Checkerboard Microdilution Method Research Articles

Antimicrobial and synergistic effects of lemongrass and geranium essential oils against Streptococcus mutans, Staphylococcus aureus, and Candida spp.

The oral cavity harbors more than 700 species of bacteria, which play crucial roles in the development of various oral diseases including caries, endodontic infection, periodontal infection, and diverse oral diseases. To investigate the antimicrobial action of Cymbopogon Schoenanthus and Pelargonium graveolens essential oils against Streptococcus mutans, Staphylococcus aureus, Candida albicans, Ca. dubliniensis, and Ca. krusei. Minimum microbicidal concentration was determined following Clinical and Laboratory Standards Institute documents. The synergistic antimicrobial activity was evaluated using the Broth microdilution checkerboard method, and the antibiofilm activity was evaluated with the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide assay. Data were analyzed by one-way analysis of variance followed by the Tukey post-hoc test (P ≤ 0.05). C. schoenanthus and P. graveolens essential oils were as effective as 0.12% chlorhexidine against S. mutans and St. aureus monotypic biofilms after 24 h. After 24 h P. graveolens essential oil at 0.25% was more effective than the nystatin group, and C. schoenanthus essential oil at 0.25% was as effective as the nystatin group. C. schoenanthus and P. graveolens essential oils are effective against S. mutans, St. aureus, Ca. albicans, Ca. dubliniensis, and Ca. krusei at different concentrations after 5 min and 24 h.

Antimicrobial Effect of Lippia citriodora Extract in Combination with Gallic Acid or Octyl Gallate on Bacteria from Meat.

Chicken meat and its derivatives are easily alterable. They are a nutritionally healthy food, and their consumption has seen a remarkable increase worldwide in recent years. At the same time, consumer demand for the use of natural products to control microbial growth is increasing. In this context, the antimicrobial capacity of a commercial extract of the lemon verbena (Lippia citriodora) plant, (LCE) was tested in binary combination with gallic acid or octyl gallate against two strains of lactic acid bacteria (LAB) of meat origin: Carnobacterium divergens ATCC 35677 and Leuconostoc carnosum ATCC 49367. First, the antimicrobial potential was evaluated by the checkerboard microdilution method at the optimal growth temperature of each and at 4 °C, pH 5.7 and 6.7, in culture medium. Octyl gallate was the most effective antimicrobial against the two bacteria under all study conditions. At 4 °C, the combination of LCE with octyl gallate had a similar antimicrobial effect on the two LAB, being bactericidal at pH 6.7. In chicken breast, this effective combination was tested in normal or modified atmosphere and refrigerated (4-8 °C) for 9 days. LCE + OG in modified atmosphere reduced the different microbial groups studied, including the lactic acid bacteria as the main microorganisms responsible for the spoilage of fresh meat. Further research could pave the way for the development of novel strategies contributing to the technological stability, security, and functional properties of chicken meat.

Study on synergistic effects of curcumin and bixin against foodborne pathogens.

Various studies have shown that natural colorants, in addition to their coloring attributes, have valuable biological effects such as antioxidant, anti-inflammation, and anticarcinogenic properties. Moreover, their use as a food colorant can restrict the potential disadvantages of synthetic additives and turn foods into functional products. In this study, invitro antimicrobial activities of two natural colorants of bixin and curcumin against some important foodborne pathogens: Staphylococcus aureus (S. aureus), Listeria innocua (L. innocua), and Escherichia coli (E. coli) were investigated by disk diffusion method. Minimum inhibitory concentration and minimum bactericidal concentration values were determined by agar dilution and broth microdilution methods. The synergistic activity of the colorants against selected microorganisms was assayed by the checkerboard microdilution method. The results showed that the inhibitory effects of bixin against S. aureus were more pronounced than E. coli and L. innocua. The lowest concentration of curcumin (0.6 mg/mL) in the disk diffusion method was not inhibited by any tested bacteria. However, it was effective at the higher concentrations against three microorganisms, but its diameter of inhibition zones was lower than gentamicin in all concentrations. Synergetic effects were observed by curcumin and bixin combination against S. aureus (FICI ≤ 0.5), but they act as an antagonist against E. coli and L. innocua. The results of the synergy test were confirmed by the isobologram curves.

The Therapeutic Role and Mechanism of Glabridin Under Aspergillus fumigatus Infection.

Purpose: To characterize the efficiency of glabridin alone and in combination with clinical antifungals in Aspergillus fumigatus keratitis. Methods: The broth microdilution method was performed to investigate whether glabridin exerted an antifungal role on planktonic cells and immature and mature biofilm. Antifungal mechanism was evaluated by Sorbitol and Ergosterol Assays. The synergistic effect of glabridin and antifungals was assessed through the checkerboard microdilution method and time-killing test. Regarding anti-inflammatory role, inflammatory substances induced by A. fumigatus were assessed by real-time quantitative polymerase chain reaction, western blot, and enzyme-linked immunosorbent assay. Drug toxicity was assessed by Draize test in vivo. Macrophage phenotypes were examined by flow cytometry. Results: Regarding antifungal activity, glabridin destroyed fungal cell wall and membrane on planktonic cells and suppressed immature and mature biofilm formation. After combining with natamycin or amphotericin B, glabridin possessed a potent synergistic effect against A. fumigatus. Regarding anti-inflammatory aspects, Dectin-1, toll‑like receptor (TLR)-2 and TLR-4 expression of human corneal epithelial cells were significantly elevated after A. fumigatus challenge and reduced by glabridin. The elevated expression of interleukin-1β and tumor necrosis factor-alpha induced by A. fumigatus or corresponding agonists were reversed by glabridin, equivalent to the effect of corresponding inhibitors. Glabridin could also contribute to anti-inflammation by downregulating inflammatory mediator expression to suppress macrophage infiltration. Conclusions: Glabridin contributed to fungal clearance by destroying fungal cell wall and membrane, and disrupting biofilm. Combining glabridin with clinical antifungals was superior in reducing A. fumigatus growth. Glabridin exerted an anti-inflammatory effect by downregulating proinflammatory substance expression and inhibiting macrophage infiltration, which provide a potential agent and treatment strategies for fungal keratitis.

Pythium insidiosum: In vitro oomicidal evaluation of telithromycin and interactions with azithromycin and amorolfine hydrochloride

This study evaluated the repositioning of the ketolide antibacterial telithromycin (TLT) against the oomycete Pythium insidiosum and verified the combination of TLT and the antimicrobials azithromycin (AZM) and amorolfine hydrochloride (AMR), which have known anti-P. insidiosum activity. Susceptibility tests of P. insidiosum isolates (n = 20) against the drugs were carried out according to CLSI protocol M38-A2, and their combinations were evaluated using the checkerboard microdilution method. The minimum inhibitory concentrations were 0.5–4 µg/mL for TLT, 2–32 µg/mL for AZM, and 16–64 µg/mL for AMR. For the TLT+AZM combination, 52.75 % of interactions were indifferent, 43.44 % were antagonistic, and 9.70 % were synergistic. As for interactions of the TLT+AMR combination, 60.43 % were indifferent, 39.12 % were antagonistic, and 10.44 % synergistic interactions. This study is the first to evaluate the repositioning of the antibacterial TLT against mammalian pathogenic oomycetes, and our results show that its isolated action is superior to its combinations with either AZM or AMR. Therefore, we recommend including TLT in future research to evaluate therapeutic approaches in different clinical forms of human and animal pythiosis.

Effects of chitooligosaccharide on the in vitro antibacterial activity against avian Escherichia coli and the pharmacokinetics of florfenicol in healthy chickens

This study investigates the combined effects of chitooligosaccharide (COS) and florfenicol (FLO) on the inhibition of Escherichia coli in vitro, as well as the pharmacokinetic interactions between these compounds in healthy chickens. The minimum inhibitory concentration (MIC) of COS and FLO alone and the fractional inhibitory concentration index (FICI) after combined treatment were determined using the broth microdilution method and checkerboard method, respectively. Additionally, we evaluated the pharmacokinetic interactions between the 2 types of COS and FLO in healthy chickens. Thirty chickens were randomly divided into 3 groups: Florfenicol group (30 mg/kg), COS J85 group (COS J85 20 mg/kg + florfenicol 30 mg/kg), COS H85 group (COS H85 20 mg/kg + florfenicol 30 mg/kg). Either FLO or COS was orally administered by gavage. The concentrations of FLO in chicken plasma were measured at different time points after the drug withdrawal using high-performance liquid chromatography (HPLC), and pharmacokinetic parameters were calculated by a compartmental method. The results showed that COS J85 and COS H85, when combined with FLO, had FICI values of 0.1875 to 0.75 and 0.3125 to 1, respectively, indicating good synergistic or additive effects against Escherichia coli. The pharmacokinetics of FLO alone and in combination with COS followed a 1-compartment model with first-order absorption and elimination. Furthermore, the pharmacokinetic analysis revealed that the elimination half-life (t1/2ke) of florfenicol was significantly increased in the COS H85 group compared to oral administration of florfenicol alone (P < 0.05). Other pharmacokinetic parameters did not show significant changes (P > 0.05), except between the 2 combined treatment groups, where statistical differences were observed for various parameters, excluding the area under the concentration-time curve from the time of dosing to infinity (AUC) and peak concentration (Cmax).

Antifungal Biofilm Inhibitory Effects of Combinations of Diclofenac and Essential Oils.

Systemic fungal infections have risen in recent decades and most of them are caused by Candida species, which are becoming increasingly resistant to conventional antifungal drugs. Biofilm production has been considered the most common growth form of Candida cells and is associated with a high level of antifungal resistance. At present, international research reports on the antifungal activity of non-traditional antimicrobial drugs and their potential use against life-threatening resistant fungal infections. Indeed, drug repurposing has led to the consideration of well-known compounds as a last-line therapy. The goal of this work is to evaluate the potential synergistic antifungal biofilm activity of new combinations between diclofenac sodium salt (DSS), a widely used non-steroidal anti-inflammatory drug (NSAID), with the essential oils (EOs) of Mentha piperita, Pelargonium graveolens, and Melaleuca alternifolia, whose antifungal activity has been well documented over the years. The in vitro antifungal activity of DSS and EOs was determined on different Candida strains. Susceptibility testing and the synergism of DSS and EOs versus biofilm cells was performed by using the broth microdilution assay and checkerboard methods. Minimum inhibitory concentrations (sMIC50) of DSS alone ranged from 1.25 to 2.05 mg/mL for all the strains considered. These values significantly decreased when the drug was used in combination with the EOs. The fractional inhibitory concentration index (FICI) was lower than 0.5 for almost all the associations, thus indicating a significant synergism, particularly for the DSS-Pelargonium graveolens combination towards the Candida strains examined. These preliminary results show that the combination of the EOs with DSS improves the antifungal activity on all the tested Candida strains, significantly lowering the concentrations of the components used and thus allowing any toxic effects to be overcome.

Rapamycin and caspofungin show synergistic antifungal effects in caspofungin-susceptible and caspofungin-resistant Candida strains in vitro.

Caspofungin is an echinocandin antifungal agent that inhibits synthesis of glucan required for the fungal cell wall. Resistance is mediated by mutation of Fks1 glucan synthase, among which S645P is the most common resistance-associated polymorphism. Rapamycin is a macrolide that inhibits the mechanistic target of rapamycin (mTOR) protein kinase activity. This study investigated the interaction between rapamycin and caspofungin in inhibiting the growth of WT Candida albicans and Fks1 S645P mutant clinical isolate, and WT Candida lusitaniae and genetically engineered isogenic strain with Fks1 S645P mutation at equivalent position. Interactions between caspofungin and rapamycin were evaluated using the microdilution chequerboard method in liquid medium. The results were analysed using the Loewe additivity model (FIC index, FICI) and the Bliss independence model (response surface, RS, analysis). Synergy between rapamycin and caspofungin was shown for C. albicans and C. lusitaniae strains by RS analysis of the chequerboard tests. Synergy was observed in strains susceptible and resistant to caspofungin. Weak subinhibitory concentrations of rapamycin were sufficient to restore caspofungin susceptibility. We report here, for the first time, synergy between caspofungin and rapamycin in Candida species. Synergy was shown for strains susceptible and resistant to caspofungin. This study highlights the possible implication of the TOR pathway in sensing antifungal-mediated cell wall stress and in modulating the cellular response to echinocandins in Candida yeasts.

Molecular Epidemiology of Carbapenem Resistant Klebsiella Pneumoniae in Northern China: Clinical Characteristics, Antimicrobial Resistance, Virulence and Geographic Distribution.

In this article, we studied in detail 74 Carbapenem Resistant Klebsiella pneumoniae (CRKP) in Shanxi to provide essential insight into development of effective strategies for control of CRKP. From 2018 to 2021, we collected 74 clinical CRKP from 11 hospitals in Shanxi Province. Clinical data were obtained from medical records, and all isolates were subjected to antimicrobial susceptibility testing, multi locus sequence typing, capsular serotypes, resistant gene profiles and virulence gene profiles. The synergistic activity was performed by microdilution checkerboard method. Our study found differences in the clinical characteristics of CRKP between regions in Shanxi. Sequence type (ST) 11 was the dominant ST in Shanxi; however, the ST types in Shanxi had become more diverse over time and the proportion of STs showed a more balanced distribution with a significant decrease in ST11. NDM was the most common carbapenemase in Shanxi. In addition, the STs, carbapenemases, serotypes and virulence gene distribution varied by region in Shanxi. Moreover, tigecycline in combination with carbapenems and aztreonam had an excellent synergistic effect on CRKP in vitro. The results of this study provide essential insight into development of effective strategies for control of CRKP in Shanxi.

Photodynamic Activity of Chlorophyllin and Polyethylenimine on Pseudomonas aeruginosa Planktonic, Biofilm and Persister Cells.

Antimicrobial photodynamic inactivation is considered a promising antimicrobial approach that may not develop resistance in the near future. Here, we investigate the influence of the photosensitizer chlorophyllin (CHL) and the cationic permeabilizer polyethylenimine (PEI), exposed to a red light-emitting diode, on the human pathogen Pseudomonas aeruginosa free-living planktonic cells, the sessile biofilm and persister cells. The broth microdilution checkerboard method was used to test antimicrobial susceptibility. As a substrate for biofilms, the Calgary biofilm device was used, and the quantification of the biofilm biomass was carried out using a crystal violet assay. Serine hydroxamate was used for the induction of persisters. Our findings reveal that PEI ameliorates the antimicrobial activity of CHL against P. aeruginosa planktonic and biofilm states, and the concentration required to eradicate the bacteria in the biofilm is more than fourfold that is required to eradicate planktonic cells. Interestingly, the persister cells are more susceptible to CHL/PEI (31.25/100 µg mL-1) than the growing cells by 1.7 ± 0.12 and 0.4 ± 0.1 log10 reduction, respectively, after 15 min of illumination. These data demonstrate that CHL excited with red light together with PEI is promising for the eradication of P. aeruginosa, and the susceptibility of P. aeruginosa to CHL/PEI is influenced by the concentrations and the exposure time.

Synergistic activity of the combination of falcarindiol and itraconazole in vitro against dermatophytes.

Previous studies have shown that natural polyacetylene alcohols, such as falcarindiol (FADOH), have good antifungal effects on plant fungi. While its effect on fungi that infect humans remains to be explored. In our study, checkerboard microdilution, drop-plate assay, and time-growth method were employed to analyze the interactions between FADOH and itraconazole (ITC) in vitro against dermatophytes, including 12 Trichophyton rubrum (T. rubrum), 12 Trichophyton mentagrophytes (T. mentagrophytes), and 6 Microsporum canis (M. canis). The results showed that the combination of FADOH and ITC exhibited synergistic and additive activity against 86.7% of all tested dermatophytes. FADOH had an excellent synergistic effect on ITC against T. rubrum and T. mentagrophytes; the synergistic rates were 66.7% and 58.3%, respectively. On the contrary, FADOH combined with ITC showed poor synergistic inhibitory activity (16.7%) against M. canis. Moreover, the additive rates of these two drugs against T. rubrum, T. mentagrophytes, and M. canis were 25%, 41.7%, and 33.3%, respectively. No antagonistic interactions were observed. The drop-plate assay and time-growth curves confirmed that the combination of FADOH and ITC had a potent synergistic antifungal effect. The in vitro synergistic effect of FADOH and ITC against dermatophytes is reported here for the first time. Our findings suggest the potential use of FADOH as an effective antifungal drug in the combined therapy of dermatophytoses caused especially by T. rubrum and T. mentagrophytes.

Lower Concentrations of Amphotericin B Combined with Ent-Hardwickiic Acid Are Effective against Candida Strains

Life-threatening Candida infections have increased with the COVID-19 pandemic, and the already limited arsenal of antifungal drugs has become even more restricted due to its side effects associated with complications after SARS-CoV-2 infection. Drug combination strategies have the potential to reduce the risk of side effects without loss of therapeutic efficacy. The aim of this study was to evaluate the combination of ent-hardwickiic acid with low concentrations of amphotericin B against Candida strains. The minimum inhibitory concentration (MIC) values were determined for amphotericin B and ent-hardwickiic acid as isolated compounds and for 77 combinations of amphotericin B and ent-hardwickiic acid concentrations that were assessed by using the checkerboard microdilution method. Time-kill assays were performed in order to assess the fungistatic or fungicidal nature of the different combinations. The strategy of combining both compounds markedly reduced the MIC values from 16 µg/mL to 1 µg/mL of amphotericin B and from 12.5 µg/mL to 6.25 µg/mL of ent-hardwickiic acid, from isolated to combined, against C. albicans resistant to azoles. The combination of 1 µg/mL of amphotericin B with 6.25 µg/mL of ent-hardwickiic acid killed all the cells of the same strain within four hours of incubation.

665. Modified Superposition in vitro Susceptibility of Aztreonam and Ceftazidime/Avibactam for an NDM-5 Metallo-β-Lactamase-Producing Klebsiella pneumoniae Isolate

Abstract Background We describe a novel in vitro susceptibility testing approach of aztreonam and ceftazidime/avibactam using a superposition method paired with confirmatory testing and molecular sequencing of a New Delhi metallo-β-lactamase (NDM)-5-producing Klebsiella pneumoniae. Methods A modified strip superposition method with Epsilometer test (Etest) strips and Kirby Bauer disks was utilized to test the in vitro activity of aztreonam and ceftazidime/avibactam against the NDM-producing K. pneumoniae. The isolate was also sent for whole genome sequencing and tested for the in vitro activity of aztreonam and ceftazidime in combination with avibactam using a broth microdilution checkerboard assay. Aztreonam and ceftazidime/avibactam in vitro testing with modified superposition method Results All zones of inhibition from the Etest superposition method were measured to be 22 mm in diameter (Clinical and Laboratory Standards Institute [CLSI] zone of inhibition for aztreonam susceptibility is ≥ 21 mm for Enterobacterales). The results of whole genome sequencing showed the K. pneumoniae isolate carried a metallo-β-lactamase (blaNDM-5), extended-spectrum β-lactamase (blaCTX-M-15), and other β-lactamases (blaSHV-11, blaTEM-1). Through the broth microdilution checkerboard method, the isolate demonstrated inhibition by aztreonam/avibactam (fixed concentration of 4mg/L) at a minimum inhibitory concentration (MIC) of 0.25 mg/L but was resistant to ceftazidime/avibactam (MIC &amp;gt; 16 mg/L) and aztreonam alone (MIC &amp;gt; 16 mg/L) according to CLSI breakpoints. Conclusion The in vitro activity of aztreonam and ceftazidime/avibactam against an NDM-5-producing K. pneumoniae was demonstrated with a modified superposition method. This may serve as a simplistic and readily available technique for rapidly determining antimicrobial activity in these difficult to treat pathogens. Disclosures Justin Schmetterer, PharmD, PhC, BCIDP, AbbVie Speaker Bureau: Honoraria Yao Lin, MS, Abbvie Inc: employee|Abbvie Inc: Stocks/Bonds.

SINERGISTIC COMBINATION OF ALOE VERA AND KERSEN LEAF EXTRACTS FOR INHIBITION OF THE GROWTH OF Staphylococcus aureus 25923

Aloe vera leaf (Aloe vera L.) and cherry leaf (Muntingia calabura L.) contain bioactive compounds that serve as the potential antibacterial agent. The combination treatment of multiple extracts can produce a stronger effect than a single compound. This study aims to determine the synergistic combination of aloe vera leaf extract (EDLB) and cherry leaf extract (EDK) as an antibacterial agent against Staphylococcus aureus ATCC 25923. The extract was obtained by maceration using 96% ethanol. Chemical content in the extract was analyzed by tube test and thin layer chromatography (TLC). Antibacterial activity against S. aureus ATCC 25923 was measured by the checkerboard microdilution method on a 96 well microplate. The effect of the combination of extracts on antibacterial activity was based on the fractional inhibitory concentration index (FICI) value. EDLB and EDK contain flavonoids, tannins, alkaloids, and saponins. The second combination showed a synergistic effect with a FICI value of 0.5 in inhibiting the growth of S. aureus with minimal inhibitory levels in a mixture of 0.625 mg/mL EDLB and 2.5 mg/mL EDK.

In Vitro Combination of Terbinafine with Ketoconazole Against Aspergillus Species with Terbinafine High MIC Values Isolated From Otomycosis.

Otomycosis is a common mycotic infection of the external auditory canal, and Aspergillus species are one of the most frequent causative agents worldwide. The limited antifungal arsenal, the high toxicity and side effects of antifungal agents, and the growing resistance to the currently available antifungals underscore the need for new therapeutic strategies. The present study aimed to evaluate the combined in vitro efficacy of terbinafine and ketoconazole against Aspergillus species with terbinafine high MIC values isolated from patients with otomycosis.84 Aspergillus species with high MIC values to terbinafine (≥ 4µg/ml), consisting of A. flavus, A. tubingensis, A. niger, and A. terreus, were included in this study. The checkerboard microdilution method evaluated the in vitro interactions using the CLSI reference technique. Synergistic effects were observed for 66.67% (56/84) of all isolates (FICI ranging from 0.19 to 0.5). However, the interactions of terbinafine and ketoconazole exhibited indifference in 33.33% (28/84) of the isolates, and no antagonism was observed for any combination. The interaction of terbinafine and ketoconazole showed synergistic activity against Aspergillus species with high MIC values, suggesting that this is an alternative and promising approach for treating otomycosis.

Synergistic Effect of Clinically Available Beta-Lactamase Inhibitors Combined with Cefiderocol against Carbapenemase-Producing Gram-Negative Organisms.

The role of β-lactamases in reduced susceptibility or resistance to cefiderocol has been supported by recent reports. The purpose of this study was to investigate the in vitro impact of clinically available β-lactamase inhibitors on cefiderocol activity against characterized carbapenemase-producing Gram-negative isolates. A collection of 39 well-characterized Gram-negative isolates obtained from various clinical sources and countries were included. Cefiderocol antimicrobial susceptibility was evaluated via reference broth microdilution. The chequerboard microdilution method and time-kill assays were used to determine the synergy of tazobactam, avibactam, vaborbactam and relebactam in combination with cefiderocol. MICs of cefiderocol presented a 4- to 256-fold reduction against Klebsiella pneumoniae carbapenemase (KPC)-producing Gram-negative isolates (predominantly K. pneumoniae) when avibactam, vaborbactam and relebactam were combined individually. Notably, the KPC-inhibitors led to a 4- to 32-fold reduction in cefiderocol MICs in the four cefiderocol-resistant KPC-producing K. pneumoniae isolates, showing restoration of cefiderocol susceptibility (MIC ≤ 2 mg/L) in ten out of twelve cases. Tazobactam led to a 4- to 64-fold decrease in cefiderocol MICs only in K. pneumoniae strains harbouring blaKPC-41, blaKPC-31, blaKPC-53 and blaKPC-66. The synergistic effect of all serine-β-lactamase inhibitors on cefiderocol activity was also shown in OXA-48-like-producing Enterobacterales strains. Conversely, a combination of β-lactamases inhibitors with cefiderocol was not synergistic with all OXA-23-like-producing strains and most metallo-β-lactamases producers. In conclusion, the addition of clinically available serine β-lactamase inhibitors to cefiderocol might represent an important development in the formulation to increase its spectrum and therapeutic efficacy, and to limit in vivo resistance emergence.

Synergistic Action of Cinnamomum verum Essential Oil with Sertraline.

Cinnamomum verum L. essential oil (CEO), commonly known as Ceylon cinnamon or cinnamon tree, is regarded as one of the most employed essential oils in the field of aromatherapy. It is usually applied externally as astringent, antipruritic, rubefacient, and anti-septic agent. Furthermore, both in vitro and in vivo research have demonstrated its numerous pharmacological effects, including the potentiality for treating neuralgia, myalgia, headache, and migraine. Several pieces of research also corroborated its significant antiviral and antimicrobial properties. Cinnamaldehyde, eugenol, caryophyllene, cinnamyl acetate, and cinnamic acid are the most representative compounds that are generally found in greater quantities in CEO and play a pivotal role in determining its pharmacological activities. Due to the global antibiotic resistance scenario and the dwindling amount of funding dedicated to developing new antibiotics, in recent years research has concentrated on exploring specific economic approaches against microbial infections. In this context, the purpose of this study was the investigation of the synergistic antibacterial activities of commercially available and chemically characterized CEO in combination with sertraline, a selective serotonin reuptake inhibitor (SSRI), whose repositioning as a non-antibiotic drug has been explored over the years with encouraging results. In vitro effects of the titled combination were assessed toward a wide panel of both Gram-positive and Gram-negative bacteria. The antimicrobial efficacy was investigated by using the checkerboard microdilution method. The interesting preliminary results obtained suggested a synergistic effect (fractional inhibitory index, FICI &lt; 0.5) of sertraline in combination with CEO, leading to severe growth inhibition for all bacterial species under investigation.

Glyphosate and glyphosate-based herbicides (GBHs) induce phenotypic imipenem resistance in Pseudomonasaeruginosa

GBHs are the most widely used herbicides for weed control worldwide that potentially affect microorganisms, but the role of their sublethal exposure in the development of antibiotic resistance of Pseudomonasaeruginosa is still not fully investigated. Here, the effects of glyphosate acid (GLY), five glyphosate-based herbicides (GBHs), and POE(15), a formerly used co-formulant, on susceptibility to imipenem, a potent carbapenem-type antibiotic, in one clinical and four non-clinical environmental P.aeruginosa isolates were studied. Both pre-exposure in broth culture and co-exposure in solid media of the examined P.aeruginosa strains with 0.5% GBHs resulted in a decreased susceptibility to imipenem, while other carbapenems (doripenem and meropenem) retained their effectiveness. Additionally, the microdilution chequerboard method was used to examine additive/antagonistic/synergistic effects between GLY/POE(15)/GBHs and imipenem by determining the fractional inhibitory concentration (FIC) indexes. Based on the FIC index values, glyphosate acid and Total demonstrated a potent antagonistic effect in all P.aeruginosa strains. Dominator Extra 608 SL and Fozat 480 reduced the activity of imipenem in only one strain (ATCC10145), while POE(15) and three other GBHs did not have any effect on susceptibility to imipenem. Considering the simultaneous presence of GBHs and imipenem in various environmental niches, the detected interactions between these chemicals may affect microbial communities. The mechanisms of the glyphosate and GBH-induced imipenem resistance in P.aeruginosa are yet to be investigated.

S2.4d Combination antifungal effects of eugenol with voriconazole against Candida tropicalis and Candida krusei strains isolated from the genital tract of mares

S2.4 Veterinary mycology research, September 21, 2022, 3:00 PM - 4:30 PM ObjectivesThe objective of the current study was to investigate the possibility that eugenol synergizes the antifungal effects of voriconazole on genital isolates of Candida krusei and Candida tropicalis from mares.MethodsThe antifungal activity of eugenol and voriconazole was evaluated using the broth microdilution assay (CLSI- M27-A3). The synergism of eugenol and voriconazole against genital Candida isolates was evaluated by the microdilution checkerboard method.ResultsMinimum inhibitory concentration (MIC) values for eugenol and voriconazole ranged from 400 to 800 μg/ml and 1 to 8 μg/ml, respectively, for C. tropicalis isolates, and from 200 to 400 μg/ml for eugenol and 2 to 16 μg/ml for voriconazole against C. krusei isolates. Eugenol decreased the arithmetic mean of MIC for voriconazole against C. tropicalis and C. krusei isolates from 2.66 to 0.46 μg/ml and 7.77 to 0.41 μg/ml respectively. The fractional inhibitory concentration index (FICI) values for the eugenol voriconazole combination ranged from 0.25 to 0.88 and 0.19 to 0.63 for C. tropicalis and C. krusei isolates respectively. A synergistic effect of eugenol in combination with voriconazole was observed for 83.3% of C. tropicalis and 77.7% of C. krusei isolates. The antagonistic activity was not seen in any of the isolates tested.ConclusionsEugenol showed fungistatic and fungicidal effects against genital Candida isolates and, in combination, synergized the antifungal effects of voriconazole. The eugenol-voriconazole combination can lay the foundation for a therapeutic approach against isolates in which azole resistance has increased over time.

Vincetoxicum hirundinaria'nın Farklı Kısımlarının Bazı Biyolojik Aktiviteleri

In this study, it was aimed to determine the antimicrobial, antibiofilm activities and synergistic effects of ethanol extracts obtained from above-ground, root, seed and seed coat of Vincetoxicum hirundinaria Medic. Antimicrobial activity was performed against 7 bacterial and 3 fungal standard strains by microdilution method and minimum inhibition concentrations (MIC) were determined. The crystal violet method was applied to determine the prevention of biofilm formation and inhibition of preformed biofilm activities on P. aeruginosa biofilm. In addition, the synergistic effects of aerial part and seed extracts against both E. coli and E. faecalis were examined by the microdilution checkerboard method. According to the antimicrobial test results tested extracts had moderate to low efficacy against studied bacterial and yeast strains. The synergy test showed that the aerial and seed extracts had additive effect against both E. coli and E. faecalis. The extracts also showed the potential to inhibit biofilm formation and inhibit preformed biofilms. Root and seed pod extracts showed the strongest antibiofilm activity, while the aerial part extract was the weakest. In conclusion, our results prove that the tested extracts, especially obtained from root and seed pods and used in the treatment of many diseases, have potential in terms of antibiofilm activity. The literature search indicated that the antimicrobial and antibiofilm activities of V. hirundinaria was evaluated for the first time in the current study, therefore; our findings provide important preliminary data to the literature in terms of antibiofilm activity of V. hirundinaria