Antibacterial Fatty Acids Research Articles

Identification of four genes responsible for antimicrobial resistance of MEL-B against S. aureus

There is increasing interest in the antimicrobial activity of mannosylerythritol lipids-B (MEL-B) against Gram-positive bacteria such as Staphylococcus aureus (S. aureus). However, the specific molecules involved in MEL-B's antimicrobial action against S. aureus have not been identified. This study utilized the Nebraska transposon mutant library (NTML), which contains 1920 mutants, each lacking three-quarters of the genes found in S. aureus. The NTML was screened to identify mutants resistant to MEL-B. Four mutants (Accession Number: SAUSA300_0904, SAUSA300_0752, SAUSA300_0387, and SAUSA300_2311) largely unaffected by incubation with MEL-B, indicating MEL-B resistance. Despite the strong binding of MEL-B to these mutants, the four molecules encoded by the deleted genes (yjbI, clpP, pbuX, or brpS) in each mutant were not directly recognized by MEL-B. Given that these molecules are not localized on the outer surface of S. aureus and that the antibacterial activity of MEL-B against S. aureus is facilitated by the effective transfer of two antibacterial fatty acids (caprylic acid and myristoleic acid) to S. aureus via ME, the deletion of each of the four molecules may alter the peptidoglycan structure, potentially inhibiting the effective transfer of these antimicrobial fatty acids into S. aureus.

Production of a selective antibacterial fatty acid against Staphylococcus aureus by Bifidobacterium strains.

Aims: C16 monounsaturated fatty acid (C16:1) show antibacterial activity against Staphylococcus aureus, a pathogen associated with various diseases such as atopic dermatitis and bacteremia, while the compound does not exhibit antibacterial activity against Staphylococcus epidermidis, an epidermal commensal that inhibits the growth of S. aureus. In this study, we aimed to find bifidobacterial strains with the ability to produce C16:1 and to find a practical manner to utilize C16:1-producing strains in industry. Methods: Various Bifidobacterium strains were screened for their content of C16:1. The chemical identity of C16:1 produced by a selected strain was analyzed by gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS). Medium components that affect the C16:1 content of the selected strain were investigated. Antibacterial activity against staphylococci was compared between the authentic C16:1 isomers and total fatty acids (TFA) extracted from the selected strain. Results: B. adolescentis 12451, B. adolescentis 12-111, B. boum JCM 1211, and Bifidobacterium sp. JCM 7042 showed high C16:1 content among the tested strains. TFA extracted from Bifidobacterium sp. JCM 7042 contained C16:1 at 2.3% as the fatty acid constituent (2.4 mg/L of broth). Through GC-MS and LC-MS analyses, the C16:1 synthesized by Bifidobacterium sp. JCM 7042 was identified as 7-cis-hexadecenoic acid (7-cis-C16:1). The authentic 7-cis-C16:1 showed strong and selective antibacterial activity against S. aureus, similar to 6-cis-C16:1, with a minimum inhibitory concentration (MIC) of < 10 µg/mL. Components that increase C16:1 productivity were not found in the MRS and TOS media; however, Tween 80 was shown to considerably reduce the C16:1 ratio in TFA. Antibacterial activity against S. aureus was observed when the TFA extracted from Bifidobacterium sp. JCM 7042 contained high level of 7-cis-C16:1 (6.1% in TFA) but not when it contained low level of 7-cis-C16:1 (0.1% in TFA). Conclusion: The fatty acid, 7-cis-C16:1, which can selectively inhibit the S. aureus growth, is accumulated in TFA of several bifidobacteria. The TFA extracted from cultured cells of Bifidobacterium sp. JCM 7042 demonstrated antibacterial activity. From a practical viewpoint, our findings are important for developing an efficient method to produce novel skin care cosmetics, functional dairy foods, and other commodities.

A genomic island in a plant beneficial rhizobacterium encodes novel antimicrobial fatty acids and a self-protection shield to enhance its competition.

Rhizobacteria devote a relatively large percentage of their genomes to encode bioactive natural products that are important for competition in the rhizosphere. In this study, a plant beneficial rhizobacterium Bacillus velezensis SQR9 was discovered to produce novel antibacterial fatty acids, Bacillunoic acids, which are encoded on a genomic island (GI). This GI contains a hybrid type I fatty acid synthase (FAS)-polyketide synthase (PKS) system and an ABC transporter. The FAS was predicted to synthesize a primer that was transferred to the PKS to synthesize Bacillunoic acids. The synthesized Bacillunoic acids inhibit the growth of diverse bacteria, with the strongest activity against closely related Bacillus strains, the ABC transporter exported the toxic Bacillunoic acids upon their induction for protecting the producing strain. The inhibition of other Bacillus strains by Bacillunoic acids extended the antimicrobial spectrum of SQR9 and enhanced its competition with closely related root-associated bacteria. So, through the obtaining of this GI by horizontal gene transfer, strain SQR9 not only acquired a competitive weapon but also acquired a self-protecting shield, which increased its competition with other rhizobacteria.

Production of hydrocarbon-degrading microorganisms using agricultural residues of Mangifera indica L. and Carica papaya as carbon source

The aim of the present study was to evaluate the potential of oils from agricultural residues, such as Mangifera indica L. (mango) and Carica papaya (papaya) from the Papaloapan region, Mexico, as a carbon source for the production of hydrocarbon-degrading (hydrocarbonoclastic) microorganisms in an airlift bioreactor via a common metabolic pathway for hydrocarbons and fatty acids. Biomass growth and carbon source uptake were measured using optical density and gas chromatography, respectively. Gompertz, logistic, and Von Bertalanffy mathematical models were used to obtain kinetic parameters such as the lag phase, maximum specific growth, and consumption rate. The hydrocarbonoclastic consortium was able to grow using papaya (6.09 ± 0.23gL-1) and mango (2.59 ± 0.30gL-1) oils, which contain certain antibacterial fatty acids. Differences observed in maximum specific growth and consumption rates indicate that, although mango oil was consumed faster (0.33day-1 for mango and 0.25day-1 for papaya), papaya oil provided a higher rate of biomass production per microorganism (0.24day-1 for mango and 0.44day-1 for papaya). Additionally, the consortium was able to consume 13gL-1 diesel as a sole carbon source and improve its maximum specific consumption rate following growth using the oils. Furthermore, the maximum specific growth rate was decreased, indicating a change in the consortium capabilities. Nevertheless, agricultural waste oils from the Papaloapan region can be used to cultivate hydrocarbonoclastic microorganisms. The present study creates the possibility of investigating carbon sources other than hydrocarbons for the production of hydrocarbonoclastic microorganisms.

Antibacterial fatty acids destabilize hydrophobic and multicellular aggregates of biofilm in S. aureus.

Present study is based on 20 methicillin-resistant Staphylococcus aureus (MRSA) isolates recovered from different food items. These isolates were identified on the basis of colony morphology, Gram staining and growth on different selective and differential media. Studies on 16S RNA and positive reactions on DNase agar and Prolex Latex Agglutination system confirm it as Staphylococcus aureus. Oxacillin susceptibility testing and PCR with mecA gene-specific primer results showed that these isolates are MRSA-carrying mecA gene that belongs to SCCmecA type IV and also harbor agr type II. Phenotypic study revealed that these isolates adopt biofilm mode of growth after exposure to subinhibitory doses of oxacillin. The biofilm and cell surface hydrophobicity have a strong correlation. It was noticed that affinity to hexadecane (apolar-solvent) of planktonic cells was low, suggesting its hydrophilic character. However, as the cells are exposed to oxacillin, they adopt biofilm mode of life and the affinity to apolar solvent increases, indicating a hydrophobic character. In biofilm consortia, the cells with more hydrophobic surfaces show incomplete septation and produce multicellular aggregates. This is due to reduced expression of atl gene. This was confirmed by real-time PCR studies. Moreover, the planktonic or wild-type phenotypes of these isolates were more tolerant to antibacterial effect of the fatty acids used; that is, cis-2-decanoic acid and cis-9-octadectanoic acid. These fatty acids were more effective against biofilms. After exposure to these fatty acids, established biofilms were dispersed and surviving cells were unable to readopt biofilm mode of life. The planktonic or wild-type phenotypes produce fatty acid-modifying enzyme (FAME) to inactivate the bactericidal activity of fatty acids by esterification to cholesterol. The biofilm indwellers are metabolically inactive and unable to produce FAME; hence, they are vulnerable to antibiofilm effect of cis-2-decanoic acid and cis-9-octadectanoic acid.

Spectrum of Membrane Morphological Responses to Antibacterial Fatty Acids and Related Surfactants.

Medium-chain saturated fatty acids and related compounds (e.g., monoglycerides) represent one class of membrane-active surfactants with antimicrobial properties. Most related studies have been in vitro evaluations of bacterial growth inhibition, and there is limited knowledge about how the compounds in this class destabilize lipid bilayers, which are the purported target within the bacterial cell membrane. Herein, the interaction between three representative compounds in this class and a supported lipid bilayer platform was investigated using quartz crystal microbalance-dissipation and fluorescence microscopy in order to examine membrane destabilization. The three tested compounds were lauric acid, sodium dodecyl sulfate, and glycerol monolaurate. For each compound, we discovered striking differences in the resulting morphological changes of supported lipid bilayers. The experimental trends indicate that the compounds have membrane-disruptive behavior against supported lipid bilayers principally above the respective critical micelle concentration values. The growth inhibition properties of the compounds against standard and methicillin-resistant Staphylococcus aureus bacterial strains were also tested. Taken together, the findings in this work improve our knowledge about how saturated fatty acids and related compounds destabilize lipid bilayers, offering insight into the corresponding molecular mechanisms that lead to membrane morphological responses.

Comparative assessment of microalgal fatty acids as topical antibiotics

Bacteria are increasingly resistant to common antibiotics. Certain fatty acids are effective antibacterial agents and, when compared to conventional antibiotics, have fewer incidences of bacterial resistance. Algae naturally synthesize these fatty acids and can provide a sustainable source of these antibiotics. To evaluate the antibacterial properties of fatty acids, 29 acids present in algae were tested for topical antibacterial properties on Escherichia coli and Staphylococcus aureus using surface zone inhibition assay. From the set of fatty acids showing good antibacterial activity, five representative fatty acids—capric acid (10:0), palmitoleic acid (16:1), gamma-linolenic acid [18:3 (n-6)], arachidonic acid [20:4 (n-6)], and docosadienoic acid [22:2 (n-6)]—were selected for further analysis. Algal growth conditions (light, temperature, and nutrients) were manipulated to increase the content of the selected target fatty acids in five algal species (Rhodella maculata, Phaeodactylum tricornutum, Boekelovia hooglandii, Goniochloris sculpta, and Chloridella simplex). P. tricornutum contained the highest content of the target antibacterial fatty acids, with a maximum of 206 mg g−1 dry weight (dw). However, biomass productivity was low; thus, for commercial viability, initial growth should be under optimum conditions and then shifted to enhance antibacterial fatty acid content. B. hooglandii had the highest biomass productivity and a total antibacterial fatty acid content of 32.6 mg g−1 dw in Erddekokt + Salze + Peptone (ESP) medium, 18 °C, 80 μmol photons m−2 s−1, and 17:7-h light/dark cycle. This study identifies fatty acids that have effective antibacterial properties and potential use as topical antibacterial products and determined growth conditions that result in algae rich in these acids.

Inhibition of pathogenic Vibrio by the microalgae Isochrysis galbana

Vibrio alginolyticus, Vibrio campbellii, and Vibrio harveyi were inhibited by Isochrysis galbana in batch cultures. I. galbana reduced the V. alginolyticus, V. campbellii, and V. harveyi counts to undetectable levels in 2, 4, and 7 days ( 82 %) of I. galbana during the stationary growth phase were estearidonic (24.3 %), oleic (15.7 %), myristic (13.8 %), docosahexaenoic (11.0 %), palmitic (10.3 %), and α-linolenic (7.2 %) acids. These results demonstrate that I. galbana synthesizes antibacterial fatty acids that inhibit the growth of pathogenic bacteria such as V. alginolyticus, V. campbellii, and V. harveyi.

Native Efflux Pumps Contribute Resistance to Antimicrobials of Skin and the Ability of Staphylococcus aureus to Colonize Skin

Staphylococcus aureus colonizes skin in the presence of antimicrobial fatty acids and polyamines. The chromosomally encoded Tet38 efflux transporter confers resistance to tetracycline and fitness in abscesses, but its natural substrates and those of the Nor quinolone efflux pumps are unknown. Susceptibility of tet38 and other pump mutants to and pump gene induction by fatty acids and polyamines were compared. Transport of fatty acids by Tet38 was determined in membrane vesicles. Survival on skin was tested in an adapted mouse skin infection model. The tet38 expression caused a 5- to 8-fold increase in resistance to palmitoleic and undecanoic acids but not polyamines. Subinhibitory concentrations of these fatty acids induced 4-fold increases in tet38 transcripts and competitively inhibited transport of Hoechst 33 342 dye in Tet38 membrane vesicles. Colonization of skin in BALB/c mice was decreased 5-fold in a Δtet38 mutant, which was complemented by plasmid-encoded tet38. Although polyamine minimum inhibitory concentrations (MICs) decreased 4-fold in a norC::cat mutant and increased 8-fold with norC overexpression, spermidine did not induce expression of norC and other pump genes, and norC::cat exhibited wild-type colonization. Antibacterial fatty acids may be natural substrates of Tet38, which contributes to resistance and the ability of S. aureus to colonize skin.

INFLUENCE OF ETHYLENEDIAMINE‐N,N′‐DISUCCINIC ACID (EDDS) CONCENTRATION ON THE BACTERICIDAL ACTIVITY OF FATTY ACIDS IN VITRO*

ABSTRACTThe antibacterial activity of mixtures of ethylenediamine‐N,N′‐disuccinic acid (EDDS) and antibacterial fatty acids (FA) was examined using the agar diffusion assay. Solutions of caproic, caprylic, capric and lauric acids dissolved in potassium hydroxide (KOH) were supplemented with 0, 5 or 10 mM of EDDS and adjusted to pH 11.0 with citric acid. Wells made in agar media seeded with one of nine bacterial isolates were filled with FA‐KOH or FA‐KOH‐EDDS solutions. After incubation of agar plates, zones of inhibition of bacterial growth around the agar wells were measured. Results indicated that caproic‐KOH‐EDDS and caprylic‐KOH‐EDDS inhibited growth of more bacterial isolates than caproic‐KOH and caprylic‐KOH, respectively. Although, capric‐KOH and lauric‐KOH produced zones of inhibition of all nine bacterial isolates, significantly (P ≤ 0.05) larger zones of inhibition were produced by capric‐KOH‐EDDS and lauric‐KOH‐EDDS. Findings of this study indicate that mixtures of FA‐KOH‐EDDS possess greater antibacterial activity than FA‐KOH.PRACTICAL APPLICATIONSResearch demonstrates that supplementing mixtures of FA‐KOH with EDDS increases the antibacterial activity of these antimicrobial surfactants. Similar results have been produced with FA‐KOH supplemented with ethylenediaminetetraacetic acid (EDTA). However, EDTA is a man‐made, nonbiodegradable chelator, whereas EDDS is a natural biodegradable chelator. Therefore, EDDS may be used as an alternative for EDTA in formulations of FA‐KOH currently being examined as novel sanitizers for decreasing microbial contamination associated with poultry processing. Since EDDS is biodegradable, use of this chelator will not cause concerns of environmental pollution associated with the use of EDTA.

A Fatty Acid from the Diatom Phaeodactylum tricornutum is Antibacterial Against Diverse Bacteria Including Multi-resistant Staphylococcus aureus (MRSA)

Pathogenic bacteria, such as multidrug-resistant Staphylococcus aureus (MRSA), which are not susceptible to most conventional antibiotics, are causing increased concern in healthcare institutions worldwide. The discovery of novel antibacterial compounds for biomedical exploitation is one avenue that is being pursued to combat these problematic bacteria. Marine eukaryotic microalgae are known to produce numerous useful products but have attracted little attention in the search for novel antibiotic compounds. Cell lysates of the marine diatom, Phaeodactylum tricornutum Bohlin, have been reported to display antibacterial activity in vitro, but the compounds responsible have not been fully identified. In this paper, using column chromatography and reversed-phase high-performance liquid chromatography, we report the isolation of an antibacterial fatty acid. Mass spectrometry and (1)H-nuclear magnetic resonance spectroscopy revealed it to be the polyunsaturated fatty acid, eicosapentaenoic acid (EPA). We show that EPA is active against a range of both Gram-positive and Gram-negative bacteria, including MRSA, at micromolar concentrations. These data indicate that it could find application in the topical and systemic treatment of drug-resistant bacterial infections.

Isolation of antibacterial fatty acids from Schotia brachypetala

In southern Africa, the roots of Schotia brachypetala are used by traditional healers to treat dysentery and diarrhoea. Activity-directed fractionation of the ethanol extract of the dried leaves yielded 9,12,15-octadecatrienoic (linolenic) acid and methyl-5,11,14,17-eicosatetraenoate. These fatty acids displayed antibacterial activity against the Gram-positive bacteria Bacillus subtilis and Staphylococcus aureus and to a lesser extent, against the Gram-negative Escherichia coli and Klebsiella pneumoniae.

Fatty acids in honeybee-collected pollens from six endemic Western Australian eucalypts and the possible significance to the Western Australian beekeeping industry

Western Australian eucalypt pollens in this research have low levels of lipid (0.59–1.9%) when compared with many other plant species that have evolved alongside the European honeybee. Eucalypt-pollen lipid was dominant in linoleic acid (35.7–48%). The six other major fatty acids that were present in the lipid were myristic, palmitic, stearic, oleic, linolenic and arachidic acids. Linoleic acid was dominant in eucalypt pollen at average concentrations of 2.77–5.81 mg/g pollen. These results could be of significance to the Australian beekeeping industry in refining disease-management strategies in the light that other researchers have found that 2 economically damaging bee diseases (EFB and AFB) are inhibited by certain concentrations of the acid. Redgum- or marri (Corymbia calophylla)-pollen lipid was also dominated by 2 other known antibacterial fatty acids: myristic (0.25 mg/g pollen) and linolenic (1.06 mg/g pollen), when compared with the other eucalypts studied.

The farAB-encoded efflux pump mediates resistance of gonococci to long-chained antibacterial fatty acids.

Gonococci often infect mucosal surfaces bathed in antibacterial fatty acids (FAs). Resistance of gonococci to FAs and other antibacterial hydrophobic agents has been attributed to the mtrCDE-encoded efflux pump system and a heretofore undefined mechanism. This alternative resistance mechanism has been suggested to mediate gonococcal resistance to long-chained FAs independently of the mtr efflux pump. We have now identified this alternative FA resistance system in gonococci and report that it bears significant similarity to the emrAB-encoded efflux pump possessed by Escherichia coli and the vceAB-encoded pump of Vibrio cholerae. We termed the gonococcal version of this efflux pump farAB (fatty acid resistance) to signify its involvement in FA resistance expressed by gonococci and to distinguish it from the emrAB- or vceAB-encoded pumps that modulate bacterial susceptibility to uncoupling agents and certain antibiotics. Although the farAB system in gonococci was found to provide resistance to FAs independently of the mtrCDE-encoded efflux pump, its function was dependent on the MtrE outer membrane protein. Moreover, expression of the tandemly linked farA and farB genes was positively associated with the presence of the MtrR transcriptional regulatory protein that normally downregulates the expression of mtrCDE. Thus, the data presented herein suggest that, while the mtrCDE- and farAB-encoded systems act independently to mediate resistance of gonococci to host-derived, hydrophobic antimicrobial agents, their capacity to export these agents is dependent on the same outer membrane protein (MtrE), and their expression may be differentially controlled by the same transcriptional regulatory protein (MtrR).

Antibacterial constituents of the red alga cystoclonium purpureum

The red alga Cystoclonium purpreum was found to contain α-carotene, plastoquinone-9, trans-phytol, ubiquinol-9, lutein and fucoxanthin. The composition of the antibacterial fatty acid fraction was determined by GC/MS.