Synergistic Antibiofilm Efficacy of Thymol and Piperine in Combination with Three Aminoglycoside Antibiotics against Klebsiella pneumoniae Biofilms

Background. Thymol and piperine are two naturally occurring bioactive compounds with several pharmacological activities. In this study, their antibiofilm potential either alone or in combination with three aminoglycoside antibiotics was evaluated against a biofilm of Klebsiella pneumoniae. Methods. Determination of antimicrobial susceptibility was performed using the broth microdilution method. Biofilm formation was evaluated by the microtiter plate method. Antibiofilm activity was determined using 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium-bromide (MTT) assay. The combination studies were performed by the checkerboard microdilution method. Results. The minimum biofilm inhibitory concentration (MBIC) of streptomycin was reduced by 16- to 64-fold when used in combination with thymol, while the MBIC of kanamycin was reduced by 4-fold when combined with piperine. The minimum biofilm eradication concentration (MBEC) values of streptomycin, amikacin, and kanamycin were, respectively, 16- to 128-fold, 4- to 128-fold, and 8- to 256-fold higher than the planktonic minimum inhibitory concentration (MIC). Thymol combined with streptomycin or kanamycin showed synergic effects against the preformed biofilm with 16- to 64-fold reduction in the minimum biofilm eradication concentration values of each antibiotic in combination. Piperine acted also synergically with kanamycin with an 8- to 16-fold reduction in the minimum biofilm eradication concentration values of kanamycin in combination. Conclusion. The association of thymol with antibiotics showed a strong synergistic effect both in the inhibition of biofilm formation and the destruction of the preformed biofilm of K. pneumoniae. This study suggests that a combination of thymol with streptomycin, amikacin, or kanamycin could be a promising alternative therapy to overcome the problem of K. pneumoniae biofilm-associated infections.

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8-hydroxyquinoline-5-(N-4-chlorophenyl) sulfonamide and fluconazole combination as a preventive strategy for Candida biofilm in haemodialysis devices

Journal of Medical Microbiology ◽

10.1099/jmm.0.001377 ◽

2021 ◽

Vol 70 (7) ◽

Author(s):

Letícia Fernandes da Rocha ◽

Bruna Pippi ◽

Angélica Rocha Joaquim ◽

Saulo Fernandes de Andrade ◽

Alexandre Meneghello Fuentefria

Keyword(s):

Biofilm Formation ◽

Synergistic Effects ◽

Local Treatment ◽

The Other ◽

Antibiofilm Activity ◽

Preventive Strategy ◽

Clinical Issue ◽

Microdilution Method ◽

Broth Microdilution Method ◽

Time Kill

Introduction. The presence of Candida biofilms in medical devices is a concerning and important clinical issue for haemodialysis patients who require constant use of prosthetic fistulae and catheters. Hypothesis/Gap Statement. This prolonged use increases the risk of candidaemia due to biofilm formation. PH151 and clioquinol are 8-hydroxyquinoline derivatives that have been studied by our group and showed interesting anti-Candida activity. Aim. This study evaluated the biofilm formation capacity of Candida species on polytetrafluoroethylene (PTFE) and polyurethane (PUR) and investigated the synergistic effects between the compounds PH151 and clioquinol and fluconazole, amphotericin B and caspofungin against biofilm cells removed from those materials. Further, the synergistic combination was evaluated in terms of preventing biofilm formation on PTFE and PUR discs. Methodology. Susceptibility testing was performed for planktonic and biofilm cells using the broth microdilution method. The checkerboard method and the time–kill assay were used to evaluate the interactions between antifungal agents. Antibiofilm activity on PTFE and PUR materials was assessed to quantify the prevention of biofilm formation. Results. Candida albicans, Candida glabrata and Candida tropicalis showed ability to form biofilms on both materials. By contrast, Candida parapsilosis did not demonstrate this ability. Synergistic interaction was observed when PH151 was combined with fluconazole in 77.8 % of isolates and this treatment was shown to be concentration- and time-dependent. On the other hand, indifferent interactions were predominantly observed with the other combinations. A reduction in biofilm formation on PUR material of more than 50 % was observed when using PH151 combined with fluconazole. Conclusion. PH151 demonstrated potential as a local treatment for use in a combination therapy approach against Candida biofilm formation on haemodialysis devices.

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Influence of Gallic Acid and Thai Culinary Essential Oils on Antibacterial Activity of Nisin against Streptococcus mutans

Advances in Pharmacological and Pharmaceutical Sciences ◽

10.1155/2021/5539459 ◽

2021 ◽

Vol 2021 ◽

pp. 1-12

Author(s):

Pimsumon Jiamboonsri ◽

Pimpikar Kanchanadumkerng

Keyword(s):

Essential Oils ◽

Gallic Acid ◽

Streptococcus Mutans ◽

Biofilm Formation ◽

Inhibitory Concentration ◽

Microdilution Method ◽

Broth Microdilution Method ◽

Life Threatening ◽

Oral Pathogens ◽

Time Kill

Streptococcus mutans is a well-known oral pathogen commonly associated with a normal dental problem and life-threatening infection. A bacteriocin nisin and the plant-derived compounds including gallic acid (GA) and Thai culinary essential oils (EOs) have been reported to have activity against oral pathogens. However, their synergistic interaction against S. mutans has not been explored. The purposes of this study were primarily to investigate anti-S. mutans properties and the antibiofilm formation of nisin, GA, and five EOs by using the broth microdilution method. Besides, the morphological change, killing rate, and antibacterial synergism were determined by scanning electron microscopy (SEM), time-kill assay, and checkerboard method, respectively. The results demonstrated that kaffir lime leaf (KLL) oil, lemongrass (LG) oil, and GA showed a potent anti-S. mutans activity and inhibited biofilm formation with the possible mechanism targeted on the cell membrane. Additionally, KLL oil revealed anti-S. mutans synergism with GA, LG oil, and chlorhexidine with the fractional inhibitory concentration (FIC) indexes ≤ 0.5. Interestingly, GA displayed a high potential to enhance anti-S. mutans activity of nisin by lowering the minimum inhibitory concentrations (MICs) to at least 8-fold in a bacteriostatic manner. These results suggest that GA and KLL oil may be potentially used as an adjunctive therapy along with nisin and chlorhexidine to control S. mutans infection.

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Impact of Tigecycline’s MIC in the Outcome of Critically Ill Patients with Carbapenemase-Producing Klebsiella pneumoniae Bacteraemia Treated with Tigecycline Monotherapy—Validation of 2019′s EUCAST Proposed Breakpoint Changes

Antibiotics ◽

10.3390/antibiotics9110828 ◽

2020 ◽

Vol 9 (11) ◽

pp. 828

Author(s):

Matthaios Papadimitriou-Olivgeris ◽

Christina Bartzavali ◽

Alexandra Nikolopoulou ◽

Fevronia Kolonitsiou ◽

Virginia Mplani ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Primary Outcome ◽

Inhibitory Concentration ◽

Therapeutic Option ◽

Bloodstream Infections ◽

Microdilution Method ◽

Appropriate Treatment ◽

Broth Microdilution Method ◽

The Impact ◽

Treatment Mortality

Background: Tigecycline is a therapeutic option for carbapenemase-producing Klebsiella pneumoniae (CP-Kp). Our aim was to evaluate the impact of the tigecycline’s minimum inhibitory concentration (MIC) in the outcome of patients with CP-Kp bacteraemia treated with tigecycline monotherapy. Methods: Patients with monomicrobial bacteraemia due to CP-Kp that received appropriate targeted monotherapy or no appropriate treatment were included. Primary outcome was 30-day mortality. MICs of meropenem, tigecycline, and ceftazidime/avibactam were determined by Etest, whereas for colistin, the broth microdilution method was applied. PCR for blaKPC, blaVIM, blaNDM, and blaOXA genes was applied. Results: Among 302 CP-Kp bacteraemias, 32 isolates (10.6%) showed MICs of tigecycline ≤ 0.5 mg/L, whereas 177 (58.6%) showed MICs that were 0.75–2 mg/L. Colistin and aminoglycoside susceptibility was observed in 43.0% and 23.8% of isolates, respectively. The majority of isolates carried blaKPC (249; 82.5%), followed by blaVIM (26; 8.6%), both blaKPC and blaVIM (16; 5.3%), and blaNDM (11; 3.6%). Fifteen patients with tigecycline MIC ≤ 0.5 mg/L and 55 with MIC 0.75–2 mg/L were treated with tigecycline monotherapy; 30-day mortality was 20.0% and 50.9%, respectively (p = 0.042). Mortality of 150 patients that received other antimicrobials was 24.7%; among 82 patients that received no appropriate treatment, mortality was 39.0%. No difference in 30-day mortality was observed between patients that received tigecycline (MIC ≤ 0.5 mg/L) or other antimicrobials. Conclusion: Tigecycline monotherapy was as efficacious as other antimicrobials in the treatment of bloodstream infections due to CP-Kp isolates with a tigecycline’s MIC ≤ 0.5 mg/L.

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In vitro antibiofilm activity of resveratrol against avian pathogenic Escherichia coli

BMC Veterinary Research ◽

10.1186/s12917-021-02961-3 ◽

2021 ◽

Vol 17 (1) ◽

Author(s):

Xiangchun Ruan ◽

Xiaoling Deng ◽

Meiling Tan ◽

Chengbo Yu ◽

Meishi Zhang ◽

...

Keyword(s):

Escherichia Coli ◽

Biofilm Formation ◽

Inhibitory Concentration ◽

Laser Scanning Microscopy ◽

Antibiofilm Activity ◽

Dependent Manner ◽

Swarming Motility ◽

Semisolid Medium ◽

Broth Microdilution Method

Abstract Background Avian pathogenic Escherichia coli (APEC) strains cause infectious diseases in poultry. Resveratrol is extracted from Polygonum cuspidatum, Cassia tora Linn and Vitis vinifera, and displays good antimicrobial activity. The present study aimed to investigate the antibiofilm effect of resveratrol on APEC in vitro. The minimum inhibitory concentration (MIC) of resveratrol and the antibiotic florfenicol toward APEC were detected using the broth microdilution method. Then, the effect of resveratrol on swimming and swarming motility was investigated using a semisolid medium culture method. Subsequently, the minimum biofilm inhibitory concentration (MBIC) and the biofilm eradication rate were evaluated using crystal violet staining. Finally, the antibiofilm activity of resveratrol was observed using scanning electron microscopy (SEM). Meanwhile, the effects of florfenicol combined with resveratrol against biofilm formation by APEC were evaluated using optical microscopy (OM) and a confocal laser scanning microscopy (CLSM). Results The MICs of resveratrol and florfenicol toward APEC were 128 μg/mL and 64 μg/mL, respectively. The swimming and swarming motility abilities of APEC were inhibited in a resveratrol dose-dependent manner. Furthermore, resveratrol showed a significant inhibitory activity against APEC biofilm formation at concentrations above 1 μg/mL (p < 0.01). Meanwhile, the inhibitory effect of resveratrol at 32 μg/mL on biofilm formation was observed using SEM. The APEC biofilm was eradicated at 32 μg/mL of resveratrol combined with 64 μg/mL of florfenicol, which was observed using CLSM and OM. Florfenicol had a slight eradication effect of biofilm formation, whereas resveratrol had a strong biofilm eradication effect toward APEC. Conclusion Resveratrol displayed good antibiofilm activity against APEC in vitro, including inhibition of swimming and swarming motility, biofilm formation, and could eradicate the biofilm.

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Synergistic Antibiofilm Effect of Thymol and Piperine in Combination with Aminoglycosides Antibiotics against Four Salmonella enterica Serovars

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2021/1567017 ◽

2021 ◽

Vol 2021 ◽

pp. 1-9

Author(s):

Christian Ramsès Tokam Kuaté ◽

Borel Bisso Ndezo ◽

Jean Paul Dzoyem

Keyword(s):

Natural Products ◽

Synergistic Effect ◽

Salmonella Enterica ◽

Biofilm Formation ◽

Antimicrobial Agents ◽

Inhibitory Concentration ◽

Assay Method ◽

Antibiofilm Activity ◽

Microtiter Plate Assay ◽

Broth Microdilution Method

Biofilms related to human infection have high levels of pathogenicity due to their resistance to antimicrobial agents. The discovery of antibiofilm agents is necessary. One approach to overcome this problem is the use of antibiotics agents’ combination. This study aimed to determine the efficacy of the combination of natural products thymol and piperine with three aminoglycosides antibiotics, amikacin, kanamycin, and streptomycin against biofilm-forming Salmonella enterica. The microtiter plate assay method was used to evaluate the biofilm-producing capacity of the isolates. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration were determined by the broth microdilution method. The inhibition of biofilm formation and biofilm eradication was determined using the microtiter broth method. The checkerboard method was used to determine the combined effects of natural products with aminoglycosides antibiotics. All the tested isolates showed various levels of biofilm formation. Overall, combinations provided 43.3% of synergy in preventing the biofilm formation and 40% of synergy in eradicating preformed biofilms, and in both cases, no antagonism was observed. The combination of thymol with kanamycin showed a synergistic effect with 16- to 32-fold decrease of the minimum biofilm eradication concentration (MBEC) of kanamycin. The interaction of piperine with amikacin and streptomycin also revealed a synergistic effect with 16-fold reduction of the minimum biofilm inhibitory concentration (MBIC). The combination of thymol with the three antibiotics showed a strong synergistic effect in both inhibiting the biofilm formation and eradicating the preformed biofilm. This study demonstrates that thymol and piperine potentiate the antibiofilm activity of amikacin, kanamycin, and streptomycin. These combinations are a promising approach therapeutic to overcome the problem of Salmonella enterica biofilm-associated infections. In addition, these combinations could help reduce the concentration of individual components, thereby minimizing the nephrotoxicity of aminoglycosides antibiotics.

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Antibacterial and Antibiofilm Activity of Daemonorops draco Resin

Biosaintifika Journal of Biology & Biology Education ◽

10.15294/biosaintifika.v10i1.13554 ◽

2018 ◽

Vol 10 (1) ◽

pp. 138-144 ◽

Cited By ~ 2

Author(s):

Wulan Tri Wahyuni ◽

Sri Purwanti ◽

Irmanida Batubara

Keyword(s):

Antimicrobial Activity ◽

Antibacterial Activity ◽

Ethyl Acetate ◽

Biofilm Formation ◽

Inhibitory Concentration ◽

Extraction Yield ◽

Diffusion Method ◽

Antibiofilm Activity ◽

Microdilution Method ◽

Violet Crystal

Daemonorops draco has been reported for its antibacterial activity and empirically used for wound healing by Anak Dalam ethnic at Jambi Province, Sumatera-Indonesia. This study was performed to evaluate antibacterial and antibiofilm activity of D. draco resin collected from Jambi. D. draco resin was extracted using n-hexane, ethyl acetate and methanol, respectively. Antibacterial activity of the extracts was evaluated using agar diffusion method against Staphylococcus aureus and Eschericia coli, whereas the minimum inhibitory concentration (MIC) and minimum bacteriacidal concentration (MBC) was determined by microdilution method. In addition, antibiofilm activity was evaluated by violet crystal method. The result showed that extraction yield of ethyl acetate was higher than methanol and n-hexane. Ethyl acetate and methanol extracts of D. draco exhibited stronger antimicrobial activity against S. aureus compare to n-hexane extract. MIC and MBC of methanol extract and chromatographic fraction (F5.1) of ethyl acetate extract were 0.5 and 1.0 mg/mL, respectively. In addition, antibiofilm assay revealed that all extracts were inhibit initial attachment of bacteria cell in biofilm formation. This result revealed a novel information that D. draco extracts was potential as inhibitor of biofilm formation. TLC bioautography of D. draco extracts indicated that constituent with Rf of 0.71 performed antimicrobial activity against S. aureus. This finding expected to strengthen the scientific backup for utilization of D. draco by society.

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Whole Transcriptomic Analysis of Mechanisms of Tolerance and Resistance to Chlorhexidine in Clinical Strains of Klebsiella Pneumoniae Producers of Carbapenemase

10.20944/preprints202007.0579.v1 ◽

2020 ◽

Author(s):

Ines Bleriot ◽

Lucia Blasco ◽

Mercedes Delgado ◽

Ana Gual de Torrella ◽

Anton Ambroa ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Biofilm Formation ◽

Molecular Mechanisms ◽

Cross Resistance ◽

Beta Lactamase ◽

Tolerance Mechanisms ◽

Clinical Strains ◽

Microdilution Method ◽

Broth Microdilution Method ◽

High Level

Although the failure of antibiotic treatment is normally attributed to resistance, tolerance and persistence display a significant role in the lack of response to antibiotics. Due to the fact that several nosocomial pathogens show a high level of tolerance and/or resistance to chlorhexidine, in this study we analyzed the molecular mechanisms associated with chlorhexidine adaptation in two clinical strains of Klebsiella pneumoniae by phenotypic and transcriptomic studies. These two strains belong to ST258-KPC3 (high-risk clone carrying β-lactamase KPC3) and ST846-OXA48 (low-risk clone carrying β-lactamase OXA48). Our results showed that K. pneumoniae ST258-KPC3CA and ST846-OXA48CA strains exhibited a different behavior under CHLX pressure, adapting to this biocide through resistance and tolerance mechanisms, respectively. Furthermore, the appearance of cross-resistance to colistin was observed in the ST846-OXA48CA strain (tolerant to CHLX), using the broth microdilution method. Interestingly, this ST846-OXA48CA isolate contained a plasmid that encodes a novel type II toxin/antitoxin (TA) system, PemK/PemI. We characterized this PemK/PemI TA system by cloning both genes into the IPTG-inducible pCA24N plasmid, and found their role in persistence and biofilm formation. Accordingly, the ST846-OXA48CA strain showed a persistence biphasic curve in the presence of a chlorhexidine-imipenem combination, and these results were confirmed by the enzymatic assay (WST-1).

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Mortality of Pandrug-Resistant Klebsiella pneumoniae Bloodstream Infections in Critically Ill Patients: A Retrospective Cohort of 115 Episodes

Antibiotics ◽

10.3390/antibiotics10010076 ◽

2021 ◽

Vol 10 (1) ◽

pp. 76

Author(s):

Matthaios Papadimitriou-Olivgeris ◽

Christina Bartzavali ◽

Alexandra Georgakopoulou ◽

Fevronia Kolonitsiou ◽

Chrisavgi Papamichail ◽

...

Keyword(s):

Septic Shock ◽

Klebsiella Pneumoniae ◽

Critically Ill ◽

Critically Ill Patients ◽

Primary Outcome ◽

Bloodstream Infections ◽

Microdilution Method ◽

Broth Microdilution Method ◽

Carbapenemase Gene ◽

Comorbidity Index

Background: The increased frequency of bacteraemias caused by pandrug-resistant Klebsiella pneumoniae (PDR-Kp) has significant implications. The aim of the present study was to identify predictors associated with mortality of PDR-Kp bacteraemias. Methods: Patients with monomicrobial bacteraemia due to PDR-Kp were included. K. pneumoniae was considered PDR if it showed resistance to all available groups of antibiotics. Primary outcome was 30-day mortality. Minimum inhibitory concentrations (MICs) of meropenem, tigecycline, fosfomycin, and ceftazidime/avibactam were determined by Etest, whereas for colistin, the broth microdilution method was applied. blaKPC, blaVIM, blaNDM, and blaOXA genes were detected by PCR. Results: Among 115 PDR-Kp bacteraemias, the majority of infections were primary bacteraemias (53; 46.1%), followed by catheter-related (35; 30.4%). All isolates were resistant to tested antimicrobials. blaKPC was the most prevalent carbapenemase gene (98 isolates; 85.2%). Thirty-day mortality was 39.1%; among 51 patients with septic shock, 30-day mortality was 54.9%. Multivariate analysis identified the development of septic shock, Charlson comorbidity index, and bacteraemia other than primary or catheter-related as independent predictors of mortality, while a combination of at least three antimicrobials was identified as an independent predictor of survival. Conclusions: Mortality of PDR-Kp bloodstream infections was high. Administration of at least three antimicrobials might be beneficial for infections in critically ill patients caused by such pathogens.

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Action ofCoriandrum sativumL. Essential Oil upon OralCandida albicansBiofilm Formation

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2011/985832 ◽

2011 ◽

Vol 2011 ◽

pp. 1-9 ◽

Cited By ~ 38

Author(s):

V. F. Furletti ◽

I. P. Teixeira ◽

G. Obando-Pereda ◽

R. C. Mardegan ◽

A. Sartoratto ◽

...

Keyword(s):

Essential Oil ◽

Biofilm Formation ◽

Plant Material ◽

Inhibitory Concentration ◽

Chemical Characterization ◽

Synergistic Action ◽

Biofilm Growth ◽

Water Distillation ◽

Microdilution Method

The efficacy of extracts and essential oils fromAllium tuberosum, Coriandrum sativum, Cymbopogon martini, Cymbopogon winterianus,andSantolina chamaecyparissuswas evaluated againstCandidaspp. isolates from the oral cavity of patients with periodontal disease. The most active oil was fractionated and tested againstC. albicansbiofilm formation. The oils were obtained by water-distillation and the extracts were prepared with macerated dried plant material. The Minimal Inhibitory Concentration—MIC was determined by the microdilution method. Chemical characterization of oil constituents was performed using Gas Chromatography and Mass Spectrometry (GC-MS). C. sativum activity oil upon cell and biofilm morphology was evaluated by Scanning Electron Microscopy (SEM). The best activities against planktonicCandidaspp. were observed for the essential oil and the grouped F8–10fractions fromC. sativum. The crude oil also affected the biofilm formation inC. albicanscausing a decrease in the biofilm growth. Chemical analysis of the F8–10fractions detected as major active compounds, 2-hexen-1-ol, 3-hexen-1-ol and cyclodecane. Standards of these compounds tested grouped provided a stronger activity than the oil suggesting a synergistic action from the major oil constituents. The activity ofC. sativumoil demonstrates its potential for a new natural antifungal formulation.

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In VitroAntistaphylococcal Effects ofEmbelia schimperiExtracts and Their Component Embelin with Oxacillin and Tetracycline

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2015/175983 ◽

2015 ◽

Vol 2015 ◽

pp. 1-7 ◽

Cited By ~ 4

Author(s):

Johana Rondevaldova ◽

Olga Leuner ◽

Alemtshay Teka ◽

Ermias Lulekal ◽

Jaroslav Havlik ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Medicinal Plants ◽

Bacterial Infections ◽

Inhibitory Concentration ◽

Developed Countries ◽

Positive Interactions ◽

Fractional Inhibitory Concentration ◽

Broth Microdilution ◽

Microdilution Method ◽

Broth Microdilution Method

Bacterial infections are in less-developed countries traditionally treated by remedies prepared from medicinal plants.Embelia schimperi(Vatke) is a plant used as a taenicide or disinfectant in Ethiopia, very often taken mixed with another plant species. In the present study, we examined two extracts prepared from seeds and twigs with leaves ofE. schimperiand its main present secondary metabolite embelin for their antibacterial combinatory effect with oxacillin and tetracycline against sensitive and resistantStaphylococcus aureusstrains. Minimum inhibitory concentrations were determined through the broth microdilution method, whereas the combinatory effect was evaluated through fractional inhibitory concentration sum (ΣFIC) indices. Results show many positive interactions and synergy occurring in embelin and oxacillin combinations against 4 out of 9 strains (ΣFIC 0.203–0.477) and for embelin and tetracycline combination against 3 out of 9 strains (ΣFIC 0.400–0.496). Moreover, the resistance to oxacillin has been overcome in 2 strains and to tetracycline in 3 strains. According to our knowledge, this is the first study showing antimicrobial combinatory effect ofE. schimperias well as of embelin. These findings can be used for the further research targeted on the development of new antistaphylococcal agents.

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