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Antimicrobial Agents and Chemotherapy, February 2000, p. 400-404, Vol. 44, No. 2
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Influence of Shaking on Antifungal Susceptibility Testing of Cryptococcus neoformans: a Comparison of the NCCLS Standard M27A Medium, Buffered Yeast Nitrogen Base, and RPMI-2% Glucose

Juan L. Rodríguez-Tudela,1,* Francisco Martín-Díez,1 Manuel Cuenca-Estrella,1 Laura Rodero,2 Yolanda Carpintero,1 and Begoña Gorgojo1

Servicio de Micología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra. Majadahonda-Pozuelo km.2, 28220 Majadahonda, Spain,1 and Departamento Micología, Instituto Nacional de Enfermedades Infecciosas, ANLIS "Dr. Carlos G. Malbrán," Avda. Velez Sarfield 563, 1281 Buenos Aires, Argentina2

Received 17 August 1999/Returned for modification 15 October 1999/Accepted 23 November 1999

Cryptococcus neoformans is a nonfermentative yeast that requires oxygen for growth. The shaking of culture media achieves good oxygenation, promoting the growth of cryptococci. In this study, three test media (RPMI 1640, RPMI 1640-2% glucose, and buffered yeast nitrogen base [BYNB]) recommended in the National Committee for Clinical Laboratory Standards M27A standard were examined. Growth abilities and minimum inhibitory concentrations (MICs) in microplates incubated at 35°C for 48 h were determined. The results indicated that shaking and an inoculum size of 105 CFU/ml yielded optimal growth of this yeast. Compared to RPMI 1640, supplementation of RPMI 1640 with 2% glucose did not significantly improve growth of C. neoformans and resulted in an 8.7-h delay of exponential growth. Cryptococcal growth in RPMI 1640 at 24 h was notably better than that in RPMI-2% glucose, although by 48 h the growths were comparable. The MIC range of amphotericin B observed for the C. neoformans strains grown in RPMI 1640 with or without glucose was too narrow to allow the separation of susceptible and resistant strains based on clinical outcome. The widest ranges of MICs of flucytosine and fluconazole were obtained with BYNB. This work demonstrates the need for a new antifungal susceptibility test for C. neoformans.

* Corresponding author. Mailing address: Servicio de Micología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra. Majadahonda-Pozuelo km.2, 28220 Majadahonda, Spain. Phone: 34 91 5097961. Fax: 34 91 5097966. E-mail: juanl.rodriguez-tudela{at}isciii.es.

Antimicrobial Agents and Chemotherapy, February 2000, p. 400-404, Vol. 44, No. 2
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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