scholarly journals Oleate Hydratase (OhyA) Is a Virulence Determinant in Staphylococcus aureus

2021 ◽  
Author(s):  
Christopher D. Radka ◽  
Justin L. Batte ◽  
Matthew W. Frank ◽  
Jason W. Rosch ◽  
Charles O. Rock
Keyword(s):  
Fatty Acids ◽  
Staphylococcus Aureus ◽  
Unsaturated Fatty Acids ◽  
Commensal Bacteria ◽  
Hydroxy Fatty Acids ◽  
Infection Model ◽  
Infection Site ◽  
Human Pathogen ◽  
Content Type ◽  
Oleate Hydratase

The oleate hydratase protein family was discovered in commensal bacteria that utilize host unsaturated fatty acids as the substrates to produce a spectrum of hydroxylated products. These hydroxy fatty acids are thought to act as signaling molecules that suppress the inflammatory response to create a more tolerant environment for the microbiome. S. aureus is a significant human pathogen, and defining the mechanisms used to evade the immune response is critical to understanding pathogenesis. S. aureus expresses an OhyA that produces at least three 10-hydroxy fatty acids from host unsaturated fatty acids at the infection site, and an S. aureus strain lacking the ohyA gene has compromised virulence in an immunocompetent infection model.

scholarly journals Comparison of Biochemical Properties of the Original and Newly Identified Oleate Hydratases from Stenotrophomonas maltophilia

2017 ◽  
Vol 83 (9) ◽  
Author(s):  
Woo-Ri Kang ◽  
Min-Ju Seo ◽  
Kyung-Chul Shin ◽  
Jin-Byung Park ◽  
Deok-Kun Oh
Keyword(s):  
Fatty Acids ◽  
Oleic Acid ◽  
Linoleic Acid ◽  
Linolenic Acid ◽  
Stenotrophomonas Maltophilia ◽  
Unsaturated Fatty Acids ◽  
Hydroxy Fatty Acids ◽  
Plant Oils ◽  
Content Type ◽  
Oleate Hydratase

ABSTRACT Oleate hydratases (OhyAs) catalyze the conversion of unsaturated fatty acids to 10-hydroxy fatty acids, which are used as precursors of important industrial compounds, including lactones and ω-hydroxycarboxylic and α,ω-dicarboxylic acids. The genes encoding OhyA and a putative fatty acid hydratase in Stenotrophomonas maltophilia were identified by genomic analysis. The putative fatty acid hydratase was purified and identified as an oleate hydratase (OhyA2) based on its substrate specificity. The activity of OhyA2 as a holoenzyme was not affected by adding cofactors, whereas the activity of the original oleate hydratase (OhyA1) showed an increase. Thus, all characterized OhyAs were categorized as either OhyA1 or OhyA2 based on the activities of holoenzymes upon adding cofactors, which were determined by the type of the fourth conserved amino acid of flavin adenine dinucleotide (FAD)-binding motif. The hydration activities of S. maltophilia OhyA2 toward unsaturated fatty acids, including oleic acid, palmitoleic acid, linoleic acid, α-linolenic acid, and γ-linolenic acid, were greater than those of OhyA1. Moreover, the specific activity of S. maltophilia OhyA2 toward unsaturated fatty acids, with the exception of γ-linolenic acid, was the highest among all reported OhyAs. IMPORTANCE All characterized OhyAs were categorized as OhyA1s or OhyA2s based on the different properties of the reported and newly identified holo-OhyAs in S. maltophilia upon the addition of cofactors. OhyA2s showed higher activities toward polyunsaturated fatty acids (PUFAs), including linoleic acid, α-linolenic acid, and γ-linolenic acid, than those of OhyA1s. This suggests that OhyA2s can be used more effectively to convert plant oils to 10-hydroxy fatty acids because plant oils contain not only oleic acid but also PUFAs. The hydration activity of the newly identified OhyA2 from S. maltophilia toward oleic acid was the highest among the activity levels reported so far. Therefore, this enzyme is an efficient biocatalyst for the conversion of plant oils to 10-hydroxy fatty acids, which can be further converted to important industrial materials.

scholarly journals Role of the Tet38 Efflux Pump in Staphylococcus aureus Internalization and Survival in Epithelial Cells

2015 ◽  
Vol 83 (11) ◽  
pp. 4362-4372 ◽  
Author(s):  
Q. C. Truong-Bolduc ◽  
G. R. Bolduc ◽  
H. Medeiros ◽  
J. M. Vyas ◽  
Y. Wang ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Staphylococcus Aureus ◽  
Epithelial Cells ◽  
Unsaturated Fatty Acids ◽  
Efflux Pump ◽  
Fold Increase ◽  
Growth Patterns ◽  
Heart Cell ◽  
Content Type

ABSTRACTWe previously identified the protein Tet38 as a chromosomally encoded efflux pump ofStaphylococcus aureusthat confers resistance to tetracycline and certain unsaturated fatty acids. Tet38 also contributes to mouse skin colonization. In this study, we discovered a novel regulator oftet38, named tetracycline regulator 21 (TetR21), that bound specifically to thetet38promoter and repressed pump expression. A ΔtetR21mutant showed a 5-fold increase intet38transcripts and an 8-fold increase in resistance to tetracycline and fatty acids. The global regulator MgrA bound to thetetR21promoter and indirectly repressed the expression oftet38. To further assess the full role of Tet38 inS. aureusadaptability, we tested its effect on host cell invasion using A549 (lung) and HMEC-1 (heart) cell lines. We usedS. aureusRN6390, its Δtet38, ΔtetR21, and ΔmgrAmutants, and a Δtet38 ΔtetR21double mutant. After 2 h of contact, the Δtet38mutant was internalized in 6-fold-lower numbers than RN6390 in A549 and HMEC-1 cells, and the ΔtetR21mutant was internalized in 2-fold-higher numbers than RN6390. A slight increase of 1.5-fold in internalization was found for the ΔmgrAmutant. The growth patterns of RN6390 and the ΔmgrAand ΔtetR21mutants within A549 cells were similar, while no growth was observed for the Δtet38mutant. These data indicate that the Tet38 efflux pump is regulated by TetR21 and contributes to the ability ofS. aureusto internalize and replicate within epithelial cells.

scholarly journals DNA Binding and Sensor Specificity of FarR, a Novel TetR Family Regulator Required for Induction of the Fatty Acid Efflux Pump FarE in Staphylococcus aureus

2018 ◽  
Vol 201 (3) ◽  
Author(s):  
Heba Alnaseri ◽  
Robert C. Kuiack ◽  
Katherine A. Ferguson ◽  
James E. T. Schneider ◽  
David E. Heinrichs ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Staphylococcus Aureus ◽  
Fatty Acid ◽  
Dna Binding ◽  
Unsaturated Fatty Acids ◽  
Efflux Pump ◽  
Ectopic Expression ◽  
Exogenous Fatty Acid ◽  
Content Type ◽  
Tetr Family Regulator

ABSTRACT Divergent genes in Staphylococcus aureus USA300 encode the efflux pump FarE and TetR family regulator FarR, which confer resistance to antimicrobial unsaturated fatty acids. To study their regulation, we constructed USA300 ΔfarER, which exhibited a 2-fold reduction in MIC of linoleic acid. farE expressed from its native promoter on pLIfarE conferred increased resistance to USA300 but not USA300 ΔfarER. Complementation of USA300 ΔfarER with pLIfarR also had no effect, whereas resistance was restored with pLIfarER or through ectopic expression of farE. In electrophoretic mobility shift assays, FarR bound to three different oligonucleotide probes that each contained a TAGWTTA motif, occurring as (i) a singular motif overlapping the −10 element of the PfarR promoter, (ii) in palindrome PAL1 immediately in the 3′ direction of PfarR, or (iii) within PAL2 upstream of the predicted PfarE promoter. FarR autorepressed its expression through cooperative binding to PAL1 and the adjacent TAGWTTA motif in PfarR. Consistent with reports that S. aureus does not metabolize fatty acids through acyl coenzyme A (acyl-CoA) intermediates, DNA binding activity of FarR was not affected by linoleoyl-CoA. Conversely, induction of farE required fatty acid kinase FakA, which catalyzes the first metabolic step in the incorporation of unsaturated fatty acids into phospholipid. We conclude that FarR is needed to promote the expression of farE while strongly autorepressing its own expression, and our data are consistent with a model whereby FarR interacts with a FakA-dependent product of exogenous fatty acid metabolism to ensure that efflux only occurs when the metabolic capacity for incorporation of fatty acid into phospholipid is exceeded. IMPORTANCE Here, we describe the DNA binding and sensor specificity of FarR, a novel TetR family regulator (TFR) in Staphylococcus aureus. Unlike the majority of TFRs that have been characterized, which function to repress a divergently transcribed gene, we find that FarR is needed to promote expression of the divergently transcribed farE gene, encoding a resistance-nodulation-division (RND) family efflux pump that is induced in response to antimicrobial unsaturated fatty acids. Induction of farE was dependent on the function of the fatty acid kinase FakA, which catalyzes the first metabolic step in the incorporation of exogenous unsaturated fatty acids into phospholipid. This represents a novel example of TFR function.

scholarly journals Exogenous Fatty Acids Remodel Staphylococcus aureus Lipid Composition through Fatty Acid Kinase

2020 ◽  
Vol 202 (14) ◽  
Author(s):  
Zachary DeMars ◽  
Vineet K. Singh ◽  
Jeffrey L. Bose
Keyword(s):  
Fatty Acids ◽  
Staphylococcus Aureus ◽  
Oleic Acid ◽  
Fatty Acid ◽  
Lipid Composition ◽  
Unsaturated Fatty Acids ◽  
Mouse Skin ◽  
Membrane Composition ◽  
Wild Type ◽  
Content Type

ABSTRACT Staphylococcus aureus can utilize exogenous fatty acids for phospholipid synthesis. The fatty acid kinase FakA is essential for this utilization by phosphorylating exogenous fatty acids for incorporation into lipids. How FakA impacts the lipid membrane composition is unknown. In this study, we used mass spectrometry to determine the membrane lipid composition and properties of S. aureus in the absence of fakA. We found the fakA mutant to have increased abundance of lipids containing longer acyl chains. Since S. aureus does not synthesize unsaturated fatty acids, we utilized oleic acid (18:1) to track exogenous fatty acid incorporation into lipids. We observed a concentration-dependent incorporation of exogenous fatty acids into the membrane that required FakA. We also tested how FakA and exogenous fatty acids impact membrane-related physiology and identified changes in membrane potential, cellular respiration, and membrane fluidity. To mimic the host environment, we characterized the lipid composition of wild-type and fakA mutant bacteria grown in mouse skin homogenate. We show that wild-type S. aureus can incorporate exogenous unsaturated fatty acids from host tissue, highlighting the importance of FakA in the presence of host skin tissue. In conclusion, FakA is important for maintaining the composition and properties of the phospholipid membrane in the presence of exogenous fatty acids, impacting overall cell physiology. IMPORTANCE Environmental fatty acids can be harvested to supplement endogenous fatty acid synthesis to produce membranes and circumvent fatty acid biosynthesis inhibitors. However, how the inability to use these fatty acids impacts lipids is unclear. Our results reveal lipid composition changes in response to fatty acid addition and when S. aureus is unable to activate fatty acids through FakA. We identify concentration-dependent utilization of oleic acid that, when combined with previous work, provides evidence that fatty acids can serve as a signal to S. aureus. Furthermore, using mouse skin homogenates as a surrogate for in vivo conditions, we showed that S. aureus can incorporate host fatty acids. This study highlights how exogenous fatty acids impact bacterial membrane composition and function.

scholarly journals Skin-Specific Unsaturated Fatty Acids Boost the Staphylococcus aureus Innate Immune Response

2015 ◽  
Vol 84 (1) ◽  
pp. 205-215 ◽  
Author(s):  
Minh Thu Nguyen ◽  
Dennis Hanzelmann ◽  
Thomas Härtner ◽  
Andreas Peschel ◽  
Friedrich Götz
Keyword(s):  
Fatty Acids ◽  
Staphylococcus Aureus ◽  
Human Skin ◽  
Immune Activation ◽  
Pathogenic Bacteria ◽  
Unsaturated Fatty Acids ◽  
Innate Immune ◽  
Immune Stimulation ◽  
Content Type ◽  
Lipid Moiety

Antimicrobial fatty acids (AFAs) protect the human epidermis against invasion by pathogenic bacteria. In this study, we questioned whether human skin fatty acids (FAs) can be incorporated into the lipid moiety of lipoproteins and whether such incorporation would have an impact on innate immune stimulation in the model organismStaphylococcus aureusUSA300 JE2. This organism synthesized only saturated FAs. However, when feeding USA300 with unsaturated FAs present on human skin (C16:1, C18:1, or C18:2), those were taken up, elongated stepwise by two carbon units, and finally found in the bacterial (phospho)lipid fraction. They were also observed in the lipid moiety of lipoproteins. When USA300 JE2 was fed with the unsaturated FAs, the cells and cell lysates showed an increased innate immune activation with various immune cells and peripheral blood mononuclear cells (PBMCs). Immune activation was highest with linoleic acid (C18:2). There are several pieces of evidence that the enhanced immune stimulating effect was due to the incorporation of unsaturated FAs in lipoproteins. First, the enhanced stimulation was dependent on Toll-like receptor 2 (TLR2). Second, anlgtmutant, unable to carry out lipidation of prolipoproteins, was unable to carry out immune stimulation when fed with unsaturated FAs. Third, the supplied FAs did not significantly affect growth, protein release, or expression of the model lipoprotein Lpl1. AlthoughS. aureusis unable to synthesize unsaturated FAs, it incorporates long-chain unsaturated FAs into its lipoproteins, with the effect that the cells are better recognized by the innate immune system. This is an additional mechanism how our skin controls bacterial colonization and infection.

scholarly journals Cyclobacterium qasimii sp. nov., a psychrotolerant bacterium isolated from Arctic marine sediment

2012 ◽  
Vol 62 (Pt_9) ◽  
pp. 2133-2139 ◽  
Author(s):  
S. Shivaji ◽  
P. Vishnu Vardhan Reddy ◽  
S. S. S. Nageshwara Rao ◽  
Zareena Begum ◽  
Poorna Manasa ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Marine Sediment ◽  
Dna Hybridization ◽  
Type Species ◽  
Unsaturated Fatty Acids ◽  
Rrna Gene ◽  
Respiratory Quinone ◽  
Content Type ◽  
Link Type ◽  
Sequence Similarities

A novel Gram-stain-negative, horseshoe-shaped, non-motile bacterium, designated strain M12-11BT, was isolated from a marine sediment sample collected at a depth of 200 m from Kongsfjorden, Svalbard. The colony colour was orangish red due to the presence of carotenoids. Fatty acids were dominated by branched and unsaturated fatty acids (90.8 %), with a high abundance of iso-C15 : 0 (14.9 %), anteiso-C15 : 0 (11.4 %), iso-C15 : 1 G (13.1 %), C15 : 1ω6c (5.4 %), C17 : 1ω6c (6.7 %), summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c; 9.3 %) and summed feature 9 (10-methyl C16 : 0 and/or iso-C17 : 1ω9c; 5.9 %). Strain M12-11BT contained MK-7 as the major respiratory quinone. The polar lipids consisted of phosphatidylcholine, phosphatidylethanolamine, one unidentified aminolipid and three unidentified lipids. Based on 16S rRNA gene sequence similarities, the type strains of Cyclobacterium amurskyense , Cyclobacterium marinum and Cyclobacterium lianum were most closely related to M12-11BT with sequence similarities of 98.2, 96.8 and 93.3 %, respectively. Other members of the family Cyclobacteriaceae had sequence similarities of <92.0 %. However, DNA–DNA hybridization with Cyclobacterium amurskyense KCTC 12363T and Cyclobacterium marinum DSM 745T showed relatedness values of only 24.5 and 32.5 % with respect to strain M12-11BT. Based on the results of DNA–DNA hybridization experiments and phenotypic and chemotaxonomic data, it appears that strain M12-11BT represents a novel species of the genus Cyclobacterium , for which the name Cyclobacterium qasimii sp. nov. is proposed; the type strain is M12-11BT ( = KCTC 23011T = NBRC 106168T) and it has a DNA G+C content of 40.5 mol%.

scholarly journals Decreased Vancomycin Susceptibility in Staphylococcus aureus Caused by IS256Tempering of WalKR Expression

2013 ◽  
Vol 57 (7) ◽  
pp. 3240-3249 ◽  
Author(s):  
Christopher R. E. McEvoy ◽  
Brian Tsuji ◽  
Wei Gao ◽  
Torsten Seemann ◽  
Jessica L. Porter ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Fluorescent Protein ◽  
Site Directed Mutagenesis ◽  
Infection Model ◽  
Content Type ◽  
Reverse Transcription Pcr ◽  
Phenotype Analysis ◽  
Dosing Regimens ◽  
Mrsa Strains ◽  
Green Fluorescent

ABSTRACTVancomycin-intermediateStaphylococcus aureus(VISA) strains often arise by mutations in the essential two-component regulatorwalKR; however their impact onwalKRfunction has not been definitively established. Here, we investigated 10 MRSA strains recovered serially after exposure of vancomycin-susceptibleS. aureus(VSSA) JKD6009 to simulated human vancomycin dosing regimens (500 mg to 4,000 mg every 12 h) using a 10-day hollow fiber infection model. After continued exposure to the vancomycin regimens, two isolates displayed reduced susceptibility to both vancomycin and daptomycin, developing independent IS256insertions in thewalKR5′ untranslated region (5′ UTR). Quantitative reverse transcription-PCR (RT-PCR) revealed a 50% reduction inwalKRgene expression in the IS256mutants compared to the VSSA parent. Green fluorescent protein (GFP) reporter analysis, promoter mapping, and site-directed mutagenesis confirmed these findings and showed that the IS256insertions had replaced two SigA-likewalKRpromoters with weaker, hybrid promoters. Removal of IS256reverted the phenotype to VSSA, showing that reduced expression of WalKR did induce the VISA phenotype. Analysis of selected WalKR-regulated autolysins revealed upregulation ofssaAbut no change in expression ofsakandsceDin both IS256mutants. Whole-genome sequencing of the two mutants revealed an additional IS256insertion withinagrCfor one mutant, and we confirmed that this mutation abolishedagrfunction. These data provide the first substantial analysis ofwalKRpromoter function and show that prolonged vancomycin exposure can result in VISA through an IS256-mediated reduction inwalKRexpression; however, the mechanisms by which this occurs remain to be determined.

scholarly journals Role of BrnQ1 and BrnQ2 in Branched-Chain Amino Acid Transport and Virulence in Staphylococcus aureus

2014 ◽  
Vol 83 (3) ◽  
pp. 1019-1029 ◽  
Author(s):  
Julienne C. Kaiser ◽  
Sameha Omer ◽  
Jessica R. Sheldon ◽  
Ian Welch ◽  
David E. Heinrichs
Keyword(s):  
Staphylococcus Aureus ◽  
Virulence Gene ◽  
Defined Medium ◽  
Infection Model ◽  
Content Type ◽  
Virulence Gene Expression ◽  
Branched Chain Amino Acid ◽  
Branched Chain

The branched-chain amino acids (BCAAs; Ile, Leu, and Val) not only are important nutrients for the growth ofStaphylococcus aureusbut also are corepressors for CodY, which regulates virulence gene expression, implicating BCAAs as an important link between the metabolic state of the cell and virulence. BCAAs are either synthesized intracellularly or acquired from the environment.S. aureusencodes three putative BCAA transporters, designated BrnQ1, BrnQ2, and BrnQ3; their functions have not yet been formally tested. In this study, we mutated all threebrnQparalogs so as to characterize their substrate specificities and their roles in growthin vitroandin vivo. We demonstrated that in the community-associated, methicillin-resistantS. aureus(CA-MRSA) strain USA300, BrnQ1 is involved in uptake of all three BCAAs, BrnQ2 transports Ile, and BrnQ3 does not have a significant role in BCAA transport under the conditions tested. Of the three, only BrnQ1 is essential for USA300 to grow in a chemically defined medium that is limited for Leu or Val. Interestingly, we observed that abrnQ2mutant grew better than USA300 in media limited for Leu and Val, owing to the fact that this mutation leads to overexpression ofbrnQ1. In a murine infection model, thebrnQ1mutant was attenuated, but in contrast,brnQ2mutants had significantly increased virulence compared to that of USA300, a phenotype we suggest is at least partially linked to enhancedin vivoscavenging of Leu and Val through BrnQ1. These data uncover a hitherto-undiscovered connection between nutrient acquisition and virulence in CA-MRSA.

scholarly journals Antimicrobial Activity of Red Fruit (Pandanus conoideus Lam) Paste against Staphylococcus aureus and Escherichia coli in Burger Patties

2021 ◽  
Vol 16 (2) ◽  
pp. 132-143
Author(s):  
Selvia Tharukliling ◽  
Lilik Eka Radiati ◽  
Imam Thohari ◽  
Agus Susilo
Keyword(s):  
Fatty Acids ◽  
Staphylococcus Aureus ◽  
Antimicrobial Activity ◽  
Unsaturated Fatty Acids ◽  
Saturated Fatty Acids ◽  
Ethanol Extract ◽  
Plate Count ◽  
Significant Difference ◽  
The Mean ◽  
Paste Content

This study aims to determine the antimicrobial activity of red fruit paste against Staphylococcus aureus FNCC-0047 and Eschericia coli FNCC-0091 by using ethanol and n-hexane as well as the total plate count value of the patty with red fruit paste added on different observations day. The concentration of paste extract for antimicrobial activity test was 6.25%, 12.5%, 25%, 50%. The calculation of the total value of the burger patty plate is calculated on the 3rd, 7th and 14th d with the red fruit paste content in the formula as much as 0%, 5%, 10%, 15%. The results showed that the fatty acids of red fruit paste were dominated by oleic acid and linoleic acid, which are unsaturated fatty acids and palmitic acid, which are saturated fatty acids. The yield of red fruit paste in n-hexane solvent was higher than ethanol solvent. There was a significant difference (P <0.05) from the use of different solvents to the mean clear zone of the two bacteria tested. The antibacterial activity shown by the ethanol extract and n-hexane extract of red fruit paste was in the inactive category at the 6.12% level, the moderate category at the 12.5% to 25% level and the strong category at the 50% level. There was a significant difference (P <0.05) from the total microbial value in each treatment where the higher the red fruit paste content in the patty, the lower the total microbial value found on each d of observation. The use of red fruit paste at a level of 10% to 15% can withstand the rate of microbial growth

Enterotoxin B production by Staphylococcus aureus under controlled fatty acid nutrition induced by cerulenin

1977 ◽  
Vol 23 (9) ◽  
pp. 1145-1150 ◽  
Author(s):  
Robert A. Altenbern
Keyword(s):  
Fatty Acids ◽  
Staphylococcus Aureus ◽  
Fatty Acid ◽  
Membrane Fluidity ◽  
Unsaturated Fatty Acid ◽  
Unsaturated Fatty Acids ◽  
Basal Medium ◽  
Aureus Strain ◽  
Enterotoxin B ◽  
Fatty Acid Supplement

Cells of Staphylococcus aureus, strain S-6, can grow in the presence of 100 μg of cerulenin/ml if the basal medium is supplemented with certain saturated or unsaturated fatty acids. The production of enterotoxin B (SEB) is markedly influenced by both the ratio of saturated to unsaturated fatty acid and by the melting point of the unsaturated fatty acid supplement. The results presented suggest that a certain degree of membrane fluidity promotes maximum SEB production and that greater or lesser degrees of membrane fluidity prohibit substantial SEB formation but fail to affect final growth density.

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