P1963 Effects of MG3290, a selective histone deacetylase inhibitor, on azole resistance in Candida glabrata

S564 P1961 Effects of vasoactive amines and albumin upon yeast susceptibility to antifungals S. Costa-de-Oliveira, A. Silva Dias, C. Pina-Vaz, A. Gon¸calves Rodrigues (Porto, PT) Critically-ill patients are often administered a variety of therapeutical drugs namely vasoactive amines and albumin. Following a prospective study about fungaemia conducted at our University hospital, several risk factors for invasive fungal disease and unfavourable outcome were detected. A significant correlation was found between fungaemia and those prescriptions. Previous studies showed the promotion of germination of A. fumigatus and the increase of antifungal resistance following incubation with albumin (Rodrigues et al, 2005). Objective: To evaluate the effect of albumine and vasoactive amines like adrenaline, noradrenaline, dopamine and dobutamine upon the susceptibility of Candida to several antifungals. Methods: 15 clinical strains (3 C. glabrata, 4 C. parapsilosis, 5 C. albicans and 3 C. tropicalis) plus 3 control strains belonging to American Type Culture Collection (ATCC) were studied. Susceptibility tests, according by the microdilution protocol M27-A2 from CLSI, were performed to amphotericin B, fluconazole, voriconazole and posaconazole; minimal inhibitory concentrations (MIC) were determined. MIC values were re-determined in the presence of 2% and 4% of albumin (Octapharma, Viena, Austria), 10, 25 and 100 mg/mL of adrenaline (Braun), 2.5, 5 and 100 mg/mL of noradrenaline (Braun) and 10, 50 and 200 mg/mL of both dobutamine (Sigma, Germany) and dopamine (Sigma). Results: All strains showed initial low MIC values to the antifungals tested. In the presence of 2% and 4% of albumin, MIC of all antifungal drugs increased from 2 to 10 dilutions. The MIC values of azoles increased from 2 to 4 dilutions in the presence of adrenaline and noroadrenaline. These effects were more consistent observed with the non-albicans strains tested. Conclusion: Vasoactive and inotropic amines have the potential to promote antifungal resistance when administered to patients simultaneously receiving antifungal therapy, especially in non-albicans species. Such fact may help to explain the increasing incidence of invasive fungal disease caused by such species among critical care patients. Reference(s) Rodrigues et al, Med Mycol 43: 711−7 2005. P1962 Comparison of Etest and microdilution method for antifungal susceptibility testing of C. neoformans to four azoles A.I. Aller, R. Claro, E. L´opez-Oviedo, A. Romero, E. Mart´ın-Mazuelos (Seville, ES) Objective: The purpose of this study is to compare the results obtained by the reference broth microdilution method (MDM) with those obtained with Etest system (ET) (Ab Biodisk, Solna, Sweden) for antifungal susceptibility testing of C. neoformans to fluconazole (Pfizer Central Research, UK), itraconazole (Janssen Pharmaceutica, UCA), voriconazole (Pfizer Central Research, UK) and posaconazole (SheringPlough Research Institute, USA). Methods: We have studied retrospectively 80 clinical isolates of C. neoformans. The susceptibility to four azoles were performed by the MDM according to the CLSI guidelines (M27-A2 document) modified by Ghannoum et al (J Clin Microbiol 1992; 30:2881−86). The final concentrations were of 0.12−64 mg/L for fluconazole and 0.015−8 mg/L for the rest of antifungal agents. The final inoculum size was 104 CFU/mL. The MICs were defined as the lowest concentration in which a prominent decrease in turbidity was observed. The ET was performed according to manufacturer’s instructions. The inoculum suspensions of C. neoformans isolates matched the turbidity of nº1 McFarland standard. The incubation time was 48−72 h. The MIC was read where the border of the elliptical inhibition zone intersected the scale on the antifungal strip. C. krusei ATCC 6258, C. parapsilosis

17th ECCMID / 25th ICC, Posters ATCC 22019, C. neoformans 90112 and C. neoformans 90113 were included as control strains. Essential agreement (EA) was defined as MIC results by ET and MDM in exact agreement or within two dilutions. Results: The MICs obtained by the two methods after different incubation times varied by no more than one two-fold dilutions. The MIC ranges, MIC50 and MIC90 obtained at 72 h are summarised in table 1. The EA was 75% for fluconazole, 63.4% for posaconazole, 30% for voriconazole and 6.3% for itraconazole. Table 1 Antifungal agent

Fluconazole Itraconazole Posaconazole Voriconazole

Method

MDM ET MDM ET MDM ET MDM ET

MIC (mg/L) Range

MIC50

MIC90

0.5−32 0.032–256 0.06−1 <0.002–0.25 0.03−4 <0.02−1 0.25−1 0.008−1.5

4 4 0.5 0.064 0.25 0.023 0.5 0.094

8 32 1 0.25 0.5 0.125 0.5 0.19

Conclusions: 1. Fluconazole MICs by ET showed good correlation with MDM (75%) 2. Itraconazole showed the lowest agreement (6.3%) 3. It would be necessary to carry out further studies including interlaboratory agreement and correlation of MICs by different methods with “in vivo” response. P1963 Effects of MG3290, a selective histone deacetylase inhibitor, on azole resistance in Candida glabrata B. Yeung, N. Georgopapadakou (Montreal, CA) Objectives: To examine whether MG3290, a highly active potentiator of azole antifungals against Aspergillus and Candida species, potentiates azoles against azole-resistant C. glabrata mutants and lowers resistance frequency when combined with azoles. Methods: Synergy of MG3290 with azoles was determined by twofold dilutions of each compound alone and in combination in 96-well plates (checkerboard format). Ergosterol synthesis was measured by the absorbance of lipid extracts at 281 nm; azole transport by the transport of Rhodamine123 (excitation max, 485 nm; emission max, 535 nm). Results: C. glabrata mutants resistant to itraconazole and fluconazole were detected at 3×10−7 and 5×10−6 frequencies respectively on agar plates containing 4×MIC of the relevant azole. Mutants resistant to itraconazole/MG3290 were detected at 1×10−7 frequency on plates containing 4×MIC of itraconazole/MG3290 (none at 2×MIC), while mutants resistant to fluconazole/MG3290, at 2×10−7 frequency at 4×MIC of fluconazole/MG3290. Three of seven itraconazole- and 13 of 14 fluconazole-resistant mutants isolated and examined had altered transport and were cross resistant. Basal ergosterol levels in the azole-resistant mutants were unchanged from the parent. MG3290, at
4mg/mL, lowered itraconazole MICs against itraconazole-resistant mutants and fluconazole MICs against fluconazole-resistant mutants. Conclusion: MG3290 potentiates azoles currently used in the clinic against resistant mutants and lowers resistance frequency when combined with azoles. It is therefore an attractive candidate for further development in combination with azoles. P1964 Enhancement of yeast susceptibility to antifungals J. Subik, J. Cernicka, Z. Kozovska, M. Valachovic, I. Hapala, G. Hajos (Bratislava, Ivanka pri Dunaji, SK; Budapest, HU) Objectives: Decreased susceptibilities to azole antifungals in Candida species were resulting from combination of several molecular mech-