± 0.00 | |||||||||
a
*, membrane leaflet; SCFA, straight chain fatty acid; BCFA, branched chain fatty acid; UI, unsaturation index.
Acknowledgments
This research was supported by National Institutes of Health grants AI-39108 to A.S.B., AI-74832 to G.Y.L., and AI-39001 and AI-48031 to M.R.Y.
REFERENCES
1.
Bayer, A. S., et al. 2000. In vitro resistance of Staphylococcus aureus to thrombin-induced microbicidal protein is associated with alterations in membrane fluidity. Infect. Immun. 68:3548-3553.
2.
Bayer, A. S., et al. 2006. Low-level resistance of Staphylococcus aureus to thrombin-induced platelet microbicidal protein 1 in vitro associated with qacA gene carriage is independent of multidrug efflux pump activity. Antimicrob. Agents Chemother. 50:2448-2454.
3.
Bourbigot, S., et al. 2009. Antimicrobial peptide RP-1 structure and interactions with anionic versus zwitterionic micelles. Biopolymers 91:1-13.
4.
Clauditz, A., A. Resch, K. P. Wieland, A. Peschel, and F. Götz. 2006. Staphyloxanthin plays a role in the fitness of Staphylococcus aureus and its ability to cope with oxidative stress. Infect. Immun. 74:4950-4953.
5.
Cui, L., E. Tominaga, H. M. Neoh, and K. Hiramatsu. 2006. Correlation between reduced daptomycin susceptibility and vancomycin resistance in vancomycin-intermediate Staphylococcus aureus. Antimicrob. Agents Chemother. 50:1079-1082.
6.
El-Agamey, A., et al. 2004. Carotenoid radical chemistry and antioxidant/pro-oxidant properties. Arch. Biochem. Biophys. 430:37-48.
7.
Gruszecki, W. I. 2004. Carotenoids in health and disease, p. 151-163. In N. I. Krinsky, S. T. Mayne, and H. Sies (ed.), Carotenoid orientation: role in membrane stabilization. Marcel Dekker, New York, NY.
8.
Gruszecki, W. I. 1999. The photochemistry of carotenoids, p. 363-379. In H. A. Frank, A. J. Young, G. Britton, and R. J. Cogdell (ed.), Carotenoid in membranes. Kluwer Academic, Dordrecht, Netherlands.
9.
Gruszecki, W. I., and K. Strzalka. 1991. Does the xanthophyll cycle take part in the regulation of fluidity of the thylakoid membrane? Biochim. Biophys. Acta 1060:310-314.
10.
Hartmann, W., and H. J. Galla. 1978. Binding of polylysine to charged bilayer membranes: molecular organization of a lipid peptide complex. Biochim. Biophys. Acta 509:474-490.
11.
Jeng, A., et al. 2003. Molecular genetic analysis of a group A Streptococcus operon encoding serum opacity factor and a novel fibronectin-binding protein, SfbX. J. Bacteriol. 185:1208-1217.
12.
Jones, T., et al. 2008. Failures in clinical treatment of Staphylococcus aureus infection with daptomycin are associated with alterations in surface charge, membrane phospholipid asymmetry, and drug binding. Antimicrob. Agents Chemother. 52:269-278.
13.
Joyce, G. H., R. K. Hammond, and D. C. White. 1970. Changes in membrane lipid composition in exponentially growing Staphylococcus aureus during the shift from 37° to 25°C. J. Bacteriol. 104:323-330.
14.
Katzif, S., D. Danavall, S. Bowers, J. T. Balthazar, and W. M. Shafer. 2003. The major cold shock gene, cspA, is involved in the susceptibility of Staphylococcus aureus to an antimicrobial peptide of human cathepsin G. Infect. Immun. 71:4304-4312.
15.
Katzif, S., E. H. Lee, A. B. Law, Y. L. Tzeng, and W. M. Shafer. 2005. CspA regulates pigment production in Staphylococcus aureus through a SigB-dependent mechanism. J. Bacteriol. 187:8181-8184.
16.
Kim, J., M. Mosior, L. A. Chung, H. Wu, and S. McLaughlin. 1991. Binding of peptides with basic residues to membranes containing acidic phospholipids. Biophys. J. 60:135-148.
17.
Koo, S.-P., M. R. Yeaman, C. C. Nast, and A. S. Bayer. 1997. The cytoplasmic membrane is a primary target for the staphylocidal action of thrombin-induced platelet microbicidal protein. Infect. Immun. 65:4795-4800.
18.
Lennette, E. H., A. Balows, W. J. Hausler, Jr., and H. J. Shadomy. 1985. Manual of clinical microbiology, 4th ed., p. 146. American Society for Microbiology, Washington, DC.
19.
Liu, C. I., et al. 2008. Cholesterol biosynthesis inhibitor blocks Staphylococcus aureus virulence. Science 319:1391-1394.
20.
Liu, G. Y., et al. 2005. Staphylococcus aureus golden pigment impairs neutrophil killing and promotes virulence through its antioxidant activity. J. Exp. Med. 202:209-215.
21.
Liu, G. Y., and V. Nizet. 2009 Color me bad: microbial pigments as virulence factors. Trends Microbiol. 17:406-413.
22.
Marshall, J. H., and G. J. Wilmoth. 1981. Pigments of Staphylococcus aureus, a series of triterpenoid carotenoids. J. Bacteriol. 147:900-913.
23.
Mishra, N. N., et al. 2009. Analysis of cell membrane characteristics of in vitro-selected daptomycin-resistant strains of methicillin-resistant Staphylococcus aureus. Antimicrob. Agents Chemother. 53:2312-2318.
23a.
Mishra, N. N., G. Y. Liu, M. R. Yeaman, S. J. Yang, and A. S. Bayer. 2009. Carotenoid induced alteration of cell membrane (CM) fluidity impacts Staphylococcus aureus susceptibility to innate host defense peptides in vitro, abstr. C1-1362, p. 75. Abstr. 49th Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.
24.
Mukhopadhyay, K., et al. 2007. Reduced in vitro susceptibility of Staphylococcus aureus to thrombin-induced platelet microbicidal protein-1 (tPMP-1) is associated with alterations in cell membrane phospholipid composition and asymmetry. Microbiology 153:1187-1197.
25.
Pelz, A., et al. 2005. Structure and biosynthesis of staphyloxanthin from Staphylococcus aureus. J. Biol. Chem. 280:32493-32498.
26.
Rohmer, M., P. Bouvier, and G. Ourisson. 1979. Molecular evolution of membranes: structural equivalents and phylogenetic precursors of sterols. Proc. Natl. Acad. Sci. U. S. A. 76:847-851.
27.
Subczynski, W. K., E. Markowska, W. I. Gruszecki, and J. Sielewiesiuk. 1992. Effects of polar carotenoids on dimyristoylphosphatidylcholine membranes: a spin-label study. Biochim. Biophys. Acta 1105:97-108.
28.
Subczynski, W. K., and A. Wisniewska. 2000. Physical properties of lipid bilayer membranes: relevance to membrane biological functions. Acta Biochim. Pol. 47:613-625.
29.
Widomska, J., A. Kostecka-Gugała, D. Latowski, W. I. Gruszecki, and K. Strzałka. 2009. Calorimetric studies of the effect of cis-carotenoids on the thermotropic phase behavior of phosphatidylcholine bilayers. Biophys. Chem. 140:108-114.
30.
Wisniewska, A. J., J. Widomska, and W. K. Subczynski. 2006. Carotenoid-membrane interactions in liposomes: effect of dipolar, monopolar, and nonpolar carotenoids. Acta Biochim. Pol. 53:475-484.
31.
Wisniewska, A., and W. K. Subczynski. 1998. Effects of polar carotenoids on the shape of the hydrophobic barrier of phospholipid bilayers. Biochim. Biophys. Acta 1368:235-246.
32.
Xiong, Y. Q., A. S. Bayer, and M. R. Yeaman. 2002. Inhibition of intracellular macromolecular synthesis in Staphylococcus aureus by thrombin-induced platelet microbicidal proteins. J. Infect. Dis. 185:348-356.
33.
Xiong, Y. Q., K. Mukhopadhyay, M. R. Yeaman, J. Adler-Moore, and A. S. Bayer. 2005. Functional interrelationships between cell membrane and cell wall in antimicrobial peptide-mediated killing of Staphylococcus aureus. Antimicrob. Agents Chemother. 49:3114-3121.
34.
Yang, S. J., et al. 2009. Enhanced expression of dltABCD is associated with the development of daptomycin nonsusceptibility in a clinical endocarditis isolate of Staphylococcus aureus. J. Infect. Dis. 200:1916-1920.
35.
Yeaman, M. R., K. D. Gank, A. S. Bayer, and E. P. Brass. 2002. Synthetic peptides that exert antimicrobial activities in whole blood and blood-derived matrices. Antimicrob. Agents Chemother. 46:3883-3891.
36.
Yeaman, M. R., Y. Q. Tang, A. J. Shen, A. S. Bayer, and M. E. Selsted. 1997. Purification and in vitro activities of rabbit platelet microbicidal proteins. Infect. Immun. 65:1023-1031.
Information & Contributors
Information
Published In
Antimicrobial Agents and Chemotherapy
Volume 55 • Number 2 • February 2011
Pages: 526 - 531
PubMed: 21115796
Copyright
© 2011 American Society for Microbiology.
History
Received: 19 May 2010
Revision received: 7 July 2010
Accepted: 20 October 2010
Published online: 20 January 2011
Contributors
Metrics & Citations
Metrics
Note:
- For recently published articles, the TOTAL download count will appear as zero until a new month starts.
- There is a 3- to 4-day delay in article usage, so article usage will not appear immediately after publication.
- Citation counts come from the Crossref Cited by service.
Citations
Citation
2011. Carotenoid-Related Alteration of Cell Membrane Fluidity Impacts Staphylococcus aureus Susceptibility to Host Defense Peptides . Antimicrob Agents Chemother 55:.
If you have the appropriate software installed, you can download article citation data to the citation manager of your choice. For an editable text file, please select Medlars format which will download as a .txt file. Simply select your manager software from the list below and click Download.