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DOI: 10.1093/jac/dkg018
1Ctedra
de Medicina Preventiva y Salud Pblica, Facultad de Medicina, Universidad de la Laguna,
38071 La Laguna, Tenerife; 2Departamento de Microbiologa, ACIA, Barcelona;
3Hospital Ntra. Sra. de Candelaria, Tenerife; 4Departamento de Immunologa, Microbiologa y Parasitologa,
Facultad de Medicina, Universidad del Pas Vasco, Bilbao, Spain; 5Medical College of Virginia,
Campus Virginia Commonwealth University, Richmond, VA, USA
This study further evaluated the in vitro activity of anidulafungin (VER002, Versicor Inc.)
(LY303366) against 460 clinical yeast isolates. MICs of anidulafungin, fluconazole and itra-
conazole were determined by following the NCCLS M27-A guidelines. Minimum fungicidal
concentrations (MFCs) of anidulafungin were determined for 230 isolates of Candida spp. The
activity of anidulafungin in vitro was significantly superior (P < 0.05) to those of itraconazole and
fluconazole against Candida albicans, Candida tropicalis, Candida glabrata and Candida krusei,
but anidulafungin was less active for Candida famata and Candida parapsilosis. The differences
were not significant for the other species evaluated.
163
2002 The British Society for Antimicrobial Chemotherapy
M. P. Arvalo et al.
form of Pneumocystis carinii; it has no activity against C. Sabouraud dextrose agar. The colonies were suspended in
neoformans.7 Kill-curves of anidulafungin are similar to 5 mL of sterile saline (0.85% NaCl). The inoculum suspen-
those produced by amphotericin B for C. albicans, Candida sions were shaken for 15 s and the inoculum density was
glabrata and Candida krusei.8 The activity in vivo of this adjusted to the turbidity of a 0.5 McFarland Standard (equiva-
echinocandin has been good in animal models (per os) of lent to 15 106 cfu/mL) with sterile saline. The suspensions
systemic candidiasis and in experimental pneumonia infec- were diluted 1:1000 in RPMI 1640 to give a final inoculum
tions due to P. carinii, as well as in infections produced by suspension equivalent to 0.52.5 103 cfu/mL.
Aspergillus fumigatus (intravenous).5,9 Ongoing Phase II
clinical trials will focus on the safety and efficacy of the intra- Microdilution tests
venous treatment with anidulafungin for patients suffering
invasive candidiasis. The present study evaluated the fungi- The broth microdilution assay was carried out following the
static (MICs) antifungal activities in vitro of anidulafungin, standard conditions described by the NCCLS3 with sterile,
fluconazole and itraconazole for Candida spp. The fungicidal 96-well, round-bottomed plates (Corning, New York, NY,
activity (MFCs) of anidulafungin was evaluated for 230 of the USA) and standard RPMI 1640 buffered to pH 7 with MOPS
460 isolates included in the study. as the test medium. On the day of the test, each microdilution
well containing 100 L of the corresponding two-fold drug
dilution was inoculated with 100 L of the two-fold-diluted
Materials and methods
cell suspension. Growth and sterility controls were included
Anidulafungin, as the standard powder LY-303366, was Incubation and scoring of MIC wells
kindly provided by Lilly Pharmaceuticals (Indiana, IN, USA),
fluconazole by Pfizer (Sandwich, Kent, UK) and itraconazole The microdilution trays were incubated at 35C and MICs
by Janssen Research Foundation (Beerse, Belgium) as stand- were determined visually after 48 h of incubation, using an
ard powders. Stock solutions of anidulafungin and itra- inverted mirror. The growth in each MIC well was compared
conazole (1600 mg/L) were prepared in 100% dimethyl with the growth in the control (antifungal-free) well. MICs of
sulphoxide (DMSO) (Merck, Darmstad, Germany) and anidulafungin were the lowest drug concentration wells that
fluconazole stock solutions were prepared in sterile distilled were optically clear (without any visible growth/turbidity).
water. Additive two-fold drug dilutions were prepared in the MICs of fluconazole and itraconazole were the lowest drug
corresponding solvents (100% strength) followed by further concentration wells that showed a prominent reduction
dilutions in standard RPMI 1640 broth to twice the strength (approximately 50%) in growth/turbidity.3
required for the final concentrations of 0.00316 mg/L
(anidulafungin, itraconazole) and 0.12564 mg/L (flucon- MFCs of anidulafungin
azole).
MFCs of anidulafungin were also determined for 215 C. albi-
cans and five isolates each of C. krusei, C. glabrata and
Strains
C. tropicalis recovered from a single clinical trial in the USA
A total of 460 clinical yeast isolates were evaluated: from oropharyngeal infections. Briefly, 20 L aliquots were
C. albicans (n = 317), Candida dubliniensis (n = 38), Candida subcultured from each well that showed complete inhibition
tropicalis (n = 27), C. glabrata (n = 25), C. krusei (n = 18), (optically clear) and from the growth control onto Sabouraud
Candida lusitaniae (n = 15), Candida famata (n = 10) and dextrose agar plates. The plates were incubated at 35C until
Candida parapsilosis (n = 10). The NCCLS quality control growth was seen in the growth control subculture (usually
(QC) Candida parapsilosis ATCC 22019 and C. krusei 48 h). The MFC was the lowest drug concentration that
ATCC 6358 strains3 were included as controls each time a set resulted in either no growth or fewer than three colonies.
of isolates was tested. The 460 isolates were recovered from
blood, vaginal, urine and oropharyngeal clinical specimens; Data analysis
210 from patients in the USA and 250 from Spain. These iso-
lates were maintained at room temperature in sterile distilled The MIC50 and MIC90 values were calculated as the con-
water and subcultured before testing. centrations of antifungal agent that were able to inhibit 50%
and 90% of the isolates, respectively. Geometric mean MICs
Inoculum preparation were obtained to facilitate comparisons of the activities of the
three drugs. The results were processed statistically using
Yeast inoculum suspensions were obtained by taking five SPSS PC+ version 9.0 for IBM PC. The criterion for statistical
colonies (>1 mm diameter) from 24-h-old cultures grown on significance was P < 0.05.
164
In vitro activity of anidulafungin against yeast
Table 1. In vitro susceptibilities (mg/L) of 460 clinical yeast isolates to anidulafungin, fluconazole and itraconazole
Organism (n) Antifungal agent Geometric mean MIC MIC50 MIC90 Range
165
M. P. Arvalo et al.
the infection, by impaired host defence mechanisms or by son with itraconazole and amphotericin B against Aspergillus spp.
other hostfungus and antifungal agent interactions. Antimicrobial Agents and Chemotherapy 42, 272630.
6. Powderly, W. G., Kobayashi, G. S., Herzig, G. P. & Medoff, G.
(1988). Amphotericin B-resistant yeast infection in severely immuno-
Acknowledgements compromised patients. American Journal of Medicine 84, 82632.
7. Espinel-Ingroff, A. (1998). Comparison of in vitro activities of
This study was partially supported by Lilly Pharmaceuticals.
the new triazole SCH56592 and the echinocandin MK-0991
(L-743,872) and LY303366 against opportunistic filamentous and
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