Carotenoid Content and Composition in Exponential, Stationary and Biofilm States of Staphylococcus aureus and their Influence on Membrane Biophysical Properties

AbstractStaphyloxanthin (STX) is a saccharolipid derived from a carotenoid in Staphylococcus aureus involved in oxidative-stress tolerance and antimicrobial peptide resistance. In this work, a targeted metabolomics and biophysical study was carried out on native and knock-out S. aureus strains to investigate the biosynthetic pathways of STX and related carotenoids. Identification of 34 metabolites at different growth phases (8, 24 and 48h), reveal shifts of carotenoid populations during progression towards stationary phase. Six of the carotenoids in the STX biosynthetic pathway and three menaquinones (Vitamin K2) were identified in the same chromatogram. Furthermore, other STX homologues with varying acyl chain structures reported herein for the first time, which reveal the extensive enzymatic activity of CrtO/CrtN. Fourier Transform infrared spectroscopy show that STX increases acyl chain order and shifts the cooperative melting of the membrane indicating a more rigid lipid bilayer. This study shows the diversity of carotenoids in S. aureus, and their influence on membrane biophysical properties.

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Impact of Bicarbonate on PBP2a Production, Maturation, and Functionality in Methicillin-Resistant Staphylococcus aureus (MRSA)

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02621-20 ◽

2021 ◽

Author(s):

Selvi C. Ersoy ◽

Henry F. Chambers ◽

Richard A. Proctor ◽

Adriana E. Rosato ◽

Nagendra N. Mishra ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Ex Vivo ◽

Carotenoid Content ◽

Methicillin Resistant ◽

Genes Encoding ◽

Regulatory Impact ◽

Mrsa Strains ◽

The Impact

Certain methicillin-resistant Staphylococcus aureus (MRSA) strains exhibit β-lactam-susceptibility in vitro, ex vivo and in vivo in the presence of NaHCO3 (NaHCO3-responsive MRSA). Herein, we investigate the impact of NaHCO3 on factors required for PBP2a functionality. Prototype NaHCO3-responsive and -nonresponsive MRSA strains (as defined in vitro) were assessed for the impact of NaHCO3 on: expression of genes involved in PBP2a production-maturation pathways (mecA, blaZ, pbp4, vraSR, prsA, sigB, and floA); membrane PBP2a and PrsA protein content; and membrane carotenoid content. Following NaHCO3 exposure in NaHCO3-responsive (vs - nonresponsive) MRSA, there was significantly reduced expression of: i) mecA and blaZ; ii) the vraSR-prsA gene axis; and iii) pbp4. Carotenoid production was reduced, while floA expression was increased by NaHCO3 exposure in all MRSA strains. This work underscores the distinct regulatory impact of NaHCO3 on a cadre of genes encoding factors required for maintenance of the MRSA phenotype through PBP2a functionality and maturation.

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Carotenoid-Related Alteration of Cell Membrane Fluidity ImpactsStaphylococcus aureusSusceptibility to Host Defense Peptides

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00680-10 ◽

2010 ◽

Vol 55 (2) ◽

pp. 526-531 ◽

Cited By ~ 117

Author(s):

Nagendra N. Mishra ◽

George Y. Liu ◽

Michael R. Yeaman ◽

Cynthia C. Nast ◽

Richard A. Proctor ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Cell Membrane ◽

Host Defense ◽

Phospholipid Composition ◽

Carotenoid Content ◽

Clinical Strain ◽

Cationic Peptides ◽

Multicopy Plasmid ◽

Host Defenses ◽

Host Defense Peptides

ABSTRACTCarotenoid pigments ofStaphylococcus aureusprovide 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, andin vitrosusceptibility 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-susceptibleStaphylococcus aureus(MSSA) isogenic clinical strain set was used, including a parental isolate with an intact carotenoid biosynthetic operon (crtOPQMN) containing thecrtMgene encoding early steps in staphyloxanthin biosynthesis, acrtMdeletion mutant, and acrtMNmulticopy plasmid-complemented variant. Compared to the parental andcrtMknockout strains, thecrtMN-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 ofS. aureusto subvert nonoxidative host defenses mediated by cationic peptides, potentially by increasing target membrane rigidity.

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Combination Antibiotic Exposure Selectively Alters the Development of Vancomycin Intermediate Resistance in Staphylococcus aureus

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02100-17 ◽

2017 ◽

Vol 62 (2) ◽

Cited By ~ 5

Author(s):

Xuting Zheng ◽

Andrew D. Berti ◽

Sue McCrone ◽

Melanie Roch ◽

Adriana E. Rosato ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Membrane Fluidity ◽

Susceptibility Testing ◽

Carotenoid Content ◽

Antibiotic Exposure ◽

Rational Selection ◽

Content Type ◽

Selection Of

ABSTRACTInvasive methicillin-resistantStaphylococcus aureus(MRSA) treated with vancomycin (VAN) is associated with reduced VAN susceptibility and treatment failure. VAN combination therapy is one strategy to improve response, but comprehensive assessments of combinations to prevent resistance are limited. This study identifies optimal combinations to prevent the emergence of VAN-intermediateStaphylococcus aureus(VISA). Two standard MRSA and two heterogeneous VISA (hVISA) strains were exposed for 28 daysin vitroto VAN alone, VAN with cefazolin (CFZ), fosfomycin, gentamicin, meropenem, rifampin, piperacillin-tazobactam (TZP), or trimethoprim-sulfamethoxazole. In addition to VAN susceptibility testing, cell wall thickness (CWT), carotenoid content, and membrane fluidity were determined for Mu3. VAN plus any β-lactam limited the VAN MIC increase to 1 to 4 mg/liter throughout the 28-day exposure, with CFZ and TZP being the most effective agents (VAN MIC = 1 to 2 mg/liter). Similar MIC trends occurred with the lipo-/glycopeptide agents daptomycin and telavancin, where β-lactam combinations with VAN prevented MIC increases to these agents as well. Combinations with non-β-lactams were ineffective in preventing VAN MIC increases with VAN MICs of 4 to 16 mg/liter emerging during weeks 2 to 4 of treatment. VAN plus β-lactam decreased CWT significantly, whereas VAN plus other antibiotics significantly increased the CWT. No correlation was observed between carotenoid content or membrane fluidity and antibiotic exposure. Only the combination exposures of VAN plus β-lactam suppress the development of VISA. Rational selection of VAN plus β-lactam should be further explored as a long-term combination treatment of MRSA infections due to their ability to suppress VAN resistance.

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Cytochemical Studies of Cell Wall Biosynthesis in Staphylococcus Aureus

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100094905 ◽

1972 ◽

Vol 30 ◽

pp. 328-329

Author(s):

Masaatsu Koike ◽

Koichi Nakashima ◽

Kyoko Iida

Keyword(s):

Staphylococcus Aureus ◽

Periodate Oxidation ◽

Early Time ◽

The Other ◽

Penicillin G ◽

Cell Wall Biosynthesis ◽

Septal Wall ◽

Peptidoglycan Biosynthesis ◽

Gram Positive Bacteria ◽

Cross Linkage

Penicillin exerts the activity to inhibit the peptide cross linkage between each polysaccharide backbone at the final stage of wall-peptidoglycan biosynthesis of bacteria. Morphologically, alterations of the septal wall and mesosome in gram-positive bacteria, which were occurred in early time after treatment with penicillin, have been observed. In this experiment, these alterations were cytochemically investigated by means of silver-methenamine staining after periodate oxidation, which is applied for detection of localization of wall mucopolysaccharide.Staphylococcus aureus strain 209P treated with 100 u/ml of penicillin G was divided into two aliquotes. One was fixed by Kellenberger-Ryter's OSO4 fixative at 30, 60 and 120 min after addition of the antibiotic, dehydrated through alcohol series, and embedded in Epon 812 (Specimen A). The other was fixed by 21 glutaraldehyde, dehydrated through glycolmethacrylate series and embedded in glycolmethacrylate mixture, according to Bernhard's method (Specimen B).

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Gold labeling of teichoic acid on adjacent surfaces of staphylococcus aureus

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100160443 ◽

1990 ◽

Vol 48 (3) ◽

pp. 578-579

Author(s):

Margaret Hukee

Keyword(s):

Staphylococcus Aureus ◽

Teichoic Acid ◽

Protein Labeling ◽

Minimal Amount ◽

Section Thickness ◽

Double Labeling ◽

Parallel Surfaces ◽

Supporting Membrane

Gold labeling of two antigens (double labeling) is often done on two section surfaces separated by section thickness. Whether labeling is done on both sides of the same section or on two parallel surfaces separated by section thickness (PSSST), comparable results are dependent on an equal number of epitopes being exposed at each surface. We propose a method to study protein labeling within the same field of proteins, by examining two directly adjacent surfaces that were split during sectioning. The number of labeling sites on adjacent surfaces (AS) were compared to sites on PSSST surfaces in individual bacteria.Since each bacteria needed to be recognizable in all three section surfaces, one-hole grids were used for labeling. One-hole grids require a supporting membrane and excessive handling during labeling often ruptures the membrane. To minimize handling, a labeling chamber was designed that is inexpensive, disposable, minimizes contamination, and uses a minimal amount of solution.

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