We have compared the activities of posaconazole and other currently available antifungal agents against a collection of 3,378 clinical isolates of yeasts and filamentous fungi. A total of 1,997 clinical isolates of Candida spp., 359 of other yeast species, 697 strains of Aspergillus spp., and 325 nondermatophyte non- Aspergillus spp. were included. The average geometric means of the MICs of agents that were tested against Candida spp. were 0.23 μg/ml for amphotericin B, 0.29 μg/ml for flucytosine, 0.97 μg/ml for fluconazole, 0.07 μg/ml for itraconazole, 0.04 μg/ml for voriconazole, 0.15 μg/ml for caspofungin, and 0.03 μg/ml for posaconazole. Voriconazole and posaconazole were active in vitro against the majority of isolates, with resistance to fluconazole and itraconazole, and against Cryptococcus neoformans and other Basidiomycota yeasts. Posaconazole was the most active of antifungal agents tested against Aspergillus spp., with an average geometric mean of 0.10 μg/ml. It was active against Paecilomyces spp., Penicillium spp., Scedosporium apiospermum , and some black fungi, such as Alternaria spp. Multiresistant filamentous fungi, such as Scedosporium prolificans , Scopulariopsis brevicaulis , and Fusarium solani , were also resistant to voriconazole, caspofungin, and posaconazole. Amphotericin B and posaconazole were found to be active against most of the Mucorales strains tested. Posaconazole and currently available antifungal agents exhibit a potent activity in vitro against the majority of pathogenic fungal species.

The glucan synthase complex of the human pathogenic mold Aspergillus fumigatus has been investigated. The genes encoding the putative catalytic subunit Fks1p and four Rho proteins of A. fumigatus were cloned and sequenced. Sequence analysis showed that AfFks1p was a transmembrane protein very similar to other Fksp proteins in yeasts and in Aspergillus nidulans . Heterologous expression of the conserved internal hydrophilic domain of AfFks1p was achieved in Escherichia coli . Anti-Fks1p antibodies labeled the apex of the germ tube, as did aniline blue fluorochrome, which was specific for β(1–3) glucans, showing that AfFks1p colocalized with the newly synthesized β(1–3) glucans. AfRHO1 , the most homologous gene to RHO1 of Saccharomyces cerevisiae , was studied for the first time in a filamentous fungus. AfRho proteins have GTP binding and hydrolysis consensus sequences identical to those of yeast Rho proteins and have a slightly modified geranylation site in AfRho1p and AfRho3p. Purification of the glucan synthase complex by product entrapment led to the enrichment of four proteins: Fks1p, Rho1p, a 100-kDa protein homologous to a membrane H ⁺ -ATPase, and a 160-kDa protein which was labeled by an anti-β(1–3) glucan antibody and was homologous to ABC bacterial β(1–2) glucan transporters.

The commercial technique Vitek 2 system for antifungal susceptibility testing of yeast species was evaluated. A collection of 154 clinical yeast isolates, including amphotericin B- and azole-resistant organisms, was tested. Results were compared with those obtained by the reference procedures of both the CLSI and the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Two other commercial techniques approved for clinical use, the Etest and the Sensititre YeastOne, were included in the comparative exercise as well. The average essential agreement (EA) between the Vitek 2 system and the reference procedures was >95%, comparable with the average EAs observed between the reference procedures and the Sensititre YeastOne and Etest. The EA values were >97% forCandidaspp. and stood at 92% forCryptococcus neoformans. Intraclass correlation coefficients (ICC) between the commercial techniques and the reference procedures were statistically significant (P< 0.01). Percentages of very major errors were 2.6% between Vitek 2 and the EUCAST technique and 1.6% between Vitek 2 and the CLSI technique. The Vitek 2 MIC results were available after 14 to 18 h of incubation for allCandidaspp. (average time to reading, 15.5 h). The Vitek 2 system was shown to be a reliable technique to determine antifungal susceptibility testing of yeast species and a more rapid and easier alternative for clinical laboratories than the procedures developed by either the CLSI or EUCAST.

The role of Aspergillus fumigatus 14α-sterol demethylase (Cyp51A) in azole drug susceptibility was assessed. Targeted disruption of cyp51A in azole-susceptible and -resistant strains decreased MICs from 2- to 40-fold. The cyp51A mutants were morphologically indistinguishable from the wild-type strain, retaining the ability to cause pulmonary disease in neutropenic mice.

The antifungal susceptibility results for 32 clinical isolates ofScopulariopsis brevicaulisare presented. Flucytosine and itraconazole were inactive in vitro, and MICs of amphotericin B, voriconazole, and terbinafine for all isolates were high, with geometric means of 13, 25.8, and 14.4 μg/ml, respectively.