Antimicrobial and Synergistic Activity of Metformin Against Helicobacter pylori

Background: The escalation of multidrug-resistant (MDR) Klebsiella pneumoniae infections poses a severe global health threat, necessitating the exploration of alternative therapeutic strategies such as drug repurposing. This study aims to investigate the intrinsic antibacterial activity of the antidiabetic drug metformin and its adjuvant potential when combined with conventional antibiotics against K. pneumoniae. Materials and Methods: The antimicrobial effects of metformin were evaluated against the reference strain K. pneumoniae ATCC 700603 and an MDR clinical isolate obtained from an intensive care unit. Minimum Inhibitory Concentration (MIC) values were determined using the microdilution method. Interaction studies between metformin and three antibiotics (Levofloxacin, Amikacin, and Ceftazidime) were conducted using the checkerboard assay. The results were interpreted based on the Fractional Inhibitory Concentration Index (FICI). Results: Metformin monotherapy exhibited weak intrinsic antibacterial activity (MIC ≥ 500 µg/mL) against both the reference strain and the clinical isolate. In combination assays, metformin demonstrated an antagonistic effect with Levofloxacin, while its interactions with Amikacin and Ceftazidime were classified as indifferent. The observed antagonism with Levofloxacin is attributed to metformin’s antioxidant properties, which likely mitigate the oxidative stress required for the bactericidal action of fluoroquinolones. Furthermore, the lack of synergy with Amikacin suggests that metformin-induced disruption of the Proton Motive Force (PMF) may hinder the intracellular uptake of aminoglycosides. Conclusion: The findings indicate that metformin is ineffective as a standalone antibacterial agent against K. pneumoniae. Moreover, its adjuvant potential is highly dependent on the antibiotic class and strain-specific resistance mechanisms. Contrary to some reports of synergy, this study highlights the risk of antagonism, particularly with fluoroquinolones. Further standardized mechanistic studies are required to define the specific therapeutic windows for metformin as an antimicrobial adjuvant.

The global proliferation of multidrug-resistant pathogens, particularly methicillin-resistant Staphylococcus aureus (MRSA), constitutes a severe threat to public health, rendering many conventional antibiotics obsolete. In the post-antibiotic era, where the pace of bacterial resistance evolution far exceeds that of new drug discovery, novel therapeutic strategies are urgently required. This study investigates the antibacterial potential of hyper-branched poly-L-lysine (HBPL), a synthetic antimicrobial polymer, against MRSA. We elucidate a multi-modal, physically disruptive mechanism of action initiated by electrostatic binding to the bacterial envelope, followed by rapid membrane permeabilization and cellular collapse, as visualized by electron and confocal microscopy. HBPL demonstrated potent, concentration-dependent bactericidal activity against clinical and standard MRSA strains, with a minimum inhibitory concentration (MIC) of 0.5 mg/mL and a minimum bactericidal concentration (MBC) of 1.0 mg/mL. Furthermore, HBPL inhibited biofilm formation by three MRSA strains (87, 73, and 81%) at a concentration of 1.0 mg/mL, which is a great advantage against the persistent and recalcitrant nature of biofilm-associated infections. Combination therapy studies using checkerboard assays revealed mechanistically dependent interactions. A synergistic effect [fractional inhibitory concentration index (FICI) ≤0.5] was observed with levofloxacin, attributed to HBPL-mediated membrane permeabilization enhancing intracellular drug access. Conversely, antagonism (FICI > 4) was noted with daptomycin, likely due to competitive binding at the bacterial membrane. These findings underscore the importance of rational drug pairing. While prolonged exposure induced stable, low-level resistance in MRSA, this was associated with adaptive cell envelope remodeling rather than target-site mutation. Collectively, this research establishes HBPL as a promising membrane-active agent and adjuvant therapy, capable of not only direct bactericidal action but also of restoring the efficacy of existing antibiotics against formidable pathogens like MRSA.

According to traditional Persian medicine, clove and marjoram have antibacterial properties. This study investigates their potential synergistic effects against oral pathogens. Ethanol extracts of clove and marjoram were tested for antimicrobial activity against Streptococcus mutans, Streptococcus salivarius, Streptococcus sanguinis, and Candida albicans using minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), time-kill assays, and fractional inhibitory concentration (FIC) indices. Clove extract demonstrated superior activity, with MIC/MBC values of 0.52/1 mg/mL for S. mutans, 0.2/1 mg/mL for S. salivarius and S. sanguinis, and 4/8 mg/mL for C. albicans, outperforming metronidazole and Persica. Marjoram extract also showed notable activity, especially against S. salivarius (MIC/MBC: 0.52/2.15 mg/mL). FIC analysis revealed additive effects for clove-marjoram combinations against S. mutans, S. sanguinis, and C. albicans, though a reducing effect was observed for S. salivarius. Time-kill studies confirmed enhanced bactericidal effects at 24 hours compared to reference treatments. The additive effects of clove and marjoram extracts show promise as supplementary agents to existing antimicrobial products. Further research is needed to evaluate their safety and efficacy in vivo.

Treatment of multidrug resistant (MDR) Acinetobacter baumannii infections causes some problems as a result of possessing various antibacterial resistance mechanisms against available antibiotics. Combination of antibiotics, acting by different mechanisms, is used for the treatment of MDR bacterial infections. It is an important factor to determine synergy or antagonism between agents in the combination for the constitution of effective therapy. The study aimed to determine in vitro interactions interpreted according to calculated fractional inhibitory concentration (FIC) index between sulbactam and ceftazidime, ceftriaxone, cefepime, ciprofloxacin, gentamicin, meropenem, tigecycline, and colistin. Ten clinical isolates of A. baumannii were tested for determination of synergistic effects of sulbactam with different antimicrobial combinations. Minimal inhibitory concentration (MIC) values of both sulbactam and combined antibiotics decreased 2- to 128-fold. Synergy and partial synergy were determined in combination of sulbactam with ceftazidime and gentamicin (FIC index: ≤0·5 or >0·5 to <1) and MIC values of both ceftazidime and gentamicin for five isolates fell down below the susceptibility break point. Similarly, MIC value of ciprofloxacin for six ciprofloxacin resistant isolates was determined as below the susceptibility break point in combination. However, all isolates were susceptible to colistin and tigecycline, MIC values of both were decreased in combination with sulbactam. Although synergistic and partial synergistic effects were observed in the combination of sulbactam and ceftriaxone, all isolates remained resistant to ceftriaxone. The effect of cefepime–sulbactam combination was synergy in five, partial synergy in one and indifferent in four isolates. Meropenem and sulbactam showed a partial synergistic effect (FIC index: >0·5 to <1) in three, an additive effect (FIC index: 1) in one and an indifferent effect (FIC index: >1–2) in six isolates. Antagonism was not determined in any combination for clinical A. baumannii isolates in the study. In conclusion, sulbactam is a good candidate for combination treatment regimes for MDR A. baumannii infections.

Methicillin-resistant Staphylococcus aureus (MRSA) is a deadly pathogen that initially was limited to hospital and healthcare facilities but has gradually became a growing problem in healthy children and adults. Pterostilbene belongs to the phenylpropanoid phytoalexin which is involved in plant response to various pathogen and herbivores attack. The aim of this study is to evaluate the anti-MRSA action of pterostilbene in combination with selected antibiotics; vancomycin, linezolid and oxacillin against ATCC 43300 and ATCC 33591. The minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and fractional inhibitory concentration (FIC) index values were determined. Microbroth dilution technique and microdilution checkerboard (MDC) assay were employed. The MIC and MBC of pterostilbene against ATCC 33591 was 31.25 and 62.50 µg mL-1, respectively. While for ATCC 43300, the MBC value was also twice (62.50 µg mL-1) its MIC value of 31.25 µg mL-1. This indicated that pterostilbene was bacteriostatic against both MRSA strains. Our MIC/MBC study also showed that linezolid exhibited bacteriostatic action but, oxacillin and vancomycin were bactericidal. MDC study showed that pterostilbene-oxacillin combination exhibited lowest FIC value (0.56) against both MRSA strains which indicated partial synergistic interaction. On the other hand, pterostilbene was additive (FIC 1.00) in combination with vancomycin whereas pterostilbene-linezolid combination displayed indifference effect with FIC of 1.25 against both MRSA strains. Pterostilbene in combination with oxacillin partially enhanced anti-MRSA activity with twofold reduction in MIC of oxacillin by acting at different site at the bacterial cell wall from that of oxacillin but more specific to the site of action of vancomycin.

Antibiotics are natural or synthetic substances used to inhibit or kill susceptible microorganisms. Due to the increase rate of bacterial resistance to antibiotics, there is the need to venture in other methods involving the coalescence effect of antibiotics and other substances such as cinnamon and alum to effectively eradicate these pathogenic organisms using less concentrated conventional antibiotics. The minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of the selected antimicrobial agents were determined using the broth dilution method, following standards and procedures set by the national council of clinical laboratory standards (NCCLS). The synergistic potentials of all the antimicrobics used were analyzed by the checkerboard assay. The MIC values of 25, 0.25, 0.75,25mg/ml were obtained for cinnamon extracts, gentamicin, penicillin G, and aq. Alum respectively against S. aureus. For E. coli O157H:H7, the MIC values of cinnamon extracts, gentamicin, penicillin G, and aq. Alum were 12.5, 0.25, 0.75 and 25mg/ml respectively,s while the MIC values of cinnamon extracts alum and fluconazole for C. albicans were 12.5, 25 and 0.05mg/ml respectively. There is a reduction in the MIC values for the various combinations against the same clinical microbes. All the antimicrobics used in this study inhibited the growth of the clinical microbes. All the microbes used in this study inhibited the tested microbes in dose dependent manner. The combinations of these agents resulted in reduction in MIC and MBC values, with an additive, indifference, and antagonistic effects using checkerboard assay which resulted to a fractional inhibitory concentration index which ranged from 0.56 to 7.9. Therefore, application of cinnamon, alum, and their combinations with conventional antibiotics can be used as treatment measure against infections caused by S. aureus, E. coli O157H:H7, and C. albicans.

BackgroundRapid emergence of multidrug resistant (MDR) organisms in hospital and community settings often result into treatment failure, thus leading the clinicians with fewer treatment options. Cyathea gigantea, an ethnomedicinally important fern used in cuts and wound infections. So, if this medicinal plant is used in treating the MDR infections then it might bring certain relief in future treatment options.MethodsAntibacterial activity of C. gigantea against MDR bacteria was assed using well diffusion and broth microdilution methods to determine the diameters of growth inhibition zones, minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). Synergistic activity was also determined with the conventional antibiotics by disc diffusion method followed by FIC index of each of the tested antibiotic was calculated. The active extract was then subjected to fractionation by column chromatography and antibacterial activity was done with each of the collected fractions.ResultsCrude extract of C. gigantea was found to be active against all the tested organisms. The MIC was 200 μg/ml against Gram-positive i.e., Staphylococcus aureus ATCC 25923 and 400 μg/ml against Gram-negative i.e., Escherichia coli ATCC 25922 and Pseudomonas aeruginosa PAO1, while the MBC was 400 μg/ml in case of Gram-positive and 800 μg/ml for Gram-negative. The synergistic activity revealed that the plant extract increased the antibacterial property of the studied antibiotics and the FIC index showed that significant synergistic activity was shown by ciprofloxacin followed by tetracycline, ampicillin and oxacillin. Antibacterial activity with the fractionated extract showed that the FR II, FR III and FR IV were active against both Gram-positive and Gram-negative bacteria, whereas FR I, FR V and FR VI did not show antibacterial property against any of the tested bacteria.ConclusionsExtracts of C. gigantea was found active against both selected Gram-positive and Gram-negative organisms and thus offers the scientific basis for the traditional use of the fern. The present study also provides the basis for future study to validate the possible use against multidrug resistant organisms.

The antibacterial activity of geraniol and its effect in combination with ampicillin, amoxicillin and clarithromycin against Staphylococcus aureus, Escherichia coli and Helicobacter pylori was tested. The minimum inhibitory concentrations (MICs) and combinatory effects of geraniol against the bacteria were assessed by using the modified broth microdilution and checkerboard assay, respectively. The combinatory effect is expressed as fractional inhibitory concentration index (FICI). The MIC of geraniol against S. aureus, E. coli and H. pylori was found to be 11200, 5600, and 7325 μg/mL, respectively. A significant synergistic effect was observed with geraniol and ampicillin against S. aureus with FICI in the range 0.19 to 0.32. Geraniol and ampicillin exhibited a partial synergistic effect against E. coli. A similar effect was observed with geraniol and clarithromycin against S. aureus. A partial synergistic effect was observed with clarithromycin and geraniol against H. pylori with the FICI value in the range 0.86 to 0.89. An additive effect was observed with geraniol and amoxicillin combination against H. pylori. However, the amoxicillin and clarithromycin dose was reduced by thirty-two fold when combined with geraniol against H. pylori. The anti- H. pylori effect of geraniol with clarithromycin and amoxicillin could be of potential interest in the treatment of H. pylori infection and associated ulcers in humans. Further, geraniol, in combination with other antibiotics, has substantial therapeutic potential against S. aureus and E.coli infection.

BackgroundMycoplasma genitalium has rapidly developed resistance to first-line azithromycin and second-line moxifloxacin. Third-line pristinamycin is difficult to source and only 80% effective. Consequently, there is an urgent need for alternative...

Helicobacter pylori is a Gram negative, helix-shaped and microaerophilic bacteria that colonizes the human gastric mucosa, causing chronic infections, gastritis, peptic ulcer, lymphomas associated with lymphoid mucosa tissue, and gastric cancer. H. pylori is considered as Type 1 human carcinogen by WHO. The prevalence of H. pylori infection is estimated in more than half of the world population. Treatment of H. pylori infection includes antibiotics and proton pump inhibitors, but the increasing antibiotic resistance promotes the research of novel, more effective and natural antibacterial compounds. The aim of this work was to study the effect of the partially purified proteolytic extract (granulosain) from the fruits of Solanum granuloso-leprosum (Dunal), a South American native plant, against Helicobacter pylori; in order to obtain a natural food additive or ingredient for the production of anti-H. pylori functional foods. Furthermore, granulosain could also be used as a natural adjunct to conventional therapies. Granulosain antibacterial activity was evaluated as minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against H. pylori NCTC 11638 (reference strain) and four H. pylori wild strains; using a micro dilution plating technique (Clinical and Laboratory Standards Institute). All the strains tested were susceptible to granulosain with MIC and MBC values of 312.5  0.01 µg/mL. The effect of granulosain on the transcription of H. pylori virulence genes: omp18, ureA and flaA, with respect to a housekeeping gene (16S rRNA), was evaluated by RT-PCR technique. The band intensity between virulence and control genes was correlated under treated/untreated conditions, using the ImageJ program. Granulosain significantly decreased the expression of the omp18, ureA, flaA genes (p < 0.05). Finally, the combined inhibitory effect of granulosain and a conventional antibiotic, such as: amoxicillin (AML, 10 g), clarithromycin (CLA, 15 g), levofloxacin (LEV, 5 g) or metronidazole (MTZ, 5 g) was evaluated, using the agar diffusion technique. Granulosain showed significant synergistic effect on AML, CLA and LEV, but none effect on MTZ. These pioneering results indicate that granulosain, could be used as natural food additive for the production of anti-H. pylori functional foods and as natural adjuvant compound for conventional therapies against H. pylori strains.

目的探讨反-2-癸烯醛体外抑制和杀灭幽门螺杆菌（Helicobacter pylori, H. pylori）的能力及潜在机制。方法采用微量肉汤稀释法得到反-2-癸烯醛对临床分离H. pylori以及标准菌株ATCC43504的最低抑菌浓度（minimum inhibitory concentration, MIC）和最低杀菌浓度（minimum bactericidal concentration, MBC）；通过观察MIC值的变化，分析反-2-癸烯醛连续诱导H. pylori后的耐药情况。采用分数抑制浓度指数评估反-2-癸烯醛与抗生素甲硝唑（metronidazole, MTZ）、克拉霉素（clarithromycin, CLA）、阿莫西林（amoxicillin, AMX）和左氧氟沙星（levofloxacin, LEV）的协同抑菌作用。用反-2-癸烯醛处理H. pylori标准菌株，通过扫描电镜与透射电镜、半固体穿刺和Berthelot法观察c对H. pylori形态及细胞结构、迁移力以及脲酶活性的影响。结果不同H. pylori 临床耐药株（C907、C101、R1、R2、R4、R10、R16、R24、R36和R40）MIC范围为8～16 μg/mL，MBC范围为16～32 μg/mL。H. pylori标准菌株ATCC 43504 MIC为16 μg/mL，MBC为16 μg/mL。反-2-癸烯醛能在低浓度、短时间内抑制H. pylori，且不易诱导H. pylori产生耐药性。与MTZ和LEV联用可降低抗生素使用剂量并呈相加效应。反-2-癸烯醛通过改变H. pylori形状以及诱导菌体破裂、抑制其生物膜形成、减少成熟生物膜数量、降低其脲酶活性和H. pylori迁移力发挥作用。结论反-2-癸烯醛可能通过多种机制在体外发挥抑制H. pylori的作用且不易导致H. pylori产生耐药性。

Background and Aim:The gradual loss of efficacy of conventional antibiotics is a global issue. Plant material extracts and green-synthesized nanoparticles are among the most promising options to address this problem. Therefore, the aim of this study was to assess the antibacterial properties of aqueous and hydroalcoholic extracts of grapefruit peels as well as their inclusion in green-synthesized silver nanoparticles (AgNPs).Materials and Methods:Aqueous and hydroalcoholic extracts (80% v/v) were prepared, and the volume and mass yields were determined. The synthesis of AgNPs was done in an eco-friendly manner using AgNO3 as a precursor. The nanoparticles were characterized by ultraviolet–vis spectrometry and photon cross-correlation spectroscopy. The antibacterial activity of the extracts was tested on three Gram-positive bacteria (Staphylococcus aureus ATCC 6538, clinical Enterococcus faecalis, and S. aureus) and two Gram-negative bacteria (two clinical Escherichia coli) using various concentrations of extracts (100, 50, 25, 12, and 5 mg/mL and 5% dimethyl sulfoxide as negative control). Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined using the microdilution method. Modulation of cefazoline and ampicillin on resistant E. coli and S. aureus strains was added to the mixture design response surface methodology with extreme vertices design, with the diameters of inhibition and the fractional inhibitory concentration index as responses and factors, respectively. The antibiotic, the ethanolic extract, and water varied from 0.1 MIC to 0.9 MIC for the first two and from 0 to 0.8 in proportion for the third. Validating the models was done by calculating the absolute average deviation, bias factor, and accuracy factor.Results:The volume yield of the EE and aqueous extract (AE) was 96.2% and 93.8% (v/v), respectively, whereas their mass yields were 7.84% and 9.41% (m/m), respectively. The synthesized AgNPs were very uniform and homogeneous, and their size was dependent on the concentration of AgNO3. The antibacterial activity of the two extracts was dose-dependent, and the largest inhibition diameter was observed for the Gram-positive bacteria (S. aureus ATCC 6538; AE, 12; EE, 16), whereas AgNPs had a greater effect on Gram-negative bacteria. The MICs (mg/mL) of the AEs varied from 3.125 (S. aureus ATCC 6538) to 12.5 (E. coli 1 and E. coli 2), whereas the MICs of the EEs varied from 1.5625 (S. aureus 1, S. aureus ATCC 6538, and E. faecalis) to 6.25 (E. coli 1). There was a significant difference between the MICs of AEs and EEs (p=0.014). The MBCs (mg/mL) of the AEs varied from 12.5 (S. aureus ATCC 6538) to 50 (S. aureus 1), whereas those of the EEs varied from 6.25 (S. aureus 1) to 25 (E. coli 1 and E. faecalis). Ethanolic grapefruit extracts demonstrated an ability to modulate cefazolin on E. coli and S. aureus but were completely indifferent to ampicillin on E. coli.Conclusion:Grapefruit peel extracts and their AgNPs exhibit antibacterial properties that can be exploited for the synthesis of new antimicrobials and their EEs may be efficiently used synergistically with other antibiotics against bacteria with intermediate susceptibility.

IntroductionDiabetes mellitus is a chronic metabolic disorder characterized by persistent hyperglycemia due to impaired insulin production or utilization, leading to severe health complications. Diabetic foot ulcers (DFUs) represent a major complication, often exacerbated by polymicrobial infections involving Staphylococcus aureus and Acinetobacter baumannii. These pathogens, notorious for their resistance to antibiotics, complicate treatment efforts, especially due to biofilm formation, which enhances bacterial survival and resistance. This study explores the synergistic effects of combining gentamicin, imipenem, and fucoidan, a sulfated polysaccharide with antimicrobial properties, against both planktonic and biofilm forms of S. aureus and A. baumannii.MethodsIsolates of S. aureus and A. baumannii were collected from DFUs and genetically confirmed. Methicillin resistance in S. aureus was identified through disk diffusion and PCR. Biofilm formation, including dual-species biofilms, was analyzed using the microtiter plate method. The antimicrobial efficacy of gentamicin, imipenem, and fucoidan was assessed by determining the minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), minimum biofilm inhibitory concentration (MBIC), and minimum biofilm eradication concentration (MBEC). Synergistic interactions were evaluated using the fractional inhibitory concentration index (FICi) and fractional bactericidal concentration index (FBCi). The expression of biofilm-associated genes (icaA in S. aureus and bap in A. baumannii) was analyzed, and the cytotoxicity of fucoidan was assessed.ResultsThe study revealed that 77.4% of S. aureus and all A. baumannii isolates showed multidrug resistance. Among 837 tested conditions for dual-species biofilm formation, 72 resulted in strong biofilm formation and 67 in moderate biofilm formation. The geometric mean MIC values for gentamicin were 12.2 µg/mL for S. aureus, 22.62 µg/mL for A. baumannii, and 5.87 µg/mL for their co-culture; for imipenem, they were 19.84, 9.18, and 3.70 µg/mL, respectively, and for fucoidan, 48.50, 31.20, and 19.65 µg/mL, respectively. The MBC values for gentamicin were 119.42, 128, and 11.75 µg/mL; for imipenem, they were 48.50, 14.92, and 8 µg/mL; and for fucoidan, they were 88.37, 62.62, and 42.48 µg/mL. The MBIC values were 55.71, 119.42, and 18.66 µg/mL for gentamicin; 68.59, 48.50, and 25.39 µg/mL for imipenem; and 153.89, 101.49, and 53.53 µg/mL for fucoidan. The MBEC values were 315.17, 362.03, and 59.25 µg/mL for gentamicin; 207.93, 157.58, and 74.65 µg/mL for imipenem; and 353.55, 189.46, and 99.19 µg/mL for fucoidan. When cultured in planktonic form, the geometric mean FICi and FBCi values indicated additive effects, while co-culture showed FICi values of ≤ 0.5, suggesting a synergistic interaction. Treatment with gentamicin and fucoidan led to significant downregulation of the icaA and bap genes in both single-species and dual-species biofilms of S. aureus and A. baumannii. The reductions in gene expression were more pronounced in dual-species biofilms compared to single-species biofilms. Additionally, treatment with imipenem and fucoidan also resulted in significant downregulation of these genes in both biofilm types. Cytotoxicity assessments indicated that higher concentrations of fucoidan were toxic, yet no harmful effects were noted at the optimal synergistic concentrations used with antibiotics.ConclusionIn our investigation, we found that combining gentamicin, imipenem, and fucoidan had a synergistic effect on dual-species biofilms of S. aureus and A. baumannii, suggesting potential benefits for treating such infections effectively. This underscores the importance of understanding microbial interactions, antibiotic susceptibility, and biofilm formation in DFUs.

Relevance. Urinary tract infections pose a growing threat to humanity due to the rise of antibiotic resistance in uropathogens. Exploring natural sources for alternative treatments has become a prominent approach. The aim of the research was to investigate the antibacterial effects of clove (Syzygium aromaticum L.) against uropathogenic Escherichia coli (E. coli). Materials and Methods. The research was performed on three clinical multidrug-resistant uropathogenic E. coli isolates and E. coli ATCC 25922. Clove hydroalcoholic extract was obtained by cold maceration technique. To evaluate the antibacterial activity of the extract, agar well diffusion method was performed. Minimum inhibitory and minimum bactericidal concentrations of the extract were determined by microbroth dilution method. Light microscopy was used to investigate morphological changes in uropathogenic E. coli after exposure to clove extract. Checkerboard assay was used to assess synergism between clove extract and antibiotics. All obtained data were statistically processed. Results and Discussion. In well diffusion method, bacterial responses to clove extract were concentration-dependent with inhibition zone diameter of 7-10/10-15 mm for uropathogenic strains and E. coli ATCC 25922, respectively. Minimum inhibitory and minimum bactericidal concentrations of clove extract against uropathogenic strains were 25 mg/mL. The extract showed a lower minimum inhibitory concentration against E. coli ATCC 25922 (6.25 mg/ mL) with minimum bactericidal concentration being 25 mg/mL. Minimum inhibitory and minimum bactericidal concentrations ratio showed that clove extract tends to be bactericidal agent. Synergy test revealed that the combination of clove extract and nitrofurantoin or ciprofloxacin resulted in no interaction. However, minimum inhibitory concentrations of all tested agents in combinations exhibited varying degrees of decrease. Incubation of uropathogenic strains with the extract transformed them to unstable spherical L-form in percentage of 96-99 %. Conclusion. This study highlights clove as a potential natural antibacterial agent against multidrug-resistant uropathogenic E. coli, warranting further investigations into its antibacterial properties.

To investigate the antimicrobial efficacy of an alkaloid, harmaline alone and in combination with chlorhexidine digluconate (CHG) against clinical isolates of Staphylococcus aureus (S.aureus) grown in planktonic and biofilm cultures. Minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) were determined for each micro-organism grown in suspension and in biofilm using microbroth dilution method. Chequerboard assays were used to determine synergistic, indifferent or antagonistic interactions between harmaline and CHG, and the some of results were verified by confocal laser scanning microscopy. Harmaline and CHG showed effective antimicrobial activity against suspensions and biofilm cultures of S.aureus, respectively. As determined by fractional inhibitory concentration index (FICI), synergistic antimicrobial effects between harmaline and CHG were observed in nine and 11 of the 13 S.aureus strains when in suspension and in biofilm, respectively. FICI values were from 0·375 to 1·25 when in suspension and from 0·25 to 1·25 when in biofilm. Synergistic activity of harmaline and CHG against clinical isolates of S.aureus (in suspension and in biofilm) was observed in vitro. This study might provide alternative methods to overcome the problem of drug-resistance of S.aureus both in suspension and in biofilm.