The activities of fluconazole and voriconazole against isolates of Candida spp. (n ‫؍‬ 400) were tested by the E-test, disk diffusion, and the National Committee for Clinical Laboratory Standards (NCCLS) M27-A2 broth microdilution-based reference methods. More than 96% of isolates found to be susceptible to fluconazole by the reference method were identified as susceptible by the agar-based methods. Lesser degrees of correlation with the reference method were seen for isolates identified as resistant by the agar-based methods. Interpretive categories are not available for voriconazole, but results qualitatively similar to those for fluconazole were seen. The agar-based E-test and disk diffusion methods are reliable alternatives to the NCCLS M27-A2 reference microdilution method for isolates that test susceptible to fluconazole.The development of standardized antifungal susceptibility testing methods has been the subject of numerous studies during the last decade. Reference methods for yeasts (the National Committee for Clinical Laboratory Standards [NCCLS] M27-A2 method) and molds (the NCCLS M38-A method) are now available (16). Agar-based susceptibility testing methods have been a focus of interest for many researchers and include the classical disk diffusion (DD) methods and the E-test (ET) method (3, 6-10, 13, 14, 16-18). Those tests are very attractive due to their simplicity, reproducibility, and lack of requirements for specialized equipment (11,16). Recent studies have documented comparable results between those methods and the results of standard reference broth microdilution (MD) susceptibility testing (7,11,13).In this study, we compared the NCCLS M27-A2 MD method with the ET and DD methods for determination of the susceptibilities of 400 Candida species isolates to fluconazole and voriconazole. The ET and DD methods are well studied for fluconazole (3,(6)(7)(8)(9)(10)(11)14), and this work extends their usage to include voriconazole. MATERIALS AND METHODSIsolates. Four hundred bloodstream isolates of Candida species were randomly selected for testing. These included 205 isolates of Candida albicans, 56 isolates of C. tropicalis, 39 isolates of C. glabrata, 66 isolates of C. parapsilosis, 24 isolates of C. krusei, and 10 isolates of other species. The isolates were identified with the API 20C AUX system (Biomerieux Vitek, Hazelwood, Mo.) and were subsequently stored in sterile distilled water at room temperature until susceptibility tests were performed. Each isolate was subcultured at least twice on Sabouraud dextrose agar and incubated at 35°C prior to testing to ensure purity and optimal growth.Inoculum suspensions. Yeast inoculum suspensions were prepared as described for the NCCLS M27-A2 method (12). The turbidity was measured with a spectrophotometer at 530 nm and was adjusted to match a 0.5 McFarland density standard, resulting in a concentration of 1 ϫ 10 6 to 5 ϫ 10 6 yeast cells/ml. This inoculum was used directly for inoculation of agar plates (see below) or was diluted as needed for the MD ...

The in vitro activities of amphotericin B, itraconazole, fluconazole, voriconazole, posaconazole, and ravuconazole against 39 isolates of Trichosporon spp. were determined by the NCCLS M27-A microdilution method. The azoles tested appeared to be more potent than amphotericin B. Low minimal fungicidal concentration/MIC ratios were observed for voriconazole, posaconazole, and ravuconazole, suggesting fungicidal activity.

(1-->3)-beta-d-glucan is a well known cell wall constituent of fungal isolates that can be detected by assays in vivo and in vitro. Previous studies have shown that different fungal isolates may show different levels of reactivity with an assay for beta glucan. In this study we evaluated the in vitro reactivity of 127 clinical fungal isolates belonging to 40 different genera, with the Glucatell assay. The majority of the fungal isolates released high levels of beta glucan. Beta glucan test reactivity appears to be species-specific and this may reflect the beta glucan content of the organism.

In this study, we evaluated the in vitro activity of anidulafungin against selected mold isolates. Anidulafungin showed promising activity against Bipolaris spicifera, Exophiala jeanselmei, Fonsecaea pedrosoi, Madurella mycetomatis, Penicillium marneffei, Phialophora verrucosa, Pseudallescheria boydii, Sporothrix schenckii, and Wangiella dermatitidis.The incidence of invasive fungal infections has increased in the past two decades. Candida species and Aspergillus species are the most common etiologic agents causing invasive fungal infections. Although rare overall, Fusarium species are the second most common mold isolate after Aspergillus species (14,15). Some other less common filamentous fungi, like Penicillium, Bipolaris, Pseudallescheria, and Scedosporium species, have also been emerging as causative agents of opportunistic infections in immunocompromised patients (7,14,24,26,31).Amphotericin B has been historically accepted as the "gold standard" for the treatment of most fungal infections, although it is known to have poor outcomes in immunocompromised patients with severe mold infections (13). Alternative therapeutic agents, like new azoles and echinocandins, are under clinical evaluation (18). The relatively recent approvals of caspofungin for aspergillosis and voriconazole for aspergillosis, fusariosis, and scedosporidiasis have revolutionized the field of antifungal therapy (2, 4, 5, 8-11, 19, 29, 30).Anidulafungin is an echinocandin with excellent in vivo and in vitro activities against Candida spp. and Aspergillus spp. (17, 20-23, 27, 28). The in vitro activity of anidulafungin against less common but clinically emerging filamentous fungi has been evaluated in a limited number of studies (6,22,25). We sought to evaluate its in vitro activity against selected mold isolates in comparison with the activities of voriconazole and amphotericin B.A collection of 74 clinical mold isolates (Table 1) were tested. Isolates were obtained from the Department of Pathology, University of Texas, Medical Branch, Galveston. All isolates were stored at Ϫ80°C, and each isolate was subcultured on potato dextrose agar slants (Becton, Dickinson and Company, Sparks, Md.) at least twice to ensure purity and viability. The quality control strains were Candida parapsilosis ATCC 22019 and Candida krusei ATCC 6258.Anidulafungin (Vicuron Pharmaceuticals, Inc., King of Prussia, Pa.), voriconazole (Pfizer Pharmaceutical Group, New York, N.Y.), and amphotericin B (Bristol-Myers Squibb, Wallingford, Conn.) were dissolved in 100% dimethyl sulfoxide (Fisher Chemicals, Fair Lawn, N.J.) and then were further diluted (1:50) in 2ϫ RPMI 1640 medium (Sigma Chemical Company, St. Louis, Mo.) buffered to pH 7.0 with 0.165 M morpholinepropanesulfonic acid (MOPS) buffer according to the recommendations of National Committee for Clinical Laboratory Standards (NCCLS) approved standard M38-A (16) to yield two times the final strength required for the test. Amphotericin B was diluted 1:50 with 2ϫ antibiotic medium 3 (AM3; BBL, Cockeysville, Md.) buf...

The in vitro interactions of anidulafungin with itraconazole, voriconazole, and amphotericin B were evaluated by using the checkerboard method. For Aspergillus spp., anidulafungin with amphotericin B showed indifference for 16/26 isolates, while anidulafungin with either azole showed a synergy trend for 18/26 isolates. All drug combinations showed indifference for 7/7 Fusarium sp. isolates.Invasive fungal infections due to molds are becoming more prevalent in immunocompromised patients (21). Among the invasive mold infections, Aspergillus spp. and Fusarium spp. are particularly challenging to manage, due to aggressive courses and high mortality (11,20).Since 1959, amphotericin B deoxycholate (AMBD) had been considered the "gold standard" for the treatment of fungal infections. However, due to high failure rates and significant toxicity (6), other agents are being explored today both singly and in combination therapy. Among the azoles, itraconazole (ITR) continues to show some promise against Aspergillus spp. (3). Voriconazole (VOR), a novel azole (11), is perhaps the current "gold standard" for the treatment of invasive aspergillosis, although success rates are still less than optimal (5, 8). The echinocandins (caspofungin, anidulafungin [ANID], and micafungin) inhibit 1,3-␤-D-glucan synthesis and have in vitro and in vivo activity against Candida and Aspergillus spp. (4,16,17). In the clinical setting, caspofungin appears to be at least as effective as AMBD for salvage therapy of invasive aspergillosis compared to historical controls (13).Due to the high mortality and lack of an ideal drug for these diseases, combination therapy has been an attractive possibility that has recently received much attention in medical mycology. Early studies have shown in vitro and in vivo advantages of several combinations. Arikan et al. (2) showed additive to synergistic effects of caspofungin with AMBD in vitro against Aspergillus and Fusarium spp. In a guinea pig model, colony counts of Aspergillus spp. and the number of culture-positive tissues were reduced after treatment with VOR and caspofungin compared with either of the agents alone (10). Similarly, Petraitis et al. showed that the combination of micafungin and ravuconazole in a rabbit model had synergistic effects against invasive aspergillosis (18).The purpose of this study was to evaluate the in vitro interactions of ITR, VOR, and AMBD with ANID against Aspergillus spp. and Fusarium spp. as preliminary work to support further in vivo and clinical research on these combinations.Isolates. Twenty-six clinical isolates of Aspergillus spp. and seven clinical isolates of Fusarium spp. were used. The species distribution was as follows: eight isolates of Aspergillus flavus,