In vitro and ex vivo biofilms of dermatophytes: a new panorama for the study of antifungal drugs

Abstract

This study describes an ex vivo model that creates an environment for dermatophyte biofilm growth, with features that resemble those of in vivo conditions, designing a new panorama for the study of antifungal susceptibility. Regarding planktonic susceptibility, MIC ranges were 0.125-1?µg ml^?1 for griseofulvin and 0.000097-0.25?µg ml^?1 for itraconazole and terbinafine. sMIC50 ranges were 2->512?µg ml^?1 for griseofulvin and 0.25->64?µg ml^?1 for itraconazole and terbinafine. CLSM images demonstrated a reduction in the amount of cells within the biofilm, but hyphae and conidia were still observed and biofilm biomass was maintained. SEM analysis demonstrated a retraction in the biofilm matrix, but fungal structures and water channels were preserved. These results show that ex vivo biofilms are more tolerant to antifungal drugs than in vitro biofilms, suggesting that environmental and nutritional conditions created by this ex vivo model favor biofilm growth and robustness, and hence drug tolerance.     

Comparative study of in vitro methods to analyse the antifungal activity of propolis against yeasts isolated from patients with superficial mycoses

AIM: To test a total of 15 strains belonging to four species of yeasts by different in vitro methods against propolis and itraconazole (ITC). METHODS AND RESULTS: Three methods were compared for susceptibility testing of yeast isolates to propolis: disc diffusion method, agar dilution method and National Committee for Clinical Laboratory Standards (NCCLS, M27A) broth microdilution method. ITC was selected as the antifungal agent for comparison study. Using the broth microdilution method, the geometric mean for MIC (microg ml(-1)) with regard to all isolates was < or =0.06 for propolis and < or =0.35 for ITC. The broth microdilution and the agar dilution methods were in good agreement (75%) for propolis against yeasts isolated from patients with superficial mycoses. Using the diffusion method, all strains showed a broad zone of inhibition at the first available reading time (24 or 48 h). An increase of MIC values was accompanied by a decrease of growth inhibition zone diameter. A favourable correlation was found between MIC and inhibition zone around the disc for propolis sample and the correlation coefficient was: r = -0.626 (P < 0.01). CONCLUSIONS: This study suggests the potential value of the agar dilution and disc diffusion method as a convenient alternative method for testing of yeasts to propolis. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated that propolis and ITC were very active against yeasts from patients with superficial mycoses. The other prominent finding in this study is that RPMI 1640 with L-glutamine was the available broth for the in vitro susceptibility testing of yeasts.     

Evaluation of a new method for antifungal susceptibility testing for azoles

The interpretation of the end points in azole antifungal drug susceptibility testing is challenging, in part due to incomplete growth inhibition of Candida species. Since the reference Clinical and Laboratory Standards Institute (CLSI) broth microdilution method have limitation with azoles, a new modification of the CLSI microdilution protocol was evaluated. We measure the decrease in growth rate (μ) of exponentially growing cultures in accordance to different azole concentrations at time intervals up to 10 h. Using 15 different Candida strains, an overall agreement within ± 2 dilutions by the CLSI method at 24 h in RPMI and the μ-dependent method for three antifungal agents (fluconazole- itraconazole and voriconazole) was achieved. MIC measurement by the new method was less sensitive to the medium used or the inoculum size applied. The presented data suggested that, measuring the in vitro inhibition kinetics at the logarithmic phase could have advantages for addressing susceptibility testing toward azoles.     

In vitro susceptibility of dermatomycoses agents to six antifungal drugs and evaluation by fractional inhibitory concentration index of combined effects of amorolfine and itraconazole in dermatophytes

To investigate the antifungal drug susceptibility of fungi responsible for dermatomycoses, minimum inhibition concentration (MIC) tests were performed in 44 strains of dermatophytes, including Trichophyton rubrum, Trichophyton mentagrophytes, Trichophyton verrucosum, Trichophyton tonsurans, Microsporum canis, Microsporum gypseum and Epidermophyton floccosum, with six antifungal drugs (amorolfine, terbinafine, butenafine, ketoconazole, itraconazole and bifonazole) by broth microdilution assay according to Clinical Laboratory Standard Institute protocols. Six possible dermatomycosis‐causing non‐dermatophytic fungi were also tested. The two major causes of tinea, T. rubrum and T. mentagrophytes, showed significantly different sensitivities to ketoconazole and bifonazole. Clinically derived dermatophytes were sensitive to the six antifungal drugs tested. However, non‐dermatophytes, especially Fusarium spp., tended to be resistant to these antifungal drugs. In Trichophyton spp., the MICs of non‐azole drugs had narrower distributions than those of azoles. To evaluate the effects of antifungal drug combinations, the fractional inhibitory concentration index was calculated for the combination of amorolfine and itraconazole as representative external and internal drugs for dermatophytes. It was found that this combination had synergistic or additive effects on most dermatophytes, and had no antagonistic effects. The variation in susceptibility of clinically derived fungal isolates indicates that identification of causative fungi is indispensable for appropriately choosing effective antifungal drugs in the early stages of infection. The results of combination assay suggest that multiple drugs with different antifungal mechanisms against growth of dermatophytes should be used to treat refractory dermatomycoses, especially onychomycosis.     

In vitro susceptibility of dematiaceous fungi to ten antifungal drugs using an agar diffusion test

We assessed the usefulness of an agar diffusion method, NeoSensitabs, to determine in vitro sensitivity of 52 isolates of dematiaceous filamentous fungi against ten antifungal agents: amphotericin B, 5-fluorocytosine, ketoconazole, fluconazole, itraconazole, terbinafine, bifonazole, miconazole, clotrimazole, and griseofulvin. For the preparation of the inoculum, a spectrophotometric method including both Shadomy and Casitone agar (CAS) culture media was used. Dematiaceous filamentous fungi were sensitive to itraconazole, terbinafine and bifonazole. Ketoconazole (90.4%), miconazole (71%), and clotrimazole (46%) showed a variable susceptibility pattern. Most species were resistant to griseofulvin and fluconazole (96%). All isolates were resistant to 5-fluorocytosine. Sixty-three percent of strains were susceptible to amphotericin B and 28.8% resistant. Inhibition zones in the antifungal susceptibility testing did not vary according to culture medium, although fungal growth was better in CAS. Variations in antifungal sensitivity in Exophiala spinifera and Fonsecaea pedrosoi spp. would justify an in vitro susceptibility study when indicating antifungal therapy. These results show that NeoSensitabs agar diffusion method is simple, rapid, and low-cost and can be available to many clinical laboratories for the study of in vitro sensitivity of dematiceous moulds.     

Potentiation of antifungal activity of terbinafine by dihydrojasmone and terpinolene against dermatophytes

Dermatophytoses are infections that affect keratinized tissues. Their main etiologic agents are fungi of the genera Microsporum and Trichophyton. The emergence of resistant fungi and the clinical relevance of dermatophytosis have encouraged studies that aim to increase the arsenal of drugs or act on mechanisms that confer multiple drug resistance. This study investigated the modulating activity of terbinafine promoted by dihydrojasmone and terpinolene against Microsporum canis LM 216, Trichophyton interdigitale H6 and T. interdigitale Δmdr2. The minimum inhibitory concentration (MIC) of test drugs was determined by broth microdilution. The effect of the drugs tested on plasma membrane functionality was analysed. Terbinafine MIC was determined in sub-inhibitory concentrations of monoterpenes. Finally, it was performed an association study with terbinafine and monoterpenes. Dihydrojasmone presented lower MIC values than terpinolene. All fungi were sensitive to terbinafine, starting at 1 μg ml⁻¹. All tested drugs increased K⁺ release (P < 0·05), affecting the functionality of the plasma membrane. Dihydrojasmone modulated the sensitivity of all strains against terbinafine, and terpinolene modulated the sensitivity of M. canis LM 216 and T. interdigitale Δmdr2. The monoterpenes and terbinafine drug associations presented synergism. In conclusion, the results suggest that the dihydrojasmone and terpinolene are promising antifungal agents that potentiate the antifungal activity of terbinafine against dermatophytes.     

Susceptibility Screening of Hyphae-Forming Fungi with a New, Easy, and Fast Inoculum Preparation Method

In vitro susceptibility testing of clinically important fungi becomes more and more essential due to the rising number of fungal infections in patients with impaired immune system. Existing standardized microbroth dilution methods for in vitro testing of molds (CLSI, EUCAST) are not intended for routine testing. These methods are very time-consuming and dependent on sporulating of hyphomycetes. In this multicentre study, a new (independent of sporulation) inoculum preparation method (containing a mixture of vegetative cells, hyphae, and conidia) was evaluated. Minimal inhibitory concentrations (MIC) of amphotericin B, posaconazole, and voriconazole of 180 molds were determined with two different culture media (YST and RPMI 1640) according to the DIN (Deutsches Institut für Normung) microdilution assay. 24 and 48?h MIC of quality control strains, tested per each test run, prepared with the new inoculum method were in the range of DIN. YST and RPMI 1640 media showed similar MIC distributions for all molds tested. MIC readings at 48 versus 24?h yield 1 log₂ higher MIC values and more than 90?% of the MICs read at 24 and 48?h were within ±2 log₂ dilution. MIC end point reading (log_{2 MIC-RPMI 1640}?log_{2 MIC-YST}) of both media demonstrated a tendency to slightly lower MICs with RPMI 1640 medium. This study reports the results of a new, time–saving, and easy-to-perform method for inoculum preparation for routine susceptibility testing that can be applied for all types of spore-/non-spore and hyphae-forming fungi.     

Evaluation of a Scanner-Assisted Colorimetric MIC Method for Susceptibility Testing of Gram-Negative Fermentative Bacteria

We describe the ScanMIC method, a colorimetric MIC method for susceptibility testing of gram-negative fermentative bacteria. The method is a slight modification of the National Committee for Clinical Laboratory Standards (NCCLS) recommended broth microdilution method that uses a redox indicator 2,3,5-triphenyltetrazolium chloride (TTC) to enhance the estimate of bacterial growth inhibition in a microplate and a flatbed scanner to capture the microplate image. In-house software was developed to transform the microplate image into numerical values based on the amount of bacterial growth and to generate the MICs automatically. The choice of indicator was based on its low toxicity and ease of reading by scanner. We compared the ScanMIC method to the NCCLS recommended broth microdilution method with 197 coliform strains against seven antibacterial agents. The interpretative categorical agreement was obtained in 92.4% of the assays, and the agreement for MIC differences (within ±1 log₂ dilution) was obtained in 96% for ScanMIC versus broth microdilution and 97% for a two-step incubation colorimetric broth microdilution versus the broth microdilution method. The method was found to be labor-saving, not to require any initial investment, and to show reliable results. Thus, the ScanMIC method could be useful for epidemiological surveys that include susceptibility testing of bacteria.     

Evaluation of a scanner-assisted colorimetric MIC method for susceptibility testing of gram-negative fermentative bacteria

We describe the ScanMIC method, a colorimetric MIC method for susceptibility testing of gram-negative fermentative bacteria. The method is a slight modification of the National Committee for Clinical Laboratory Standards (NCCLS) recommended broth microdilution method that uses a redox indicator 2,3,5-triphenyltetrazolium chloride (TTC) to enhance the estimate of bacterial growth inhibition in a microplate and a flatbed scanner to capture the microplate image. In-house software was developed to transform the microplate image into numerical values based on the amount of bacterial growth and to generate the MICs automatically. The choice of indicator was based on its low toxicity and ease of reading by scanner. We compared the ScanMIC method to the NCCLS recommended broth microdilution method with 197 coliform strains against seven antibacterial agents. The interpretative categorical agreement was obtained in 92.4% of the assays, and the agreement for MIC differences (within +/-1 log(2) dilution) was obtained in 96% for ScanMIC versus broth microdilution and 97% for a two-step incubation colorimetric broth microdilution versus the broth microdilution method. The method was found to be labor-saving, not to require any initial investment, and to show reliable results. Thus, the ScanMIC method could be useful for epidemiological surveys that include susceptibility testing of bacteria.     

Determination of susceptibility/resistance to antifungal drugs of Trichophyton mentagrophytes isolates by a macrodilution method

Onychomycosis is a common adult human mycosis, and dermatophytes of the Trichophyton genera are the most frequently isolated microorganism. Globally, from 3% to 10% of the human population is attacked by ony cho mycosis, and many cases involve toenails. The aim of this work was to determine the minimal inhibitory concentrations (MICs) of antifungal drugs (fluconazole, ketoconazole, itraconazole, terbinafine, and griseofulvin) often used for the treatment of ungueal dermatophytosis caused by Trichophyton mentagrophytes. The MICs were determined by the broth medium macrodilution method. The results showed that activities of terbinafine and itraconazole were significantly higher (MIC <0.007-0.015 microg.mL -1 and MIC = 0.062-1.000 microg.mL -1, respectively). All isolates had reduced susceptibility to fluconazole (MIC = 16 to >64 microg.mL -1). The MICs of ketoconazole and griseofulvin varied among strains, ranging from 0.125 to 2.000 microg.mL -1 for ketoconazole and from 0.25 to 2.00 microg.mL -1 for griseofulvin. These MICs were higher than those of other studies cited, possibly because of differences in culture medium used in the other studies.     

Occurrence of Dermatomycoses and In-Vitro Therapeutic Efficacy of Three Antifungal Drugs on the Growth of Epidermophyton Floccosum

The occurrence of dermatomycoses and the in-vitro therapeutic efficacy of some antifungal agents on dermatomycotic organisms were investigated. Of the 550 primary school children screened, the incidence was one hundred (18%), 70 were males (representing 20% of the males screened) and 30 females (15% of the females sampled). The differences between male and female prevalence were insignificant. Three species of dermatophytes were isolated and identified. These were Microsporum canis, Trichophyton tonsurans and Epidermophyton floccosum. The antifungal agents tested on E. floccosum were griseofulvin, terbinafine and ketoconazole. They produced different sized zones of inhibition against the growth of E. floccosum. Griseofulvin exhibited a 50% inhibition of growth on E. floccosum at 63.00 mg/L. Terbinafine on the other hand exhibited varying levels of inhibition of growth at varying concentrations, at 0.07 mg/L, terbinafine achieved 46% inhibition of growth on E. floccosum. The drug achieved 100% inhibition of growth on the isolate at 61.81 mg/L. In the case of ketoconazole, 50% inhibition of growth was achieved at 100 mg/L while 100% inhibition of growth was achieved at 200 mg/L. The antifungal effects of the three drugs were confirmed by broth dilution tests where terbinafine was found to be fungistatic on the growth of E. floccosum at concentrations ranging from 0.013-1.700 mg/L and was fungicidal at concentrations ranging from 0.027-1.700 mg/L. Ketoconazole was found to inhibit the growth of E. floccosum at 0.003-1.700 mg/L and was fungicidal at concentrations ranging from 0.027-1.700 mg/L. It however did not succeed in killing the isolate under the same range of concentrations. Griseofulvin exhibited fungistatic effects on the growth of E. floccusum at 0.013-1.700 mg/L. In conclusion, ketoconazole and griseofulvin were found to be fungistatic and not fungicidal while terbinafine was both fungistatic and fungicidal on the pathogen. Terbinafine was found to be the most effective drug in inhibiting E. floccosum.     

Influence of incubation time in the in vitro antifungal activity of terbinafine against Trichophyton rubrum.

Antifungal susceptibility tests are influenced by a number of technical variables, including inoculum size, temperature, medium formulation and duration of incubation. In this study, we have compared the in vitro susceptibility of 20 strains de Trichophyton rubrum against clotrimazole and terbinafine, and studied the influence of incubation time on MICs of both drugs. The assay was performed by agar dilution, the medium used was Saboraud glucose agar without an antibiotic. The MIC was evaluated at 15, 30 and 45 days' incubation. The MICs ranges of terbinafine were 0.002 to 0.0975 microg/ml, 0.0975 to 0.39 microg/ml and 0.195 to 0.39 microg/ml at 15, 30 and 45 days' incubation, respectively. The MICs ranges of clotrimazole at 15, 30 and 45 days' incubation were 3.125 to 50 microg/ml. T. rubrum was markedly more susceptible to terbinafine than to clotrimazole (p<0.001). In addition, we observed that an increase of incubation time causing an increase in the MIC value of terbinafine (p<0.001), but MIC values for clotrimazole remained constant with time (p=0.464). In conclusion, the MIC is dependent on reading time and the antifungal compound.     

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.     

Evaluation of <Emphasis Type="Italic">Fusarium solani</Emphasis> Hyphae and Conidia Susceptibility to Amphotericin B and Itraconazole: Study of a Clinical Case

Fusarium species are hyaline moulds belonging to the hyalohyphomycosis group that are usually found in the soil and plants. This organism has emerged as a cause of disseminated invasive disease. The correlation between in vitro value and clinical efficacy is low and many patients remain unresponsive to treatment despite in vitro susceptibility. We determined growth control for Fusarium solani using the BioCell-Tracer system that measures the growth rate of a single fungal hypha, and the effect of different concentrations of amphotericin B and itraconazole. The MIC for these two drugs was also determined by a broth microdilution technique, using RPMI 1640. Different MICs for amphotericin B were obtained by the two different methods. This paper describes a case of infection due to Fusarium solani in an allogeneic bone marrow transplanted patient, the microbiological diagnostic, antifungal susceptibility tests for conidia and hypha and clinical correlation.     

In vitro activity of the echinocandins. How should it be evaluated?

Echinocandins are a novel class of antifungal drugs. They have good activity against Candida spp and Aspergillus spp. Their low selective toxicity allows their administration at high doses with few secondary side effects. We have reviewed the available data on the endpoints for these drugs in their in vitro susceptibility testing on yeasts and moulds. The microdilution broth method is the most commonly used technique and MIC-1 (80% of growth inhibition) seems to be the most reliable endpoint when yeasts are tested. This endpoint also seems to be the most appropriate for the different drugs when they are combined with echinocandins using the checkerboard method for testing yeasts. By contrast, in the case of moulds, the minimum effective concentration (MEC) correlates better with the in vivo activity than the MIC when echinocandins are tested, and when these drugs are combined with other antifungals, MIC-2 (50% of growth inhibition) seems the most appropriate endpoint. Criteria based on drug pharmacodynamics is the most useful to define the echinocandin endpoints that best correlate with their in vivo efficacy.     

Microbiological Characteristics of Medically Important <Emphasis Type="Italic">Trichosporon</Emphasis> Species

Trichosporon species are opportunistic pathogens associated with a high mortality rate in immunocompromised patients. Disseminated trichosporonosis is uncommon but reports are increasing. In this study, using 16 stock clinical isolates of suspected Trichosporon species and 4 known Trichosporon strains, we investigated the morphology, physio-biochemistry, molecular biology and antifungal susceptibility characteristics of these Trichosporon spp. and discovered that ITS sequence-based identification is a rapid and accurate identification alternative to most phenotypic or physio- biochemical methods. In vitro antifungal susceptibility tests showed high amphotericin B, itraconazole and terbinafine MIC value in these Trichosporon strains.     

Evaluation of diffusion and dilution methods to determine the antibacterial activity of plant extracts

The aim of this study was to evaluate diffusion and dilution methods for determining the antibacterial activity of plant extracts and their mixtures. Several methods for measurement of the minimal inhibitory concentration (MIC) of a plant extract are available, but there is no standard procedure as there is for antibiotics. We tested different plant extracts, their mixtures and phenolic acids on selected gram-positive (Staphylococcus aureus, Bacillus cereus, and Listeria monocytogenes) and gram-negative bacteria (Escherichia coli O157:H7, Salmonella Infantis, Campylobacter jejuni, Campylobacter coli) with the disk diffusion, agar dilution, broth microdilution and macrodilution methods. The disk diffusion method was appropriate only as a preliminary screening test prior to quantitative MIC determination with dilution methods. A comparison of the results for MIC obtained by agar dilution and broth microdilution was possible only for gram-positive bacteria, and indicated the latter as the most accurate way of assessing the antimicrobial effect. The microdilution method with TTC (2,3,5-triphenyl tetrazolium chloride) or INT (2-p-iodophenyl-3-p-nitrophenyl-5-phenyl tetrazolium chloride) to indicate the viability of aerobic bacteria was found to be the best alternative approach, while only ATP determination was appropriate for microaerophilic Campylobacter spp. Using survival curves the kinetics of bacterial inactivation on plant extract exposure was followed for 24 h and in this way the MIC values determined by the microdilution method were confirmed as the concentrations of extracts that inhibited bacterial growth. We suggest evaluation of the antibacterial activity of plant extracts using the broth microdilution method as a fast screening method for MIC determination and the macrodilution method at selected MIC values to confirm bacterial inactivation. Campylobacter spp. showed a similar sensitivity to plant extracts as the tested gram-positive bacteria, but S. Infantis and E. coli O157:H7 were more resistant.     

Colorimetric broth microdilution method for the antifungal screening of plant extracts against yeasts

Screening plant extracts for antifungal activity is increasing due to demand for new antifungal agents, but the testing methods present many challenges. Standard broth microdilution methods for antifungal susceptibility testing of available antifungal agents are available now, but these methods are optimised for single compounds instead of crude plant extracts. In this study we evaluated the standard NCCLS method as well as a modification which uses spectrophotometric determination of the end-points with a plate reader. We also evaluated another standard method, the EUCAST method, which is a similar microdilution assay to the NCCLS method, but uses a larger inoculum size and a higher glucose concentration in the medium as well as spectrophotometric end-point determination. The results showed that all three methods had some drawbacks for testing plant extracts and thus we modified the NCCLS broth microdilution method by including a colorimetric indicator-resazurin for end-point determination. This modified method showed good reproducibility and clear-cut end-point, plus the end-point determination needed no instruments. It enabled us to evaluate the activity of a selection of extracts from six Combretaceous plants against three Candida spp. and thus provided pharmacological evidence for some traditional uses of these plants while assisting the identification of the active ingredients.     

Development of Reference Procedures for Broth Microdilution Antifungal Susceptibility Testing of Yeasts with Standardized Endpoint Determination

Standard guidelines for the broth microdilution antifungal susceptibility testing of amphotericin B, flucytosine, fluconazole, miconazole and itraconazole are reported. These are a modification of the method developed by the National Committee for Clinical Laboratory Standards (NCCLS) on the following two points: standardization of the means of endpoint determination and the inclusion of miconazole and itraconazole in the testing. MIC was determined to be when the positive control had a turbidity of 0.2 at the 630 nm wavelength. The endpoint was 80% inhibition for azoles and 100% inhibition for other drugs. The method provided good reproducibility, and a wide range of MIC distribution was observed in all antifungal agents except amphotericin B.     

男性尿道炎和包皮龟头炎致病真菌的分布与药敏分析

目的了解男性念珠菌性尿道炎和包皮龟头炎的菌群分布及体外抗真菌药敏试验情况。方法菌株分离均来自复旦大学附属华山医院皮肤性病门诊临床症状轻重不一、真菌直接镜检阳性的61例患者。用科玛嘉念珠菌显色培养基及API 20C AUX鉴定系统进行菌种鉴定;采用CLSIM27-A2肉汤微量稀释法对61株临床分离念珠菌作了氟康唑、两性霉素B、氟胞嘧啶、伊曲康唑、伏立康唑、特比萘芬6种抗真菌药物敏感性测定。结果对培养阳性的61例菌株,通过科玛嘉念珠菌显色培养基及API 20C AUX鉴定系统作菌种鉴定,白念珠菌52例（85.2%）,近平滑念珠菌3例,光滑念珠菌2例,热带念珠菌2例,季也蒙念珠菌1例,克柔念珠菌1例。对52株白念珠菌的药敏试验显示氟康唑98.1%敏感,1.9%剂量依赖性敏感;氟胞嘧啶96.2%敏感,3.8%耐药;伊曲康唑44.2%敏感,40.5%剂量依赖性敏感,15.3%耐药;伏立康唑84.6%敏感,15.4%耐药;两性霉素B全部敏感;特比萘芬的MIC范围为1-64μg/ml,MIC50和MIC90皆为64μg/ml。结论在男性念珠菌性尿道炎和包皮龟头炎中,白念珠菌仍是第一位致病菌,体外药敏试验显示氟康唑、伏立康唑、氟胞嘧啶、两性霉素B对男性念珠菌性尿道炎均有较好的敏感性。