Rapid Antifungal Susceptibility Determination for Yeast Isolates by Use of Etest Performed Directly on Blood Samples from Patients with Fungemia

We prospectively determined the antifungal susceptibility of yeast isolates causing fungemia using the Etest on direct blood samples (195 prospectively collected and 133 laboratory prepared). We compared the Etest direct (24 h of incubation) with CLSI M27-A3 and the standard Etest methodologies for fluconazole, voriconazole, posaconazole, isavuconazole, caspofungin, and amphotericin B. Strains were classified as susceptible, resistant, or nonsusceptible using CLSI breakpoints (voriconazole breakpoints were used for posaconazole and isavuconazole). Categorical errors between Etest direct and CLSI M27-A3 for azoles were mostly minor. No errors were detected for caspofungin, and high percentages of major errors were detected for amphotericin B. For the azoles, false susceptibility (very major errors) was found in only two (0.6%) isolates (Candida tropicalis and C. glabrata). False resistance (major errors) was detected in 46 (14%) isolates for the three azoles (in 23 [7%] after excluding posaconazole). Etest direct of posaconazole yielded a higher number of major errors than the remaining azoles, especially for C. glabrata, Candida spp., and other yeasts. Excluding C. glabrata, Candida spp., and other yeasts, the remaining species did not yield major errors. Etest direct for fluconazole, voriconazole, isavuconazole, and caspofungin shows potential as an alternative to the CLSI M27-A3 procedure for performing rapid antifungal susceptibility tests on yeast isolates from patients with fungemia. Etest direct is a useful tool to screen for the presence of azole-resistant and caspofungin-nonsusceptible strains.The incidence of fungemia continues to rise in many institutions throughout the world, and Candida is one of the leading pathogens isolated from blood (15, 28). Amphotericin B and fluconazole have been widely used for the treatment of fungemia. However, newly licensed antifungal agents (voriconazole, posaconazole, and the echinocandins) and other azoles currently under investigation (isavuconazole) have expanded the antifungal armamentarium.The mortality rate of fungemia remains high (30%) and is clearly correlated with delayed initiation of effective antifungal therapy (11, 17). Antifungal therapy is considered inappropriate when it is omitted, when the agent administered has no antifungal activity against the infecting organism, or when its serum concentrations are subtherapeutic.A growing proportion of Candida isolates obtained from blood samples have reduced antifungal susceptibility to fluconazole and other antifungal agents (14, 25). Patients with candidemia caused by Candida strains with high MICs for fluconazole or voriconazole and echinocandins can have a worse prognosis (22-24). Consequently, systematic use of empirical antifungal agents with broad-spectrum in vitro activity has led to considerable increases in the number of adverse events and in the cost of treatment (2).The combination of an increasing number of antifungal-resistant isolates and the cost of the new antifungal agents makes antifungal susceptibility testing a necessity. The reference antifungal susceptibility testing method for yeasts is the Clinical and Laboratory Standards Institute (CLSI; formerly NCCLS) testing standard M27-A3. However, this method requires pure-culture isolates, and results of antifungal susceptibility testing are not available until 48 to 72 h after the isolation of fungi in blood.The Etest performed directly on blood samples may expedite antifungal testing and provide results in 24 h. Our group has previously demonstrated that the Etest performed directly on samples from the lower respiratory tract is a rapid and accurate procedure for antimicrobial susceptibility testing of bacteria in patients with ventilator-associated pneumonia (4, 5). We compared the results of the Etest performed directly on positive blood cultures with yeasts grown in Bactec blood bottles with the results of CLSI M27-A3 in isolates from patients with fungemia and blood samples generated from previously characterized isolates.(This study was partially presented at the 20th Conference of the European Congress of Clinical Microbiology and Infectious Diseases [ECCMID] in Vienna, Austria, 2010 [abstract no. P-838] [13a].)