Antifungal Activity Of Fluconazole Research Articles

Design and Antifungal Activity of Fluconazole and Nystatin Loaded Onto Silica Mesoporous

Design and Antifungal Activity of Fluconazole and Nystatin Loaded Onto Silica Mesoporous

COVID-19-associated pulmonary aspergillosis (CAPA) in Iranian patients admitted with severe COVID-19 pneumonia.

PurposeBacterial or virus co-infections with SARS-CoV-2 have been reported in many studies; however, the knowledge on Aspergillus co-infection among patients with COVID-19 was limited. This study was conducted to identify and isolate fungal agents and to evaluate the prevalence of pulmonary aspergillosis (CAPA) as well as antifungal susceptibility patterns of Aspergillus species in patients with COVID-19 admitted to Shahid Beheshti Hospital, Kashan, Iran.MethodsThe study involved 119 patients with severe COVID-19 pneumonia referred to the Shahid Beheshti Hospital, Kashan, Iran. A total of 17 Aspergillus spp. that were isolated from COVID-19 patients suspected of CAPA were enrolled in the study. CAPA was defined using ECMM/ISHAM consensus criteria. The PCR amplification of the β-tubulin gene was used to identify the species. The antifungal activities of fluconazole, itraconazole, voriconazole, amphotericin B against Aspergillus spp. were evaluated according to the Clinical and Laboratory Standards Institute manual (M38-A3).ResultsFrom the 119 patients with severe COVID-19 pneumonia, CAPA was confirmed in 17 cases (14.3%). Of these, 12 (70.6%) were males and 5 (29.4%) were females; the mean age at presentation was 73.8 years (range: 45–88 years; median = 77; IQR = 18). Aspergillus fumigatus (9/17; 52.9%), Aspergillus flavus (5/17; 29.4%), Aspergillus oryzae (3/17, 17.6%), were identified as etiologic agents of CAPA, using the molecular techniques. Voriconazole and amphotericin B showed more activity against all isolates. Moreover, the MIC of fluconazole, itraconazole varied with the tested isolates. For 3 clinical isolates of A. fumigatus, 2 isolate of A. flavus and 3 A. oryzae, the MIC of fluconazole and itraconazole were ≥ 16 µg/mL.ConclusionsWe observed a high incidence (14.3%) of probable aspergillosis in 119 patients with COVID-19, which might indicate the risk for developing IPA in COVID-19 patients. When comparing patients with and without CAPA regarding baseline characteristics, CAPA patients were older (p =0 .024), had received more frequent systemic corticosteroids (p = 0.024), and had a higher mortality rate (p = 0.018). The outcome of CAPA is usually poor, thus emphasis shall be given to screening and/or prophylaxis in COVID-19 patients with any risk of developing CAPA.

Antibiofilm Activity of Essential Fatty Acids Against Candida albicans from Vulvovaginal Candidiasis and Bloodstream Infections.

PurposeThe biofilm formation of Candida albicans is an important virulence factor as it can increase tolerance to conventional antifungal drugs and the host immune system. The study aimed to assess the effect of essential fatty acids (EFAs) against biofilm formation and mature biofilms of C. albicans strains, which were isolated from vulvovaginal candidiasis and candidemia.MethodsThe biofilm formation ability of C. albicans and antifungal activities of fluconazole were determined. Additionally, the effects of six EFAs [α-linolenic acid (ALA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linoleic acid (LOA), γ-linolenic acid (GLA), and arachidonic acid (AA)] against C. albicans under planktonic and biofilm conditions were evaluated.Results94.1% of C. albicans exhibited biofilm formation capacity, and 98.5% of C. albicans were susceptible to fluconazole. The biofilms of C. albicans were highly resistant to fluconazole with minimum biofilm eradication concentration values ≥ 64 µg/mL. The EFAs attenuated biofilm formation in a dose-dependent manner, and GLA displayed a remarkable inhibitory activity against biofilm formation of C. albicans. In addition, EPA, DHA, and GLA at 0.1 mM could inhibit the biofilm formation of C. albicans without affecting the planktonic growth rate. Notably, EPA and AA at 1 mM had both inhibitory and eradication activities on C. albicans biofilms.ConclusionThis is the first study to directly compare different EFAs for their capacity to affect C. albicans biofilm formation as well as biofilm eradication. These results suggest EPA and AA could serve as potential new antifungal agents for the treatment of clinical infections caused by C. albicans biofilms.

Clinical impact of Candida respiratory tract colonization and acute lung infections in critically ill patients with COVID-19 pneumonia

Coronavirus disease 2019 (COVID-19), which is attributable to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has been causing a worldwide health issue. Airways colonization by Candida spp. is prevalent among patients on automatic ventilation in intensive care units (ICUs). This research aimed to ascertain the risk factors and roles of Candida spp. respiratory tract colonization, and Candida lung infection during the progression of COVID-19 pneumonia in critically ill patients. In total, Candida spp. were recovered in 69 from 100 immunosuppressed patients with COVID-19. Bronchoscopy was used to collect the Bronchoalveolar lavage (BAL) specimens. For the identification of Candida spp. PCR sequencing was done using the ITS1 and ITS4 primers. The amplification of the HWP1 gene was conducted to identify the Candida albicans complex. The antifungal activities of fluconazole, itraconazole, voriconazole, amphotericin B and caspofungin against Candida spp. were evaluated using the Clinical and Laboratory Standards Institute M60. In 63.77% of the patients, Candida respiratory colonization at D0 and D14 had no impact on the severity of COVID-19. In comparison to C. albicans strains, Candida respiratory disorder with C. glabrata had influenced the severity of COVID-19 for critically ill patients following adjustment for the risk factors of COVID-19 (P < 0.05). Amphotericin B and caspofungin showed superior activity against all Candida spp. All antifungal agents showed 100% sensitivity against the two C. africana strains. Our observation on patients who used automatic ventilation, respiratory colonization by Candida spp. was not seen to influence the infection or death caused by COVID-19. Amphotericin B and caspofungin showed superior activity against all Candida spp. and were recommended for the treatment regime of pulmonary candidiasis associated with COVID-19 infection. Although “Candida pneumonia” is rarely being reported in critically ill patients, Candida airway colonization mainly by Candida albicans is common especially among patients with diabetes, malignancies, and kidney disorders.

Evaluation of Antimicrobial and Antioxidant Activity of Newly Synthesized NMannich Bases from Methyl Salicylate

Isatin reacted with formaldehyde and a number of secondary amines in order to produce the N-Mannich bases of isatin derivatives. FT-IR, 1H-NMR, and mass spectrum data were used to demonstrate that their chemical structures were accurate. An agar diffusion method was used to examine the antibacterial activity of compounds against Gram (+) bacteria (Staphylococcus aureus and Micrococcus luteus) and Gram (-) bacteria (Escherichia coli and Klebsiella pneumonia), with streptomycin employed as control. The antifungal activity of fluconazole against Candida albicans and Aspergillus niger was measured against a benchmark of fluconazole's own performance. Compounds IVc, IVd, and IVe all exhibited potent antibacterial activity, but only compounds IVc and IVd were effective against fungal infections. Compounds IVc and IVe were found to have the highest potential as anti-oxidants among the chemicals that were investigated.

Bioactive triterpenoids from Solanum torvum fruits with antifungal, resistance modulatory and anti-biofilm formation activities against fluconazole-resistant candida albicans strains.

Vulvovaginal candidiasis (VVC) is the second most common vaginal infection that affects women of reproductive age. Its increased occurrence and associated treatment cost coupled to the rise in resistance of the causative pathogen to current antifungal therapies has necessitated the need for the discovery and development of novel effective antifungal agents for the treatment of the disease. We report in this study the anti-Candida albicans activity of Solanum torvum 70% ethanol fruit extract (STF), fractions and some isolated compounds against four (4) fluconazole-resistant strains of C. albicans. We further report on the effect of the isolated compounds on the antifungal activity of fluconazole and voriconazole in the resistant isolates as well as their inhibitory effect on C. albicans biofilm formation. STF was fractionated using n-hexane, chloroform (CHCl3) and ethyl acetate (EtOAc) to obtain four respective major fractions, which were then evaluated for anti-C. albicans activity using the microbroth dilution method. The whole extract and fractions recorded MICs that ranged from 0.25 to 16.00 mg/mL. From the most active fraction, STF- CHCl3 (MIC = 0.25-1.00 mg/mL), four (4) known compounds were isolated as Betulinic acid, 3-oxo-friedelan-20α-oic acid, Sitosterol-3-β-D-glucopyranoside and Oleanolic acid. The compounds demonstrated considerably higher antifungal activity (0.016 to 0.512 mg/mL) than the extract and fractions and caused a concentration-dependent anti-biofilm formation activity. They also increased the sensitivity of the C. albicans isolates to fluconazole. This is the first report of 3-oxo-friedelan-20α-oic acid in the plant as well as the first report of betulinic acid, sitosterol-3-β-D-glucopyranoside and oleanolic acid from the fruits of S. torvum. The present study has demonstrated the anti-C. albicans activity of the constituents of S. torvum ethanol fruit extract and also shown that the constituents possess anti-biofilm formation and resistance modulatory activities against fluconazole-resistant clinical C. albicans isolates.

Effect of Distreptaza Distrept on the specific activity of fluconazole against Candida biofilms

The ability of yeast-like fungi to form biofilms (structurally organized microbial communities) leads to chronicity of the inflammatory process and recurrent course of mycoses. Biofilms formed by the fungus Candida spp. are characterized by increased resistance to most antifungals and effectors of the immune system. Promising antibiotics are enzymes. Their specificity to the structural components of the biofilm can be used to prevent biofilm formation and destruction of the formed biofilm. The article presents the results of studying the sensitivity biofilms formed by Candida fungi to the action of the enzyme drug Distreptaza Distrept, as well as its effect on the specific antifungal activity of fluconazole.Research objective: to evaluate the Distreptaza Distrept impact on the antifungal activity of fluconazole on biofilms of Candida spp.Materials and methods. One-day cultures of C. albicans and C. glabrata were used in the experiments. To study the effect on biofilm formation Distreptaza Distrept, fluconazole solutions and microorganisms were applied simultaneously, when exposed to the formed biofilms for 24 h after application of the fungal inoculum. The experiments were carried out in accordance with conventional methods based on the sorption of gentian violet by biofilm structures with subsequent desorption of the dye in an organic solvent.Results. Experiments have shown that Distreptaza Distrept disrupts the film formation of C. glabrata, the inhibition is 85.6%. The enzyme preparation affects the biofilms formed by yeast-like fungi, biomass of C. glabrata decreases by 43.6%. It was found that Distreptaza Distrept enhances the specific antibiotic action of fluconazole at the stage of film formation against C. albicans by 13.1%, C. glabrata by 70.4%. Biomass of the one-day biofilm formed by C. glabrata is reduced by 80.2% if there was an enzyme drug and fluconazole in the incubation medium.Conclusion. Distreptaza Distrept has antibiotic biofilm activity, increases the specific antifungal effect of fluconazole on biofilms of Candida fungi. The data obtained indicate the feasibility of this drug in recurrent infections caused by yeast-like fungi, including diseases caused by azole-resistant strains of Candida spp.

Susceptibility Pattern of Different Parts of Moringa oleifera against Some Pathogenic Fungi, Isolated from Sputum Samples of HIV Positive Individuals Co-Infected with Pulmonary Tuberculosis

Background: Opportunistic fungal infection in HIV positive individualshas a high risk of morbidity and mortality as it could be a pointer to systemic infection. Moringa oleifera is one of the 14 species of the family Moringaceae with traditional effectiveness in anti–helminthic, analgesic, anti-ulcerative, anti-microbial, etc.&#x0D; Aim: This study was designed to determine the susceptibility pattern of different parts of Moringa oleifera against some pathogenic fungi using dried Morringa oleifera seed, bark, pod, and leaf extracts/fractions as agents.&#x0D; Materials and Methods: A total of 132 subjects (62 males and 70 females) attending outpatient clinics (OPD) at Nnamdi Azikiwe University Teaching Hospital Nnewi, and Holy Rossary Hospital Emekuku Owerri, Nigeria were recruited for this study. The HIV status of the test subjects was confirmed using immuno-chromatographic technique and their sputum screened for Tuberculosis (TB) using Ziehl Nelsen staining technique. The sputum samples were also cultured on Sabouaurd dextrose agar with and without chloramphenicol incubated at 25oC with examination of growth every 3 days for two weeks. Identification of isolates was done using Gram staining methods, chromogenic media and slide culture.700g, 220g, 500g, and 250g each of the different parts of the pulverized Moringa oleifera (seeds, barks, pods, and leaves) were weighed and exctracted using Soxhlet extractor and methanol as solvent.&#x0D; Results: The prevalence of fungi isolates from test subjects shows that Candida albicans has the highest frequency of 25(26.6%), with the least prevalent being Aspergillus flavus and Phialemonium curvatum at 2(2.1%) respectively. There was also a positive significant correlation between TB and fungi infection in HIV-sero positive subjects (P&lt;0.05).The invitro antifungal activity of these Moringa oleifera extracts/fractions against the test isolates showed no antifungal activity except for ethyl acetate fraction which demonstrated the most prominent activity against A. fumigatus with zones of inhibition of 3-10mm diameter with the least activity against C. krusei with zones of inhibition of 3-6mm diameter. However, the antifungal activities of fluconazole which served as a control in the antifungal susceptibility against the test isolates were more effective than the extract/fractions against the test isolates with a minimum inhibition concentration (MIC) of 50-200mg/ml against 25-100mg/ml for ethylacetate fraction.&#x0D; Conclusion: The comparable antifungi effect of ethylacetate fraction of M. oleifera leaf with that of fluconazole, suggests that ethylacetate fraction of M. oleifera leaf could aid the treatment of pathogenic fungi due to its potential applications in Pharmaceutical industry, thereby ameliorating the morbidity and mortality rate of HIV positive co-infected TB subjects.

Fluphenazine antagonizes with fluconazole but synergizes with amphotericin B in the treatment of candidiasis

Candida albicans causes a high mortality rate in immunocompromised individuals, but the increased drug resistance challenges the current antifungal therapeutics. Fluphenazine (FPZ), a commonly used antipsychotic medication, can induce the expression of drug efflux pumps in C. albicans and, thus, may interfere with the therapeutic efficacy of antifungals, such as fluconazole (FLC) and amphotericin B (AmB). Here, we investigated the combined effects of FLC/FPZ and AmB/FPZ against C. albicans in vitro and in a systemic candidiasis mouse model. The antifungal activity of FLC was significantly reduced when supplemented with FPZ. The inhibitory effects of FLC on the expression of the Candida virulence-related genes ALS3 and HWP1 were antagonized by FPZ. However, FPZ enhanced the susceptibility of C. albicans to AmB and further downregulated the expression of ALS3 and HWP1 in a synergistic manner with AmB. FPZ also enhanced the gene expression of ERG11, a key gene of the ergosterol biosynthesis pathway that has been associated with the activities of both FLC and AmB. In our mammalian infection model, mice treated with FLC/FPZ showed notably poor living status and increased fungal burden in their kidneys and brains compared with those treated with FLC alone. Conversely, the combined application of AmB/FPZ significantly improved the survival rate, attenuated the weight loss and reduced the organ fungal burdens of the infected mice. These data suggest that FPZ antagonized the therapeutic efficacy of FLC but enhanced the antifungal activity of AmB in the treatment of candidiasis.

Antifungal Susceptibility Testing For Dermatophytes Isolated From Human and Animal Dermatophytosis

This study was designed to characterize antifungal susceptibility pattern of different species of dermatophytes isolated from human and animal dermatophytosis against six commercially available antifungal agents recommended for treatment. Broth microdilution method was used to determine minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of different drugs against the tested dermatophyte species. MIC was done against fluconazole, itraconazole, terbinafine, tioconazole, miconazole and griseofulvin. Of the antifungal agents tested, the best results in terms of sensitivity were found with terbinafine for all isolates and tioconazole, miconazole for M. canis and T. verrucosum while the antifungal activity of fluconazole was found to be weak. Routine detection of MIC for antifungal agents serves as a rapid and reliable method for treatment of human and animal dermatophytosis.

Synergistic in vitro activity of sodium houttuyfonate with fluconazole against clinical Candida albicans strains under planktonic growing conditions

Context: Fluconazole resistance is an intractable problem of treating Candida albicans, calling for more antifungal agents to enhance the activity of fluconazole. Objective: This work investigates the anti-C. albicans activities of sodium houttuyfonate (SH) and/or fluconazole and the associated mechanism. Materials and methods: The minimum inhibitory concentrations (MICs) of SH and fluconazole both ranging from 0.5 to 1024 μg/mL were determined by broth microdilution method in 19 C. albicans isolates, and their fractional inhibitory concentration index (FICI) was evaluated by checkerboard assay. After MICSH and/or MICfluconazole treatments, the expressions of IFD6, PHR1, ZAP1, ADH5, BGL2, XOG1 and FKS1 were analyzed by quantitative reverse transcription polymerase chain reaction (qRT-PCR) in C. albicans 1601. Results and conclusion: The MICs of SH alone ranged from 32 to 256 μg/mL and decreased 2–16-fold in combination. SH showed strong synergism with fluconazole with FICI <0.13–0.5. In C. albicans 1601, we observed that (i) the expression of the seven genes increased notably in a range between 3.71- and 12.63-fold (p < 0.05) when SH was used alone, (ii) the combined use of SH and fluconazole slightly inhibited the expression of IFD6 and PHR1 by 1.23- and 1.35-fold (p > 0.05), but promoted evidently the expression of ZAP1, ADH5, XOG1 and FKS1 by 1.98-, 3.56-, 4.10- and 2.86-fold (p < 0.05). The results suggested SH to be a potential synergist to enhance the antifungal activity of fluconazole in C. albicans resistant isolates, and the underlying mechanism may be associated with β-1,3-glucan synthesis and transportation.

Synergic effect of combination of glycyrol and fluconazole against experimental cutaneous candidiasis due to Candida albicans.

In this study, we investigated the anti-fungal activity of glycyrol, a coumarine isolated from licorice (Glycyrrhizae Radix), in a murine model of cutaneous candidiasis caused by Candida albicans. Compared to the infected sites, located on the mice's back, of the untreated control mice, the infected sites treated with glycyrol had reduced CFU (colony forming unit) values up to 60 and 85.5% at 20 and 40μg/mouse of glycyrol, respectively (P<0.01). The antifungal activity of glycyrol was synergistically increased when glycyrol (10μg/mouse) was combined with fluconazole (10μg/mouse), demonstrating that the combination therapy is approximately 4 times more effective than fluconazole alone at 20μg/mouse (P<0.01). Additionally, the combination activity was 1.65 times greater than the antifungal activity of fluconazole alone at 40μg/mouse (P<0.05). In seeking glycyrol's antifungal mechanism, we determined that glycyrol inhibited hyphal induction and cell wall adherence of C. albicans. Thus, it is very likely that, by damaging the cell wall, glycyrol helps fluconazole invade C. albicans more readily and attack fluconazole's target in the fungus membrane. In summary, our data indicate that glycyrol may contribute to the development of a novel agent that possesses antifungal activity against cutaneous candidiasis.

The Combination of Chlorogenic Acid and Fluconazole Potentiates Antifungal Activity

근래에 폴리페놀성분(polyphenols)의 다양한 약물활성에 대한 많은 연구가 집중되고 있는 경향으로, 본 연구실에서도 폴리페놀성분의 일종인 chlorogenic acid(CRA)의 항관절염효과1)와 면역보조제효과2)에 대한 연구결과를 이미 보고한 바 있다. CRA의 항관절염효과는 Candida albicans 기인성 패혈성 관절염(septicarthritis) 동물모델에서 조사한 것으로, 이 연구과정에서 CRA의 항진균효과가 검색되었지만,2) 이 성분의 상승효과는 본 연구에서 규명되었다. 한편, 타연구자들의 보고에 의하면, CRA는 항산화 및 항암작용이 있으며,3,4) 진경과 해열작용 등도 있다는 보고가 있다.5,6) 또한, 다른 연구실에서 CRA의 항진균작용을 보고한 바가 있으나,7) CRA의 항진균활성의 상승효과에 관련된 연구는 결과는 현재까지 알려진 바가 없다. 이와 같이, 다양한 약물활성이 있는 CRA는 식물계에 광범위하게 분포되어 있는데, 이 성분은 카페인산(caffeic acid)과 퀸산(quinic acid)의 에스테르반응에 의해 형성되어 있다(Fig. 1A).3) 특히, CRA는 음식물에 함유된 flavonoid 성분들 중에서 가장 풍부하게 포함된 성분으로 알려져있다.In this present study, we determined the ability of chlorogenic acid (CRA), a polyphenolic compound, to potentiate the antifungal activity of fluconazole (FCZ) against Candida albicans. The determination was investigated by a microdilution susceptibility method and in a murine model of cutaneous candidiasis due to C. albicans. Results showed that CRA had a dose-dependent antifungal activity; and when combined with FCZ, the antifungal activity was synergistic as determined by the microdilition susceptibility method. For example, a combination of CRA at 25 μg/ml plus FCZ at 0.1 μg/ml was approximately 3 times more effective than antifungal activity of FCZ alone at the same dose (P<0.01). This potentiation by CRA was almost equivalent to antifungal activity by FCZ alone at 0.5 μg/ml, displaying that the CRA potentiates FCZ’s antifungal activity by almost 5 times (P<0.01). Moreover, this combination demonstrated synergic activity against the cutaneous disease as well, resulting in approximately 3 times more greater antifungal activity than FCZ alone at 0.1 μg/mouse (P<0.01). This suggests that the combination therapy can reduce side effects caused by FCZ in clinical stuations. In summary, CRA has a synergistic antifungal activity, which can be produced by combining CRA with FCZ. Therefore, our data strongly indicate that CRA can be a potential candidate as an antifungal agent for combination therapy.

Amino acid substitutions in Erg11p of azole-resistant Candida glabrata: Possible effective substitutions and homology modelling.

Understanding the mechanisms responsible for fluconazole resistance in Candida glabrata is not only crucial for the development of new antifungals but is also important in choosing appropriate antifungals for patients at the earliest stages. The aim of this study was to determine the Erg11p amino acid substitutions in fluconazole-resistant C. glabrata isolates. Sixty clinical isolates of C. glabrata were investigated. In vitro antifungal activities of fluconazole, itraconazole and voriconazole were determined using the broth microdilution reference method. The ERG11 gene for resistant (n=4) and susceptible (n=1) isolates were sequenced and multi-aligned using MEGA6 software. A homology model of the C. glabrata ERG11 gene was created by SWISS-MODEL software using the crystal structure of Saccharomyces cerevisiae Erg11p as a template, and the predicted binding sites to fluconazole were investigated. Fluconazole and multi-azole resistance were observed in 6.7% and 3.3% of the isolates, respectively. Several amino acid substitutions were identified, among which some were also identified in susceptible isolates. The amino acid substitution G236V was at the binding site, and substitutions H146Q and D234E were near to the binding site of triazoles according to the SWISS-MODEL. According to the homology modelling results, the amino acid substitution G236V is highly likely to play a key role in azole resistance development.

High Glucose Concentrations Decrease the in Vitro Antifungal Activity of Fluconazole against Candida Albicans as Proved by CLSI Broth Microdilution and Disk Diffusion Methods

Background: Glucose ≥ 1% was proved to decrease antifungal activity of voriconazole and amphotericin B against Candida spp. in varying degrees. This effect has not been studied with fluconazole. Objectives: Testing in vitro effect of glucose on fluconazole activity against C. albicans to investigate whether high blood glucose levels encountered in hyperglycemia can affect the response to fluconazole treatment in diabetics and whether adding glucose to culture media can affect the results of fluconazole susceptibility testing. Methodology: Clinical and Laboratory Standards Institute (CLSI) broth microdilution and disk diffusion methods were used for testing fluconazole against C. albicans ATCC 10231 and additional 5 clinical isolates in presence of wide range of glucose concentrations (0 mg/dl-8000 mg/dl). Fluconazole MIC ranges, MIC 50 , MIC 90 and inhibition zones (IZ) were determined. Results: The MIC ranges were 0.125-0.25 µg/ml, 0.125-0.50 µg/ml, 0.25-1 µg/ml, and 2-8 µg/ml for glucose concentrations 200 mg/dl, 500 mg/dl, 1000 mg/dl, 8000 mg/dl, respectively. MIC 50 increased up to sixteen fold with glucose concentration 8000 mg/dl and MIC 90 increased sixteen fold with glucose concentrations 4000 mg/ml and 6000 mg/dl, and thirty two fold with 8000 mg/dl concentration. The hyperglycemic glucose concentrations range (300 mg/dl to 600 mg/dl) showed maximum one fold increase of both MIC 50 and MIC 90 . All six tested strains showed decreases of the IZ with increase in glucose concentrations. These decreases were statistically significant in 2 strains with glucose concentrations 300 mg/dl to 600 mg/dl, in half of tested strains with glucose concentrations 700 mg/dl and 800 mg/dl, and with five strains for concentrations 1000 mg/dl and 2000 mg/dl. All tested strains expressed statistically significant decreases in IZ with concentrations 4000 mg/dl to 8000 mg/dl. Conclusion: This in vitro study showed decrease in antifungal activity of fluconazole against C. albicans by exposure to increasing glucose concentrations and this warrants special care for prescribing this antifungal drug for diabetics and caution during the interpretation of the results of its susceptibility testing against C. albicans when tested by using media containing glucose.

The roles of CDR1, CDR2, and MDR1 in kaempferol-induced suppression with fluconazole-resistant Candida albicans

Context: Fungal infections caused by fluconazole-resistant Candida albicans are an intractable clinical problem, calling for new efficient antifungal drugs. Kaempferol, an active flavonoid, has been considered a potential candidate against Candida species. Objective: This work investigates the resistance reversion of kaempferol in fluconazole-resistant C. albicans and the underlying mechanism. Materials and methods: The antifungal activities of fluconazole and/or kaempferol were assessed by a series of standard procedures including broth microdilution method, checkerboard assay and time-kill (T-K) test in nine clinical strains as well as a standard reference isolate of C. albicans. Subsequently, the morphological changes, the efflux of rhodamine 6G, and the expressions of CDR 1, CDR 2, and MDR 1 were analysed by scanning electron microscope (SEM), inverted fluorescence microscope and quantitative reverse transcription polymerase chain reaction (qRT-PCR) in C. albicans z2003. Results: For all the tested C. albicans strains, the minimum inhibitory concentrations (MICs) of fluconazole and kaempferol ranged 0.25–32 and 128–256 μg/mL with a range of fractional inhibitory concentration index of 0.257–0.531. In C. albicans z2003, the expression of both CDR 1 and CDR 2 were decreased after exposure to kaempferol alone with negligible rhodamine 6G accumulation, while the expression of CDR 1, CDR 2 and MDR 1 were all decreased when fluconazole and kaempferol were used concomitantly with notable fluorescence of rhodamine 6G observed. Discussion and conclusion: Kaempferol-induced reversion in fluconazole-resistant C. albicans might be likely due to the suppression of the expression of CDR1, CDR2 and MDR1.

Possible mechanisms of the antifungal activity of fluconazole in combination with terbinafine against Candida albicans

Context: Candidiasis is a term describing infections by yeasts from the genus Candida, the majority Candida albicans. Treatment of such infections often requires antifungals such as the azoles, but increased use of these drugs has led to selection of yeasts with increased resistance to these drugs.Objective: Combination therapy would be one of the best strategies for the treatment of candidiasis due to increased resistance to azoles.Materials and methods: The antifungal activities of fluconazole and terbinafine were evaluated in vitro alone and in combination using broth microdilution test and time kill study. Eventually the expression level of selected genes involved in ergosterol biosynthesis of Candida was evaluated using semi-quantitative RT-PCR.Results: The obtained results showed the significant MICs ranging from 0.25 to 8 µg/mL followed by FICs ranged from 0.37 to 1 in combination with fluconazole/terbinafine. Our findings have demonstrated that the combination of fluconazole and terbinafine could also significantly reduce the expression of ERG1, 3, and 11 in the cell membrane of Candida in all concentrations tested ranging from 1.73- to 6.99-fold.Discussion and conclusion: This study was undertaken with the ultimate goal of finding the probable targets of fluconazole/terbinafine in C. albicans by looking at its effects on cell membrane synthesis.

Synergistic effect of fluconazole and doxycycline against Candida albicans biofilms resulting from calcium fluctuation and downregulation of fluconazole-inducible efflux pump gene overexpression.

Candida albicans biofilms are intrinsically resistant to antimicrobial agents. Previous work demonstrated that the antifungal activity of fluconazole against C. albicans biofilms is notably enhanced by doxycycline. In order to explore the synergistic mechanism of fluconazole and doxycycline, we investigated the changes of efflux pump gene expression, intracellular calcium concentration and cell cycle distribution after drug intervention in this study. The expression levels of CDR1, CDR2 and MDR1 were determined by real-time PCR, and the results showed that fluconazole alone could stimulate the high expression of CDR1, CDR2 and MDR1, and the combination of doxycycline and fluconazole downregulated the gene overexpression induced by fluconazole. Intracellular calcium concentration was determined using Fluo-3/AM by observing the fluorescence with flow cytometry. A calcium fluctuation, which started 4 h and peaked 8 h after the treatment with fluconazole, was observed. The combined drugs also initiated a calcium fluctuation after 4 h treatment and showed a peak at 16 h, and the peak was higher than that stimulated by fluconazole alone. The cell cycle was measured using flow cytometry. Fluconazole alone and the combined drugs both induced a reduction in the percentages of S-phase cells and an elevation in the percentages of cells in the G2/M phase. The results of this research showed that the synergism of fluconazole and doxycycline against C. albicans biofilms is associated with blockade of the efflux pump genes CDR1, CDR2 and MDR1, and stimulation of high intracellular calcium concentration. The findings of this study are of great significance in the search for new antifungal mechanisms.

Thermoresponsive fluconazole gels for topical delivery: rheological and mechanical properties, in vitro drug release and anti-fungal efficacy

The aim of this study was to develop thermosensitive gels using poloxamers for topical delivery of fluconazole (FLZ). Eight different formulations containing 1% FLZ in poloxamer and a particular co-solvent (propylene glycol (PG) or Transcutol-P) of various concentrations were prepared. The gels were characterized for transition temperatures, rheological and mechanical properties. FLZ permeability and antifungal effect of the gels were also evaluated. Except for one formulation, all gels exhibited thermosensitive property, i.e. transformed from Newtonian (liquid-like) behavior at 20 °C to non-Newtonian (gel-like) behavior at 37 °C. Transcutol-P increased the transition temperature of the formulations, while the opposite effect was observed for PG. At 37 °C, formulations with high poloxamer concentrations (17%) resulted in high viscosity, compressibility and hardness. Formulations containing 17% poloxamer and 20% Transcutol-P and 10% PG, respectively, exhibited high adhesiveness. No significant differences in the in vitro antifungal activity of FLZ were observed among the formulations suggesting that the gel vehicles did not influence the biological effect of FLZ. FLZ permeability decreased with increasing poloxamer concentration. Formulations containing 17% poloxamer and 20% Transcutol-P and 10% PG seemed to be promising in situ gelling systems for the topical delivery of FLZ.

Metallic silver nanoparticle: a therapeutic agent in combination with antifungal drug against human fungal pathogen

Silver nanoparticles (Ag-NPs) are known to have inhibitory and fungicidal effects. Resistance against fungal infection has emerged as a major health problem in recent years, which needs great and immediate concern. Here, we report the extracellular biological synthesis of silver nanoparticles through a simple green route approach using a marine mangrove (Rhizophora mucronata) and silver nitrate. Aqueous extract of marine mangrove helped in reduction and was used as capping agent in biological synthesis. Nanoparticles were characterized using microscopy and spectroscopy techniques such as HRTEM, UV-Vis absorption spectroscopy and FTIR spectroscopy. X-ray diffraction analysis showed that the nanoparticles had face centered cubic structure with crystalline nature. FTIR spectroscopy showed the presence of different functional groups, such as hydroxyl and carbonyl, involved in the synthesis of nanoparticles. The antifungal activity of fluconazole and itraconazole was enhanced against the tested pathogenic fungi in the presence of Ag-NP and confirmed from increase in fold area of inhibition. This environmentally friendly method of biological synthesis can be easily integrated for various medical applications.