Indifferent Effect Research Articles

Multicenter prospective observational study of fungal keratitis in Japan: analyses of in vitro susceptibility tests for combinations of drugs.

To determine the effects of a combination of two antifungal drugs against causative fungi of fungal keratitis in Japan. Multicenter prospective observational study. Eighteen isolates of yeast-like fungi and 22 isolates of filamentous fungi collected by the Multicenter Prospective Observational Study of Fungal Keratitis in Japan were studied. Specially manufactured minimum inhibitory concentration (MIC) measurement plates were used to test the effectiveness of 10 combinations of two antifungal drugs against the isolates. The combinations were pimaricin (PMR) + voriconazole (VRCZ), PMR + fluconazole (FLCZ), PMR + miconazole (MCZ), PMR + micafungin (MCFG), VRCZ + FLCZ, VRCZ + MCZ, VRCZ + MCFG, VRCZ + amphotericin-B (AMPH-B), MCZ + FLCZ, and MCZ + MCFG. The checkerboard microdilution method was used, and the fractional inhibitory concentration (FIC) index was calculated based on the guidelines of The Clinical & Laboratory Standards Institute (CLSI). In yeast-like fungi,additive effects were observed between PMR and MCFG in 77.8% of the isolates, and they were also observed between the azoles. Synergistic effects were observed on 11.1% of the isolates for MCZ and FLCZ. On the other hand, antagonistic effects were present between PMR and azoles with 88.9% between PMR and VRCZ, 72.2% between PMR and FLCZ, and 94.4% between PMR and MCZ. In filamentous fungi,additive effects were observed between PMR and MCFG in 40.9% of the isolates, and between VRCZ and MCZ in 40.9% of the isolates. Antagonistic effects were observed for PMR and the azoles. The combination of drugs prescribed for fungal keratitis incurs a possibility of synergistic, additive, indifferent, or antagonistic effects, depending on drug combinations and fungal strains.

Gentamicin-Induced Antibacterial Activity is Attenuated by L-Captopril

Objective: Captopril is a long-acting human angiotensin-converting enzyme (ACE) inhibitor that has been used to treat hypertension and heart failure for many years. In this study, it was aimed to determine the antimicrobial activity of L-captopril on Methicillin-Resistant Staphylococcus aureus MRSA and Methicillin-Sensitive Staphylococcus aureus (MSSA) clinical isolates and, demonstrate the combination activity of captopril with ciprofloxacin (CXP) and gentamicin (GEN), which are among antistaphylococcal chemical agents. Method: The minimum inhibitor concentration (MIC) and minimum bactericidal concentration (MBC) were determined using the microdilution technique in 96-well microtiter plates. The activity of L-captopril and CXP or GEN combination against MRSA and MSSA clinical isolates was determined by the micro-broth checkerboard assay method. Results: The captopril MIC value was determined to be 2.5 mg/ml in all bacteria strains tested. The captopril/CXP combination had an indifferent effect in the other strains tested except for one MRSA isolate. The captopril/GEN combination had an antagonistic effect in all strains studied and increased the MIC 2-4 fold. The captopril/GEN combination was found to reduce the bactericidal activity of gentamicin. Conclusions: The study results suggest that exposure to these drugs may lead to multidrug resistance in S. aureus bacteria. Especially in the hypertensive patient group, the induced resistance in any S. aureus infections should be taken into consideration while using captopril and, it should be considered that this antagonism may cause an increase in S. aureus infections.

In vitro Evaluation of Synergistic Inhibitory Effect of Propolis and Chlorhexidine on Biofilm Cells of Oral Pathogens

Background: Dental plaque is a microbial biofilm that can source various oral diseases. Objectives: This study aimed to investigate the synergistic anti-biofilm effect of chlorhexidine (CHX) and propolis aqueous extract on Streptococcus mutans and Staphylococcus aureus. Methods: This study, performed in October 2021, determined the minimum biofilm inhibitory concentration (MBIC) and minimum biofilm eradication concentration (MBEC) of CHX 0.2% and propolis aqueous extract against S. mutans and S. aureus prepared from the cell bank of the Pasteur Institute of Iran. Also, the in vitro synergistic effect of these two compounds was evaluated on the biofilm inhibition and eradication of the studied bacteria using the checkerboard method. Results: The MBIC of chlorhexidine was 125 μg/mL for both S. aureus and S. mutans, while the MBIC of propolis aqueous extract was 937.5 and 3750 μg/mL, respectively. Also, the MBEC of chlorhexidine was 250 and ≥ 500 μg/mL for S. aureus and S. mutans, while the MBEC of propolis was 1875 and ≥ 7500 μg/mL, respectively. Combining the two substances showed a synergistic effect in inhibiting biofilm formation (P ≤ 0.05) while they had an indifferent effect on biofilm eradication. Conclusions: This study concluded that although CHX is one of the most common and vital oral disinfectants in oral diseases, propolis may also be used, as an alternative mouthwash with reliable natural antimicrobial properties, alone or combined with CHX as a moderator of its side effects.

Serotonin reuptake inhibitors effect on fluconazole activity against resistant Candida glabrata strains

The study aimed to evaluate four selective serotonin reuptake inhibitors (SSRIs) as modifiers of fluconazole activity against resistant strains of Candida glabrata. The effect of SSRIs on fluconazole activity was studied using the checkerboard method against C. glabrata strains (CBS 138, CBS 850821, DSY 562, DSY 565, ATCC 22553 and ATCC 90030); fractional inhibitory concentration index (FIC) was calculated and time-kill curve was used for the most prominent combination for further evaluation. All used SSRIs have antifungal activity against the C. glabrata strains tested. A combination of fluconazole with fluoxetine or fluvoxamine showed indifferent effects (fractional inhibitory concentration index [FICI] in all strains >1 but <4), whereas a paroxetine-fluconazole combination showed an additive effect against DSY565 and CBS138, known to express efflux pumps as well as on ATCC strain (0.5 < FIC < 1) with indifferent effect on other strains used. The most promising combination was that of fluconazole with sertraline (FICI ≤0.5), where a synergistic effect was observed against all resistant and susceptible dose-dependent strains, including those known to express efflux pumps. This synergistic effect was confirmed by time-kill curve assay against all resistant C. glabrata and ATCC strains with a >2-log10 CFU/mL reduction caused by combination compared with a single active agent of fluconazole after 24 hours of incubation. A sertraline-fluconazole combination produced an additive effect on the reference ATCC strain. Our data suggest that blocking active efflux pumps by sertraline may be considered the probable mechanism of synergism with fluconazole. The combination of sertraline with fluconazole could be a promising remedy for treatment of infections caused by resistant C. glabrata.

Determination of an effective agent combination using nisin against Salmonella biofilm.

This present study aims to determine the lowest concentration effects of the assayed different antibiotics; antimicrobial agents alone and their combinations with nisin were investigated to prevent the biofilm formation and break down the biofilm structure of Salmonella. While the combination of nisin and EDTA showed a synergistic effect against Salmonella strain, chlorhexidine digluconate and streptomycin with nisin showed a partial synergetic effect; citric acid and sulfonamides with nisin showed an indifferent effect. The use of citric acid and chlorhexidine digluconate alone was very effective in Salmonella inhibition. While the citric acid combined with other agents had not much effect, the use of chlorhexidine digluconate combined with nisin and EDTA inactivated the total initial count within 24h. Significantly, when citric acid and sulfonamides are used alone, they reduce by 64% and 44%, respectively. When they used nisin + EDTA, this ratio increased to 83% and 84%, respectively. For the prevention of biofilm, the most suitable conditions were determined as 97% biofilm inhibition. The results of this study can be used as a guide for the emergence of new approaches to ensure the food safety and quality of the food industry.

Chemical Composition and Synergistic Potential of Mentha pulegium L. and Artemisia herba alba Asso. Essential Oils and Antibiotic against Multi-Drug Resistant Bacteria.

The essential oils were obtained by hydrodistillation from aerial parts of Mentha pulegium L. (M. pulegium L.) and Artemisia herba alba (A. herba alba) Asso. and analyzed by gas chromatography–flame ionization detector chromatograpy (GC–FID) and gaz chromatography–mass spectrometry (GC–MS). The antibacterial activities of the oils were determined by the disk diffusion method and a microdilution broth assay against six bacteria stains. The combinations of these essential oils with antibiotics were evaluated against two multi-drug-resistant bacteria strains: imipenem-resistant Acinetobacter baumannii (IRAB S3310) and methicillin-resistant Staphylococcus aureus (MRSA S19). The chemical analysis of M. pulegium essential oil revealed the presence of pulegone (74.8%) and neoisomenthol (10.0%). A. herba alba essential oil was characterized by camphor (32.0%), α-thujone (13.7%), 1,8-cineole (9.8%), β-thujone (5.0%), bornéol (3.8%), camphene (3.6%), and p-cymene (2.1%). All strains tested except Pseudomonas aeruginosa were susceptible to these oils. The combinations of essential oils with antibiotics exerted synergism, antagonism, or indifferent effects. The best effect was observed with A. herba alba essential oil in association with cefoxitin (CX) against MRSA S19. However, for IRAB S3310, the strongest synergistic effect was observed with M. pulegium in association with amikacin (AK). This study demonstrated that M. pulegium and A. herba alba essential oils have antibacterial activities which could be potentiated by antibiotics especially in the case of IRAB S3310.

In silico and in vitro analysis of a new potential antifungal substance, 2-Bromo-Nphenylacetamide, against invasive candidiasis isolates

Introduction: invasive candidiasis is related to high rates of morbidity and mortality. There are few classes of drugs available for the treatment of this type of infection and the index of resistant strains is increasing. Such circumstances highlight that the search for new pharmacotherapeutic alternatives is increasingly necessary. This study investigated 2-Bromo-N-phenylacetamide, a substance whose antifungal activity has not yet been reported. Objective: to evaluate its activity against invasive candidiasis isolates, by determining the minimum inhibitory and fungicide concentrations. Meth-odology: molecular docking was performed to investigate the possible mechanism of action of the substance. The substance was also associated with fluconazole, to assess the viability of the combination in clinical practice. The minimum inhibitory concen-trations ranged between 4 to 32 μg/mL, and it acts in a fungicidal way. Results: molec-ular docking suggests that 2-Bromo-N-phenylacetamide possibly acts on the fungal plasma membrane. And the association of 2-Bromo-N-phenylacetamide with flucon-azole against resistant strains showed an indifferent effect. Conclusion: further studies should be carried out to elucidate the potential of this substance, which may become a future drug candidate to treat invasive candidiasis and other fungal infections.

Methicillin-Resistant &amp;lt;i&amp;gt;Staphylococcus aureus&amp;lt;/i&amp;gt; May Also Be Resistant to Clindamycin and Vancomycin

Objectives: Methicillin-resistant Staphylococcus aureus (MRSA) is a global superbug widely distributed in hospitals, communities and livestock settings. This study investigated the presence and molecular characterization of MRSA co-resistance to clindamycin and vancomycin in the southeastern region of Nigeria. The susceptibility of these organisms to other selected antibiotics was also investigated. Method: Biological samples were obtained from consenting patients from three establishments in Enugu, Nigeria and cultured for isolation and purification. The pure isolates were subjected to antimicrobial susceptibility profiling using conventional antibiotics. The genomic DNAs of the pure isolates were isolated using the Promega genomic DNA purification kit while the antibiotic resistance genes (mecA) genes were identified using a multiplex polymerase chain reaction. Also, the minimum inhibitory concentration of the clindamycin and vancomycin antibiotics was determined as well as their combined activity on the MRSA isolates. Results: A large proportion (71%) of the MRSA isolates was from urine samples and then from the High Vaginal Swab (19%). All the isolates were resistant to cloxacillin while 95% were resistant to ciprofloxacin. MRSA isolates demonstrated resistance to clindamycin (with MIC of 23.44 - 250 μg/ml) and to vancomycin (with MIC of 62.5 - 250 μg/ml). The isolated MRSA also demonstrated multidrug-resistant traits. The combined effects of vancomycin and clindamycin against different species of MRSA exhibited additive, antagonistic and indifferent effects and none had a synergistic effect. Multiplex Polymerase Chain Reaction revealed that the majority of the strains were positive for the 162-bp internal fragment of the mecA gene of MRSA and basically displayed SCCmec type III, indicating that they were multidrug-resistant and hospital-acquired. Conclusion: Clindamycin and vancomycin-resistant MRSA infections are also within the Eastern region of Nigeria as found in other countries of the world. This superbug, therefore, may require drastic and urgent measures to curtail its spread and attendant healthcare challenges like outbreaks of infections. In addition, strict adherence to antibiotic policy and continuous surveillance is highly advocated.

Evaluation of Host Defense Peptide (CaD23)-Antibiotic Interaction and Mechanism of Action: Insights From Experimental and Molecular Dynamics Simulations Studies.

Background/Aim: Host defense peptides (HDPs) have the potential to provide a novel solution to antimicrobial resistance (AMR) in view of their unique and broad-spectrum antimicrobial activities. We had recently developed a novel hybrid HDP based on LL-37 and human beta-defensin-2, named CaD23, which was shown to exhibit good in vivo antimicrobial efficacy against Staphylococcus aureus in a bacterial keratitis murine model. This study aimed to examine the potential CaD23-antibiotic synergism and the secondary structure and underlying mechanism of action of CaD23. Methods: Peptide-antibiotic interaction was evaluated against S. aureus, methicillin-resistant S. aureus (MRSA), and Pseudomonas aeruginosa using established checkerboard and time-kill assays. Fractional inhibitory concentration index (FICI) was calculated and interpreted as synergistic (FIC<0.5), additive (FIC between 0.5–1.0), indifferent (FIC between >1.0 and ≤4), or antagonistic (FIC>4). SYTOX green uptake assay was performed to determine the membrane-permeabilising action of CaD23. Molecular dynamics (MD) simulations were performed to evaluate the interaction of CaD23 with bacterial and mammalian mimetic membranes. Circular dichroism (CD) spectroscopy was also performed to examine the secondary structures of CaD23. Results: CaD23-amikacin and CaD23-levofloxacin combination treatment exhibited a strong additive effect against S. aureus SH1000 (FICI = 0.60–0.69) and MRSA43300 (FICI = 0.56–0.60) but an indifferent effect against P. aeruginosa (FIC = 1.03–1.15). CaD23 (at 25 μg/ml; 2xMIC) completely killed S. aureus within 30 min. When used at sub-MIC concentration (3.1 μg/ml; 0.25xMIC), it was able to expedite the antimicrobial action of amikacin against S. aureus by 50%. The rapid antimicrobial action of CaD23 was attributed to the underlying membrane-permeabilising mechanism of action, evidenced by the SYTOX green uptake assay and MD simulations studies. MD simulations revealed that cationicity, alpha-helicity, amphiphilicity and hydrophobicity (related to the Trp residue at C-terminal) play important roles in the antimicrobial action of CaD23. The secondary structures of CaD23 observed in MD simulations were validated by CD spectroscopy. Conclusion: CaD23 is a novel alpha-helical, membrane-active synthetic HDP that can enhance and expedite the antimicrobial action of antibiotics against Gram-positive bacteria when used in combination. MD simulations serves as a powerful tool in revealing the peptide secondary structure, dissecting the mechanism of action, and guiding the design and optimisation of HDPs.

Synergistic effect of vancomycin combined with cefotaxime, imipenem, or meropenem against Staphylococcus aureus with reduced susceptibility to vancomycin

Background/aim We investigated the synergistic effect between vancomycin and β-lactams against vancomycin-susceptible (VSSA) and nonsusceptible MRSA isolates [heterogeneous vancomycin-intermediate S. aureus (hVISA) and VISA].Materials and methods A total of 29 MRSA, including 6 VISA, 14 hVISA, and 9 VSSA isolates, were subjected to a microbroth dilution-minimum inhibitory concentration (MIC) checkerboard using vancomycin combined with cefotaxime, imipenem, or meropenem. To confirm synergistic activity, the representative strains of VISA, hVISA, and VSSA were then selected for the time-kill curve method.Results The combination of vancomycin with imipenem, meropenem, or cefotaxime exhibited synergistic effects against 17 (2 VISA, 9 hVISA, and 6 VSSA), 14 (3 VISA, 9 hVISA and 2 VSSA), and 5 (3 VISA and 2 hVISA) isolates, respectively. Additive and indifferent effects were found in the remaining isolates, but no antagonistic effect was observed. Using time-kill assay, the vancomycin combined with either imipenem or cefotaxime demonstrated synergism against both VISA and hVISA isolates, while the synergistic effect with meropenem was obtained only in the VISA isolates.Conclusion This study demonstrated in vitro enhanced antibacterial activity of vancomycin plus β-lactams against clinical hVISA or VISA isolates. These combinations may be an alternative treatment for MRSA infections in clinical practice.

Chemical composition, antifungal activity, antifungal mechanism and interaction manner of the fatty acid of Prunus mahaleb L. with fluconazole

ABSTRACT The interaction manner of Prunus mahaleb L. (P. mahaleb) seed oil (MSO) and fluconazole (FLC: antimycotic) combinations (MSO*FLC) against C. albicans and C. parapsilosis were evaluated using the microdilution technique. The most representative compounds of mahaleb oil were found to be conjugated linolenic acid (34.39%), oleic acid (31.76%), and linoleic acid (25.54%) by GC-MS. In antimicrobial activity study, P. mahaleb had an inhibition zone (IZ) of C. albicans and C. parapsilosis with 6.89 mm and 11.39 mm and a minimum inhibitory concentration (MIC) with 35.3 µgmL−1 and 23.9 µgmL−1, respectively. The strongest indifferent effect was observed as 57.14% for C. albicans and 100% for C. parapsilosis in fluconazole-mahaleb oil combinations. An increase in DNA and protein leakage was observed when yeast was exposed to the FA. The destruction on the cell surface was visualized using Scanning Electron Microscopy (SEM) analysis.

Evaluation of synergistic anticandidal activity of Galenia africana extract and fluconazole against Candida albicans and Candida glabrata

Candida infections have increased in recent years and are causing a serious public health concern. In addition, Candida species are becoming resistant to numerous antifungal drugs. It is for this reason that alternative treatment options are being explored, using medicinal plants, to curb this trend of drug resistance. The aim of this study was to evaluate the anticandidal activity of an ethanolic extract of Galenia africana (G. africana) alone and particularly in combination with fluconazole (FLC) against Candida albicans and Candida glabrata. The anticandidal activity was evaluated using various techniques. The minimum inhibitory concentration (MIC) of the extract was 6.25 mg/mL for both Candida strains. A synergistic effect was observed when G. africana was combined with FLC, against C. albicans and an indifferent effect against C. glabrata when interpreted by the fractional inhibitory concentration index (FICI) (0.36 and 1.002 for C. albicans and C. glabrata, respectively). Microscopic analysis revealed cell damage and decrease in cell size after G. africana treatment. Collectively, these results suggest that G. africana possessed antifungal activity against the Candida strains and a synergistic effect with FLC. Cell damage observed possibly contributed to this synergistic effect. This provides new information for the development of new antifungal agents.

Synergistic Antimicrobial Activities of Combinations of Vanillin and Essential Oils of Cinnamon Bark, Cinnamon Leaves, and Cloves.

Plant bioactive compounds have antimicrobial and antioxidant activities that allow them to be used as a substitute for synthetic chemical additives in both food and food packaging. To improve its sensory and bactericidal effects, its use in the form of effective combinations has emerged as an interesting possibility in the food industry. In this study, the antimicrobial activities of essential oils (EOs) of cinnamon bark, cinnamon leaves, and clove and the pure compounds vanillin, eugenol, and cinnamaldehyde were investigated individually and in combination against Listeria monocytogenes and Escherichia coli O157:H7. The possible interactions of combinations of pure compounds and EOs were performed by the two-dimensional checkerboard assay and isobologram methods. Vanillin exhibited the lowest antimicrobial activity (MIC of 3002 ppm against L. monocytogenes and 2795 ppm against E. coli O157:H7), while clove and cinnamon bark EOs exhibited the highest antimicrobial activity (402–404 against L. monocytogenes and 778–721 against E. coli O157:H7). For L. monocytogenes, pure compound eugenol, the main component of cinnamon leaves and clove, showed lower antimicrobial activity than EOs, which was attributed to the influence of the minor components of the EOs. The same was observed with cinnamaldehyde, the main component of cinnamon bark EO. The combinations of vanillin/clove EO and vanillin/cinnamon bark EO showed the most synergistic antimicrobial effect. The combination of the EOs of cinnamon bark/clove and cinnamon bark/cinnamon leaves showed additive effect against L. monocytogenes but indifferent effect against E. coli O157:H7. For L. monocytogenes, the best inhibitory effects were achieved by cinnamon bark EO (85 ppm)/vanillin (910 ppm) and clove EO (121 ppm)/vanillin (691 ppm) combinations. For E. coli, the inhibitory effects of clove EO (104 ppm)/vanillin (1006 ppm) and cinnamon leaves EO (118 ppm)/vanillin (979 ppm) combinations were noteworthy. Some of the tested combinations increased the antimicrobial effect and would allow the effective doses to be reduced, thereby offering possible new applications for food and active food packaging.

Antimicrobial synergism and antibiofilm activities of Pelargonium graveolens, Rosemary officinalis, and Mentha piperita essential oils against extreme drug-resistant Acinetobacter baumannii clinical isolates.

Rosemary officinalis L., Pelargonium graveolens L., and Mentha piperita L., essential oils are used by complementary medicine specialists simultaneously with traditional antibiotics for treatment purposes. The chemical composition of essential oils was analyzed by the gas chromatography-mass spectrometry method. In vitro antibacterial and antibiofilm activities of the essential oils were tested against extreme drug-resistant (XDR) colistin-resistant and colistin susceptible Acinetobacter baumannii clinical strains. The synergistic activities between essential oils and colistin antibiotics were investigated by the checkerboard method. The highest antibacterial effect was detected in mint essential oil (2.5-5μl/ml), followed by pelargonium essential oil (5-20μl/ml) and rosemary essential oil (5-20μl/ml). The combination of rosemary essential oil or pelargonium essential oil with colistin showed strong synergistic activity in most of the bacterial strains tested (fractional inhibitory concentration index≤0.5; synergy). As a result of the combination of mint essential oil and colistin, an indifferent effect was observed in only two bacterial strains, and other strains could not be evaluated. No antagonistic effects were observed in any of the tested essential oils. As a result of the effectiveness of the combination, the minimum inhibitory concentration (MIC) values of colistin in XDR-A.baumannii clinical isolates decreased 2-32 fold. Additionally, the sub-MIC concentration of essential oils exhibited an inhibitory effect (48-90%) against the biofilm layer of tested A.baumannii strains.

Evaluation of the Single and Combined Antibacterial Efficiency of the Leaf Essential Oils of Four Common Culinary herbs: Dill, Celery, Coriander and Fennel Grown in Jordan

This study aimed to determine the volatile oil composition of the fresh leaves of dill, celery, coriander, and fennel, grown in Jordan, and to evaluate the antimicrobial activity of the essential oils individually and in binary (50:50) and quaternary (25:25:25:25) mixtures. The volatile oils, obtained by hydrodistillation, were analysed by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). In vitro antimicrobial activities of the essential oils were evaluated against Escherichia coli, Bacillus subtilis, Pseudomonas aeruginosa, and Candida albicans. Using the microtiter plate dilution method minimal inhibitory concentrations (MICs) of the essential oils were determined and the checkerboard method was then used to investigate the antimicrobial efficacy of the essential oil combinations by means of the fractional inhibitory concentration index (FICI). Dill and celery samples were rich in monoterpene hydrocarbons, while the oils of fennel and coriander were dominated by non-terpenoid compounds. In antimicrobial experiments, the essential oil of dill yielded the best activity against all tested microorganisms. In binary combination of the essential oils, best results were detected in a combination of dill with coriander against B. subtilis and P. aeruginosa, and fennel with coriander against E. coli and P. aeruginosa. The present study is the first evaluation of the volatile oil composition and determination of the antimicrobial activities of the fresh leaves of dill, celery, coriander, and fennel in binary and quaternary combinations. Additive or synergistic effects were detected in certain binary oil combinations while indifferent and antagonist effects were noticed in other mixtures.

In vitro and in vivo Antibacterial Activity of Linezolid Plus Fosfomycin Against Staphylococcus aureus with Resistance to One Drug.

ObjectiveThe purpose of this study is to assess the in vitro/vivo activities of linezolid plus fosfomycin against Staphylococcus aureus (S. aureus) isolates with varying susceptibility to the study drugs.MethodsThe increasing concentration stepwise method was used to induce S. aureus resistant strains. The in vitro antibacterial activity of linezolid combined with fosfomycin against S. aureus in vitro was studied by time-kill curve and PAE. The transmission electron microscopy (TEM) was employed to observe the cell morphology of bacteria treated with drug, and the changes of cell wall thickness were recorded. The Galleria mellonella infection model was established to demonstrate the in vivo efficacy of linezolid and fosfomycin against S. aureus with varying susceptibility.ResultsThe antibiotic combination showed excellent synergistic or additive effects on the original and the linezolid-resistant strain, but showed indifferent effect for fosfomycin-resistant strain. TEM images showed that fosfomycin alone and in combined could reduce the cell wall thickness of the strains resistant to linezolid and cell lysis, while linezolid increases the cell wall thickness of the strains resistant to fosfomycin. In the Galleria mellonella infection model, the survival rate of the antibiotic combined was improved compared with that of the single drug. There was a good correlation between in vivo efficacy and in vitro susceptibility.ConclusionThe type of interaction expressed in the test combination was highly dependent on fosfomycin resistance.

UHPLC-ESI-OT-MS Phenolics Profiling, Free Radical Scavenging, Antibacterial and Nematicidal Activities of "Yellow-Brown Resins" from Larrea spp.

This research was designed to investigate the metabolite profiling, phenolics and flavonoids content and the potential antioxidant, antibacterial and nematicidal activities of “yellow-brown resins” from Larrea divaricata Cav (LdRe) and L. nitida Cav (LnRe). Metabolite profiling was obtained using an ultrahigh resolution liquid chromatography orbitrap MS analysis (UHPLC-ESI-OT-MS). The antioxidant properties were screened by four methods: 2,2-diphenyl-1-picrylhydrazyl assay (DPPH), trolox equivalent antioxidant activity assay (TEAC), ferric-reducing antioxidant power assay (FRAP) and lipid peroxidation in erythrocytes (LP). The antibacterial activity was evaluated according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. In addition, the potential combinatory effect was analyzed with the fractional inhibitory concentration index (FICI) values using the checkerboard design. The nematicidal activity was carried out according to a standardized protocol. LdRe and LnRe showed a strong capture of the DPPH radical withvalues around 8.4 µg resin/mL; FRAP (1.69–1.94 mgTE/ g resin), TEAC (1.08–1.09 mgTE/g resin) and LP (81–82% at 100 µg of resin/mL) assays. A strong antimicrobial activity was displayed by both resins against methicillin-sensitive Staphylococcus aureus ATCC 25923(MSSA) and methicillin-resistant S. aureus ATCC 43300(MRSA) (MICs = 16–32 µg resin/mL). Additionally, the combination of LdRe or LnRe with the antibiotic cefotaxime showed an indifferent effect (FICI values = 1–1.25), however, this combinationcould be a potential strategy to reduce the drug doses, and in this way can be a potential alternative to reduce bacterial resistance. On the other hand, the resins showed a scarce nematicidal potential toward J2 Meloidogyne incognita; an important nematode infecting horticultural crops. Phenolics compounds were identified by UHPLC-PDA-OT-MS analysis, updating the knowledge on the chemical profile of these species. These results, together with the high content of quantified phenolics and flavonoids, allow the phenolics-enriched resins of these two Larrea species to be considered as a promising sustainable source of compounds of pharmacological interest.

Effective antimicrobial combination in vivo treatment predicted with microcalorimetry screening.

ObjectivesThe worldwide emergence of antibiotic resistance calls for effective exploitation of existing antibiotics. Antibiotic combinations with different modes of action can synergize for successful treatment. In the present study, we used microcalorimetry screening to identify synergistic combination treatments against clinical MDR isolates. The synergistic effects were validated in a murine infection model.MethodsThe synergy of meropenem combined with colistin, rifampicin or amikacin was tested on 12 isolates (1 Escherichia coli, 5 Klebsiella pneumoniae, 3 Pseudomonas aeruginosa and 3 Acinetobacter baumannii) in an isothermal microcalorimeter measuring metabolic activity. One A. baumannii strain was tested with two individual pairings of antibiotic combinations. The microcalorimetric data were used to predict in vivo efficacy in a murine peritonitis/sepsis model. NMRI mice were inoculated intraperitoneally and after 1 h treated with saline, drug X, drug Y or X+Y. Bacterial load was determined by cfu in peritoneal fluid and blood after 4 h.Results In vitro, of the 13 combinations tested on the 12 strains, 3 of them exhibited a synergistic reduction in MIC (23% n = 3/13), 5 showed an additive effect (38.5% n = 5/13) and 5 had indifferent or antagonistic effects (38.5% n = 5/13). There was a significant correlation (P = 0.024) between microcalorimetry-screening FIC index values and the log reduction in peritoneal fluid from mice that underwent combination treatment compared with the most effective mono treatment. No such correlation could be found between chequerboard and in vivo results (P = 0.16).ConclusionsThese data support microcalorimetic metabolic readout to predict additive or synergistic effects of combination treatment of MDR infections within hours.

Differential expression of blaCTX-M-33 with vancomycin/trimethoprim combination in Escherichia coli-producing extended-spectrum β-lactamase isolated from intensive care unit-acquired urinary tract infection

Aim: The aim of this study was to develop combination approach for the treatment of Escherichia coli-producing extended-spectrum β-lactamases (ESBLs) isolated from intensive care unit (ICU)-acquired urinary tract infections (UTIs). Materials and Methods: The observational study was conducted between January 5, 2018- June 5, 2019 to isolate and detect E. coli from UTI patients admitted to ICUs in Shahid Rajaee hospital, Gachsaran, Iran. Morphological, biochemical and molecular methods were conducted to identify E. coli isolates. Phenotypic confirmation of E. coli producing ESBL was performed using ESBL disc diffusion test according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. The combination index assay set-up was based on CLSI guidelines to investigate antibacterial susceptibilities to vancomycin alone and in combination with trimethoprim and interpreted with the fractional inhibitory concentration (FIC) index. Eventually, the expression levels of blaCTX-M-33 gene were determined by real-time quantitative reverse transcription–polymerase chain reaction. Results: A total of 90 ICU-acquired UTIs occurred among 255 patients. The combination index assay results showed that vancomycin/trimethoprim combination would be reduced its minimal inhibitory concentration90 value. The vancomycin/trimethoprim combination revealed partial synergistic and indifferent effects (FIC = 0.52–1.50) in the isolates of E. coli-producing ESBL. The results of gene expression analysis indicated that vancomycin/trimethoprim combination caused downregulation of blaCTX-M-33 gene at negligible levels by 55.56–58.82-fold and stopped drug resistant. Conclusion: Vancomycin/trimethoprim combination may diminish resistance in the E. coli-producing ESBL isolated from ICU-acquired UTI patients.

Synergic effect of eugenol oleate with amphotericin B augments anti-leishmanial immune response in experimental visceral leishmaniasis in vitro and in vivo

Present treatment regimen on visceral leishmaniasis has multiple limitations including severe side effects, toxicity, and resistance of Leishmania strains. Amphotericin B is a well-established pharmacologically approved drug; however, mainly toxicity is a foremost issue with that drug. Recently, our group identified eugenol oleate as an anti-leishmanial immunomodulatory compound. The important objectives of this present study was to evaluate the possible synergistic effect of eugenol oleate with amphotericin B to reduce the toxicity of this approved drug.Results obtained from this study signified that combination of eugenol oleate and amphotericin B showed indifferent combinatorial effect against promastigotes with xΣFIC 1.015, while, moderate synergistic activity with xΣFIC 0.456 against amastigotes. It was also notable that eugenol oleate (2.5 μM) with low concentrations of amphotericin B (0.3125 μM) showed 96.45% parasite reduction within L. donovani-infected murine macrophages. Furthermore, eugenol oleate and amphotericin B significantly (p < 0.01) enhanced the nitrite generation, and pro-inflammatory cytokines (IL-12, IFN-γ and TNF-α) in infected macrophages in vitro and in BALB/c mice in vivo. Eugenol oleate (10 mg/Kg b. wt.) with amphotericin B (1 mg/Kg b.wt.) significantly (p < 0.01) controlled the parasite burden in liver by 96.2% and in spleen by 93.12%. Hence, this study strongly suggested the synergic potential of eugenol oleate with low concentration of amphotericin B in experimental visceral leishmaniasis through anti-leishmanial immune response.