Fluconazole-, Amphotericin-B-, Caspofungin-, and Anidulafungin-Resistant <Emphasis Type="Italic">Candida ciferrii</Emphasis>: An Unknown Cause of Systemic Mycosis in a Child

Candida ciferrii, which is known as an agent of superficial yeast infection and onychomycosis, has rarely been isolated as an agent of candidemia. Limited reports have suggested different patterns of antifungal sensitivity. We report a rare candidemia case caused by c.ciferrii in an 8-year-old child in which isolated candida species were resistant to amphotericin-B (MIC > 1 μg/ml), fluconazole, (MIC ≥ 64 μg/ml), caspofungin (MIC ≥ 32 μg/ml), and anidulafungin (MIC ≥ 32 μg/ml) but sensitive to voriconazole (MIC ≤ 0.12 μg/ml). As far as we aware, this was the first recorded C. ciferrii candidemia case in children.     

In Vitro Activities of New Triazole Antifungal Agents, Posaconazole and Voriconazole, Against Oral Candida Isolates from Patients Suffering from Denture Stomatitis

Denture stomatitis is often treated with antifungal agents but recurrences or new episodes are common, and certain episodes can be resistant. New triazoles, such as posaconazole and voriconazole, may represent useful alternatives for management. In vitro activities of amphotericin B, nystatin, miconazole, fluconazole, itraconazole, posaconazole and voriconazole against 150 oral Candida (101 C. albicans, 18 C. tropicalis, 12 C. glabrata, 11 C. guilliermondii, 4 C. parapsilosis, 2 Saccharomyces cerevisiae, 1 C. dubliniensis and 1 C. krusei) from 100 denture wearers were tested by the CLSI M27-A3 method. Resistant isolates were retested by Sensititre YeastOne and Etest. Most antifungal agents were very active. However, 4 C. glabrata (33.3%), 2 C. tropicalis (11.1%), 6 C. albicans (5.6%) and 1 C. krusei were resistant to itraconazole. Posaconazole was active against 143 yeast isolates (95.3%): 6 C. albicans (5.9%) and 1 C. tropicalis (5.6%) were resistant. Geometric mean MICs were 0.036 μg/ml for C. parapsilosis, 0.062 μg/ml for C. albicans, 0.085 μg/ml for C. tropicalis, 0.387 μg/ml for C. guilliermondii and 0.498 μg/ml for C. glabrata. Voriconazole was active against 148 isolates (98.7%) with geometric mean MICs ranging from 0.030 μg/ml for C. parapsilosis, 0.042 μg/ml for C. albicans, 0.048 μg/ml for C. tropicalis, 0.082 μg/ml for C. guilliermondii, to 0.137 μg/ml for C. glabrata. Only 2 C. albicans (2%) were resistant to voriconazole showing cross-resistance to other azoles. Posaconazole and voriconazole have excellent in vitro activities against all Candida isolates and could represent useful alternatives for recalcitrant or recurrent candidiasis.     

Susceptibility of <Emphasis Type="Italic">Candida</Emphasis> biofilms to histatin 5 and fluconazole

Candida-associated denture stomatitis has a high rate of recurrence. Candida biofilms formed on denture acrylic are more resistant to antifungals than planktonic yeasts. Histatins, a family of basic peptides secreted by the major salivary glands in humans, especially histatin 5, possess significant antifungal properties. We examined antifungal activities of histatin 5 against planktonic or biofilm Candida albicans and Candida glabrata. Candida biofilms were developed on poly(methyl methacrylate) discs and treated with histatin 5 (0.01–100 μM) or fluconazole (1–200 μM). The metabolic activity of the biofilms was measured by the XTT reduction assay. The fungicidal activity of histatin 5 against planktonic Candida was tested by microdilution plate assay. Biofilm and planktonic C. albicans GDH18, UTR-14 and 6122/06 were highly susceptible to histatin 5, with 50% RMA (concentration of the agent causing 50% reduction in the metabolic activity; biofilm) of 4.6 ± 2.2, 6.9 ± 3.7 and 1.7 ± 1.5 μM, and IC₅₀ (planktonic cells) of 3.0 ± 0.5, 2.6 ± 0.1 and 4.8 ± 0.5, respectively. Biofilms of C. glabrata GDH1407 and 6115/06 were less susceptible to histatin 5, with 50% RMA of 31.2 ± 4.8 and 62.5 ± 0.7 μM, respectively. Planktonic C. glabrata was insensitive to histatin 5 (IC₅₀ > 100 μM). Biofilm-associated Candida was highly resistant to fluconazole in the range 1–200 μM; e.g. at 100 μM only ~20% inhibition was observed for C. albicans, and ~30% inhibition for C. glabrata. These results indicate that histatin 5 exhibits antifungal activity against biofilms of C. albicans and C. glabrata developed on denture acrylic. C. glabrata is significantly less sensitive to histatin 5 than C. albicans.     

Ciprofloxacin-Induced Antibacterial Activity is Reversed by Vitamin E and Vitamin C

In the present study, we investigated the possible involvement of oxidative stress in ciprofloxacin-induced cytotoxicity against several reference bacteria including Pseudomonas aeruginosa ATCC 27853, Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 29213, and clinical isolate of methicillin-resistant Staphylococcus aureus (MRSA). Oxidative stress was assessed by measurement of hydrogen peroxide generation using a FACScan flow cytometer. The antibacterial activity of ciprofloxacin was assessed using the disk diffusion method and by measuring the minimum inhibitory concentration (MIC). Ciprofloxacin induced a dose-dependent antibacterial activity against all bacteria where the highest tested concentration was 100 ug/ml. Results revealed that E. coli cells were highly sensitive to ciprofloxacin (MIC = 0.21 μg/mL ± 0.087), P. aeruginosa and S. aureus cells were intermediately sensitive (MIC = 5.40 μg/mL ± 0.14; MIC = 3.42 μg/mL ± 0.377, respectively), and MRSA cells were highly resistant (MIC = 16.76 μg/mL ± 2.1). Pretreatment of E. coli cells with either vitamin E or vitamin C has significantly protected cells against ciprofloxacin-induced cytotoxicity. These results indicate the possible antagonistic properties for vitamins C or E when they are used concurrently with ciprofloxacin.     

In vitro antifungal activity of the berberine and its synergism with fluconazole

Berberine with and without fluconazole was tested by an agar disk diffusion assay in which clinical isolates of Candida albicans were applied onto yeast extract-peptone-dextrose agar plate. Berberine, which had no intrinsic antifungal activity at the concentration tested, exerted a powerful antifungal activity in combination of fluzonazole. Combinations of berberine and fluconazole were also tested by the checkerboard assay to determine whether they had favorable or unfavorable antifungal interactions. The MIC of fluconazole was 1.9 μg/ml when the drug was tested alone and decreased to 0.48 μg/ml in the presence of berberine concentrations of 1.9 μg/ml. However, berberine at concentrations of >1.9 μg/ml combined with a fluconazole supra-MIC (i.e., >1.9 μg/ml) eliminated the residual turbidity in the incubation wells. This endpoint fitted to the definition of MIC-0 (optically clear wells) and reflected the absence of a trailing effect, which is the result of a residual growth at fluconazole concentrations greater than the MIC.     

In vitro antifungal activities of voriconazole and reference agents as determined by NCCLS methods: Review of the literature

Voriconazole (Vfend™) is a new triazole that currently is undergoing phase III clinical trials. This review summarizes the published data obtained by NCCLS methods on the in vitro antifungal activity of voriconazole in comparison to itraconazole, amphotericin B, fluconazole, ketoconazole and flucytosine. Voriconazole had fungistatic activity against most yeasts and yeastlike species (minimum inhibitory concentrations [MICs] <2 μg/ml) that was similar or superior to those of fluconazole, amphotericin B, and itraconazole. Against Candida glabrata and C. krusei, voriconazole MIC ranges were 0.03 to 8 and 0.01 to >4 μg/ml, respectively. For four of the six Aspergillus spp. evaluated, voriconazole MICs (< 0.03 to 2 μg/ml) were lower than amphotericin B (0.25 to 4 μg/ml) and similar to itraconazole MICs. Voriconazole fungistatic activity against Fusarium spp. has been variable. Against F. oxysporum and solani, most studies showed MICs ranging from 0.25 to 8 μg/ml. Voriconazole had excellent fungistatic activity against five of the six species of dimorphic fungi evaluated (MIC₉₀s < 1.0 μg/ml). The exception was Sporothrix schenckii (MIC₉₀s and geometric mean MICs ≥ 8 μg/ml). Only amphotericin B had good fungistatic activity against the Zygomycetes species (voriconazole MICs ranged from 2 to >32 μg/ml). Voriconazole showed excellent in vitro activity (MICs < 0.03 to 1.0 μg/ml) against most of the 50 species of dematiaceous fungi tested, but the activity of all the agents was poor against most isolates of Scedosporium prolificans and Phaeoacremonium parasiticum (Phialophora parasitica). Voriconazole had fungicidal activity against most Aspergillus spp., B. dermatitidis, and some dematiaceous fungi. In vitro/in vivo correlations should aid in the interpretation of these results. This revised version was published online in June 2006 with corrections to the Cover Date.     

In Vitro susceptibility of 137 Candida sp. Isolates from HIV positive patients to several antifungal drugs

Oropharyngeal candidiasis caused by various species of Candida is one of the most common infections in HIV seropositive or AIDS patients. Drug resistance among these yeasts is an increasing problem. We studied the frequency of resistance profile to fluconazole, itraconazole, ketoconazole, amphotericin B and terbinafine of 137 isolates of Candida sp. From HIV positive or AIDS patients with oropharyngeal candidiasis at Instituto de Inmunología, U.C.V. and the Hospital “Jose Ignacio Baldó”, Caracas Venezuela, using the well diffusion susceptibility test (Magaldi et al.). We found that nearly 10% of C. albicans isolates were primarily fluconazole resistant, 45% of C. albicans isolates from patients with previous treatment were resistant to fluconazole, of which 93% showed cross-resistance to itraconazole, and even about 30% of C. tropicalis (n = 13) were resistant to fluconazole and/or itraconazole. To this respect, several recent reports have been described antifungal cross-resistance among azoles. Therefore, we consider that C. tropicalis should be added to the growing list of yeast in which antifungal drug resistance is common. This report could be useful for therapeutic aspect in AIDS patients with oral candidiasis. This revised version was published online in July 2006 with corrections to the Cover Date.     

重症监护病房真菌谱及耐药性6年变化趋势

目的 调查我院重症监护室（ICU）近6 a临床分离真菌株的菌种结构及耐药性变化.方法 2007 ～2012年从我院1435例ICU患者中分离出的1 435株真菌均为首次分离的菌株.采用Cox-Stuart趋势检验回顾性分析了分离真菌株的菌种结构和主要真菌对常用抗真菌药物耐药率的变化趋势.结果 2007 ～2012年间,我院tCU真菌检出阳性标本总数从159株/a逐年增加至396株/a.其中,白念珠菌、近平滑念珠菌、曲霉的构成比波动范围分别为34.6％ ～ 50.3％、6.3％～9.6％、1.3％～3.2％,2010 ～2012年较2007 ～2009年构成比总体均有上升趋势,P=0.02.热带念珠菌、光滑念珠菌、克柔念珠菌构成比波动范围分别为13.6％ ～ 17.3％、9.4％ ～ 22.6％、1.4％ ～5.2％,P=0.18,构成比均无明显上升或下降趋势.2010 ～2012年较2007 ～2009年,白念珠菌对氟康唑和伊曲康唑耐药率的变化均呈降低趋势,P=0.02.热带念珠菌对氟康唑和伊曲康唑耐药率的变化均呈降低趋势,P =0.02;光滑念珠菌对伊曲康唑耐药率的变化总体呈上升趋势,P =0.02.结论 我院ICU近6 a来白念珠菌仍然为临床最常见的真菌分离株,白念珠菌、近平滑念珠菌及曲霉占总真菌分离株的百分比有上升趋势.白念珠菌及热带念珠菌对氟康唑和伊曲康唑耐药率均呈降低趋势,光滑念珠菌对伊曲康唑耐药率呈上升趋势.     

Evaluation of Antifungal Susceptibility Testing with Microdilution and Etest Methods of <Emphasis Type="Italic">Candida</Emphasis> Blood Isolates

Candida species that show an increasing number of clinical and/or microbiological resistance to several antifungals and are the most common agents of invasive fungal infections. The aim of this study was to investigate the in vitro susceptibility of Candida blood isolates to antifungal agents (amphotericin B, fluconazole, itraconazole, and voriconazole) by comparative use of the CLSI reference microdilution method and Etest. Four hundred Candida blood isolates (215 Candida albicans, 185 non-albicans Candida strains) were included in the study. The broth microdilution test was performed according to the CLSI M27 A2 document. Etest was carried out according to the manufacturer’s instructions. The MIC results obtained with reference microdilution were compared with those obtained with the Etest by using percent and categorical agreements. According to MIK₉₀ values, voriconazole was the most active and itraconazole was the least active drug in vitro against all Candida species. Other than voriconazole, statistically significant differences were found when the susceptibility of Candida albicans and non-albicans Candida spp. to amphotericin B, fluconazole, and itraconazole were compared. These antifungal agents were found to be more active to C. albicans. Among the non-albicans Candida species, the lowest MIC values were obtained for Candida parapsilosis isolates. When the standard method was compared with Etest, the total agreement was higher for C. albicans than for non-albicans species, especially for fluconazole and voriconazole. In view of the findings, it was concluded that itraconazole showed the lowest activity against all Candida species. Etest could be an alternative method in assessing the in vitro antifungal susceptibility of Candida spp., but it is more convenient to use the microdilution method for studying in vitro susceptibility of non-albicans species, in particular for those possessing high MIC values against azoles.     

Wild-Type MIC Distributions and Epidemiologic Cutoff Values for Fluconazole and <Emphasis Type="Italic">Candida</Emphasis>: Time for New Clinical Breakpoints?

Antifungal susceptibility testing of Candida against fluconazole has been standardized by both the Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Both CLSI and EUCAST have developed clinical breakpoint (CBP) criteria for fluconazole, but these differ in both magnitude and target species. Studies using the EUCAST method have also defined wild-type minimum inhibitory concentration (MIC) distributions and epidemiologic cutoff values (ECVs or ECOFFs) for the common species of Candida. The ECVs serve as a sensitive means of discriminating wild-type strains from those with acquired resistance mechanisms and include MICs of 1 μg/mL for C. albicans, 2 μg/mL for C. tropicalis and C. parapsilosis, 32 μg/mL for C. glabrata, and 128 μg/mL for C. krusei. Because the CLSI CBPs may be too insensitive to detect emerging resistance among strains of C. albicans, C. tropicalis, and C. parapsilosis, and bisect the WT MIC distribution of C. glabrata, we sought to establish the wild-type MIC distribution and ECVs for fluconazole and Candida spp. The establishment of the wild-type MIC distributions and ECVs for fluconazole using CLSI methods will be useful in resistance surveillance and may prove to be an important step in the development of species-specific CBPs for this important antifungal agent.     

Minimum inhibitory concentration (MIC) of crude preparations of Brevibacillus laterosporus SA14 bioactive material compared to vancomycin and oxacillin, against clinical isolates of methicillin-resistant Staphylococcus aureus

Secondary metabolites, particularly bioactive compounds, from probiotic bacteria, are good candidates for replacing antibiotics to which bacteria have become resistant. In order to compare bioactive crude material from strain SA14 of Brevibacillus laterosporus with two antibiotics, the MICs of this bioactive crude and those of antibiotics vancomycin and oxacillin, against methicillin-resistant Staphylococcus aureus (MRSA), were determined. The result indicated that the MIC (3.6–19.2 μg/ml) of bioactive crude was higher than vancomycin (MIC = 1.28–2.56 μg/ml) when tested against MRSA. Interestingly, all tested strains of MRSA were susceptible to bioactive crude and were approximately 5.2-fold more potent than oxacillin (MIC > 100 μg/ml). Its activity against MRSA gives support for further evaluation, and the development of this substance for therapeutic use.     

Electrophoretic karyotyping and antifungal susceptibility patterns of Candida parapsilosis clinical isolates causing deep and superficial fungal infections

Forty-six isolates of Candida parapsilosis, each from a single patient, were collected from July 1993 through March 1999 at the University of Ancona Hospitals and Clinics. Twenty-eight strains were isolated from superficial lesioned sites, including skin, nails and other sources while 18 strains were isolated from blood. The isolates were typed by electrophoretickaryotyping (EK) and tested for their susceptibility to fluconazole (FLC),itraconazole (ITC), flucytosine (5-FC), and amphotericin B (AMB). Ourdata confirmed that EK is a useful technique for DNA typing of isolates ofCandida parapsilosis and showed that the source of isolation is notassociated with a given DNA type. Although strains belonging to this speciesof Candida are susceptible to the most common antifungals, including the triazoles, the degree of ITC susceptibility was dose dependent (MIC rangingfrom 0.25–0.5 μg/ml) for 98% of the isolates. This revised version was published online in June 2006 with corrections to the Cover Date.     

Antimicrobic and hemolytic activity of low-molecular metabolits of brown seaweed <Emphasis Type="Italic">Laminaria cichorioides</Emphasis> (Miyabe)

Antimicrobial and hemolytic activity of ethanol extract of brown seaweed Laminaria cichorioides (Miyabe), its lipophilic fractions, various classes of substances of lipophilic fraction, such as chlorophylls, fucoxanthin, monogalactosyldiacylglycerols, digalactosyldiacylglycerols, sulfoquinovosyldiacylglycerols, and fatty acids were investigated. The antimicrobial activity was studied by means of yeast cells Safale S-04 and Candida albicans KMM 455, fungi Aspergillus niger KMM 4634 and Fusarium oxysporum KMM 4639, bacteria Staphylococcus aureus ATCC 21027 (gram-positive) and Escherichia coli ATCC 15034 (gram-negative) which demonstrated selective sensitivity to the studied substances. Hemolytic activity was investigated at concentrations of substances in the range of 0.2–200 μg/ml at different pH of erythrocyte suspension. All investigated substances caused hemolysis. The dependence of hemolytic activity of substances on pH of media was determined.     

Effect of Azoles on the Secretion of a Candida albicans Metallopeptidase

Azole antifungals act by inhibiting the activity of a lanosterol demethylase involved in the generation of the ergosterol of the cellular membrane of fungi. These drugs could also have action on other yeast components, like secreted aspartyl proteases. We demonstrate in this study that the in vitro secretion of a metallopeptidase could be modified during the growth of Candida albicans with subinhibitory concentrations of some azoles. Eight isolates of this yeast have been cultivated in presence of MIC, MIC/2 and MIC/4 of voriconazole, fluconazole and itraconazole. The presence of voriconazole and fluconazole decreased the secretion of the metallopeptidase in the culture medium, whereas itraconazole increased this secretion for three isolates. This study points to the fact that some antifungals, given in prophylaxis, could act in an unfavourable way on some potential factors of pathogenicity.     

Anticandidal Efficacy of Cinnamon Oil Against Planktonic and Biofilm Cultures of <Emphasis Type="Italic">Candida parapsilosis</Emphasis> and <Emphasis Type="Italic">Candida orthopsilosis</Emphasis>

Candida parapsilosis is yeast capable of forming biofilms on medical devices. Novel approaches for the prevention and eradication of the biofilms are desired. This study investigated the anticandidal activity of sixteen essential oils on planktonic and biofilm cultures of C. parapsilosis complex. We used molecular tools, enumeration of colony-forming units, the colourimetric MTT assay, scanning electron microscopy (SEM) and a chequerboard assay coupled with software analyses to evaluate the growth kinetics, architecture, inhibition and reduction in biofilms formed from environmental isolates of the Candida parapsilosis complex; further, we also evaluated whether essential oils would interact synergistically with amphotericin B to increase their anticandidal activities. Of the environmental C. parapsilosis isolates examined, C. parapsilosis and C. orthopsilosis were identified. Biofilm growth on polystyrene substrates peaked within 48 h, after which growth remained relatively stable up to 72 h, when it began to decline. Details of the architectural analysis assessed by SEM showed that C. parapsilosis complex formed less complex biofilms compared with C. albicans biofilms. The most active essential oil was cinnamon oil (CO), which showed anticandidal activity against C. orthopsilosis and C. parapsilosis in both suspension (minimum inhibitory concentration—MIC—250 and 500 μg/ml) and biofilm (minimum biofilm reduction concentration—MBRC—1,000 and 2,000 μg/ml) cultures. CO also inhibited biofilm formation (MBIC) at concentrations above 250 μg/ml for both species tested. However, synergism with amphotericin B was not observed. Thus, CO is a natural anticandidal agent that can be effectively utilised for the control of the yeasts tested.     

Inhibition of Candida albicans growth by brominated furanones

Candida albicans is the most virulent Candida species of medical importance, which presents a great threat to immunocompromised individuals such as HIV patients. Currently, there are only four classes of antifungal agents available for treating fungal infections: azoles, polyenes, pyrimidines, and echinocandins. The fast spread of multidrug resistant C. albicans strains has increased the demand for new antifungal drugs. In this study, we demonstrate the antifungal activity of brominated furanones on C. albicans. Studying the structure and activity of this class of furanones reveals that the exocyclic vinyl bromide conjugated with the carbonyl group is the most important structural element for fungal inhibition. Furthermore, gene expression analysis using DNA microarrays showed that 3 μg/mL of 4-bromo-5Z-(bromomethylene)-3-butylfuran-2-one (BF1) upregulated 32 C. albicans genes with functions of stress response, NADPH dehydrogenation, and small-molecule transport, and repressed 21 genes involved mainly in cell-wall maintenance. Interestingly, only a small overlap is observed between the gene expression changes caused by the representative brominated furanone (BF1) in this study and other antifungal drugs reported in literature. This result suggests that brominated furanones and other antifungal drugs may target different fungal proteins or genes. The existence of such new targets provides an opportunity for developing new agents to control fungal pathogens which are resistant to currently available drugs.     

Identification,cloning, and functional characterization of EmrD-3, a putative multidrug efflux pump of the major facilitator superfamily from <Emphasis Type="Italic">Vibrio cholerae</Emphasis> O395

A putative multidrug efflux pump, EmrD-3, belonging to the major facilitator superfamily (MFS) of transporters and sharing homology with the Bcr/CflA subfamily, was identified in Vibrio cholerae O395. We cloned the emrD-3 gene and evaluated its role in antimicrobial efflux in a hypersensitive Escherichia coli strain. The efflux activity of this membrane protein resulted in lowering the intracellular concentration of ethidium. The recombinant plasmid carrying emrD-3 conferred enhanced resistance to several antimicrobials. Among the antimicrobials tested, the highest relative increase in minimum inhibitory concentration (MIC) of 102-fold was observed for linezolid (MIC = 256 μg/ml), followed by an 80.1-fold increase for tetraphenylphosphonium chloride (TPCL) (156.2 μg/ml), 62.5-fold for rifampin (MIC = 50 μg/ml), >30-fold for erythromycin (MIC = 50 μg/ml) and minocycline (MIC = 2 μg/ml), 20-fold for trimethoprim (MIC = 0.12 μg/ml), and 18.7-fold for chloramphenicol (MIC = 18.7 μg/ml). Among the fluorescent DNA-binding dyes, the highest relative increase in MIC of 41.7-fold was observed for ethidium bromide (125 μg/ml) followed by a 17.2-fold increase for rhodamine 6G (100 μg/ml). Thus, we demonstrate that EmrD-3 is a multidrug efflux pump of V. cholerae, the homologues of which are present in several Vibrio spp., some members of Enterobacteriaceae family, and Gram-positive Bacillus spp.     

Antifungal Activity of Antifungal Drugs,as Well as Drug Combinations Against <Emphasis Type="Italic">Exophiala dermatitidis</Emphasis>

To evaluate the in vitro efficacy of common antifungal drugs, as well as the interactions of caspofungin with voriconazole, amphotericin B, or itraconazole against the pathogenic black yeast Exophiala dermatitidis from China, the minimal inhibitory concentrations (MICs) of terbinafine, voriconazole, itraconazole, amphotericin B, fluconazole, and caspofungin against 16 strains of E. dermatitidis were determined by using CLSI broth microdilution method (M38-A2). The minimal fungicidal concentrations (MFCs) were also determined. Additionally, the interactions of caspofungin with voriconazole, amphotericin B, itraconazole or fluconazole, that of terbinafine with itraconazole, or that of fluconazole with amphotericin B were assessed by using the checkerboard technique. The fractional inhibitory concentration index (FICI) was used to categorize drug interactions as following, synergy, FICI ≤ 0.5; indifference, FICI > 0.5 and ≤4.0; or antagonism, FICI > 4.0. The MIC ranges of terbinafine, voriconazole, itraconazole, amphotericin B, fluconazole, and caspofungin against E. dermatitidis were 0.06–0.125 mg/l, 0.25–1.0 mg/l, 1.0–2.0 mg/l, 1.0–2.0 mg/l, 16–64 mg/l, and 32–64 mg/l, respectively. The in vitro interactions of caspofungin with voriconazole, amphotericin B, and itraconazole showed synergic effect against 10/16(62.5%), 15/16(93.75%), and 16/16(100%) isolates, while that of caspofungin with fluconazole showed indifference. Besides, the interaction of terbinafine with itraconazole as well as that of fluconazole with amphotericin B showed indifference. Terbinafine, voriconazole, itraconazole, and amphotericin B have good activity against E. dermatitidis. The combinations of caspofungin with voriconazole, amphotericin B or itraconazole present synergic activity against E. dermatitidis. These results provide the basis for novel options in treating various E. dermatitidis infections.     

Effects of Antifungal Agents in Sap Activity of <Emphasis Type="Italic">Candida albicans</Emphasis> Isolates

Some antifungal agents have shown to exert effects on expression of virulent factors of Candida as the production of secretory aspartyl proteinase (Sap). In this study, we sought to determine and to compare the influence of fluconazole and voriconazole in proteinase activity of this microorganism. Thirty-one isolates obtained from oral mucosa of human immunodeficiency virus positive (HIV⁺) patients were used in this study. The minimal inhibitory concentrations (MIC) of fluconazole and voriconazole were determined using the broth microdilution method with RPMI 1640 medium and with yeast carbon base–bovine serum albumin (YCB–BSA) medium. The Sap activity following by digestion of BSA as substrate was determined for four Candida albicans strains arbitrarily chosen according to susceptibility (susceptible or resistant) to fluconazole or voriconazole. Besides, the SAP1 to SAP7 genes were screened by PCR for the same isolates that were determined by the Sap activity. In vitro susceptibility testing using the two media presented similar MIC values. Increased Sap activity was observed in resistant isolates on presence of drugs, but the Sap activity by susceptible isolates to azoles showed different behavior on the presence of drug. We detected the presence of SAP1 to SAP7 genes from all susceptible or resistant C. albicans isolates. The present study provides important data about the proteinase activity and the presence of genes of SAP family in fluconazole and voriconazole susceptible or resistant C. albicans isolates.     

Candida albicans Elicits Pro-Inflammatory Differential Gene Expression in Intestinal Peyer’s Patches

The details of how gut-associated lymphoid tissues such as Peyer’s patches (PPs) in the small intestine play a role in immune surveillance, microbial differentiation and the mucosal barrier protection in response to fungal organisms such as Candida albicans are still unclear. We particularly focus on PPs as they are the immune sensors and inductive sites of the gut that influence inflammation and tolerance. We have previously demonstrated that CD11c⁺ phagocytes that include dendritic cells and macrophages are located in the sub-epithelial dome within PPs sample C. albicans. To gain insight on how specific cells within PPs sense and respond to the sampling of fungi, we gavaged naïve mice with C. albicans strains ATCC 18804 and SC5314 as well as Saccharomyces cerevisiae. We measured the differential gene expression of sorted CD45⁺ B220⁺ B-cells, CD3⁺ T-cells and CD11c⁺ DCs within the first 24 h post-gavage using nanostring nCounter^® technology. The results reveal that at 24 h, PP phagocytes were the cell type that displayed differential gene expression. These phagocytes were able to sample C. albicans and discriminate between strains. In particular, strain ATCC 18804 upregulated fungal-specific pro-inflammatory genes pertaining to innate and adaptive immune responses. Interestingly, PP CD11c⁺ phagocytes also differentially expressed genes in response to C. albicans that were important in the protection of the mucosal barrier. These results highlight that the mucosal barrier not only responds to C. albicans, but also aids in the protection of the host.