Previous Article | Next Article ![]()
Antimicrobial Agents and Chemotherapy, December 2001, p. 3474-3481, Vol. 45, No. 12
Departments of Human and Animal Infectious
Disease Research,1 and Biometrics
Research,2 Merck Research Laboratories, Rahway,
New Jersey 07065-0900
Received 15 June 2001/Returned for modification 14 August
2001/Accepted 20 September 2001
Caspofungin acetate (MK-0991) is an antifungal antibiotic that
inhibits the synthesis of 1,3-
0066-4804/01/$04.00+0 DOI: 10.1128/AAC.45.12.3474-3481.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Quantitative PCR Assay To Measure
Aspergillus fumigatus Burden in a Murine Model of
Disseminated Aspergillosis: Demonstration of Efficacy of
Caspofungin Acetate
-D-glucan, an essential
component of the cell wall of several pathogenic fungi. Caspofungin
acetate was recently approved for the treatment of invasive
aspergillosis in patients who are refractory to or intolerant of other
therapies. The activity of 1,3-
-D-glucan synthesis
inhibitors against Aspergillus fumigatus has been
evaluated in animal models of pulmonary or disseminated disease by
using prolongation of survival or reduction in tissue CFU as assay
endpoints. Because these methods suffer from limited sensitivity or
poor correlation with fungal growth, we have developed a quantitative
PCR-based (qPCR) (TaqMan) assay to monitor disease progression and
measure drug efficacy. A. fumigatus added to
naïve, uninfected kidneys as either ungerminated conidia or
small germlings yielded a linear qPCR response over at least 4 orders
of magnitude. In a murine model of disseminated aspergillosis, a burden
of A. fumigatus was detected in each of five different organs at 4 days postinfection by the qPCR assay, and the mean fungal
load in these organs was 1.2 to 3.5 log10 units greater than mean values determined by CFU measurement. When used to monitor disease progression in infected mice, the qPCR assay detected an
increase of nearly 4 log10 conidial equivalents/g of kidney between days 1 and 4 following infection, with a peak fungal burden that coincided with the onset of significant mortality. Traditional CFU
methodology detected only a marginal increase in fungal load in the
same tissues. In contrast, when mice were infected with Candida
albicans, which does not form true mycelia in tissues, quantitation of kidney burden by both qPCR and CFU assays was strongly
correlated as the infection progressed. Finally, treatment of mice with
induced disseminated aspergillosis with either caspofungin or
amphotericin B reduced the A. fumigatus burden in
infected kidneys to the limit of detection for the qPCR assay. Because of its much larger dynamic range, the qPCR assay is superior to traditional CFU determination for monitoring the progression of disseminated aspergillosis and evaluating the activity of antifungal antibiotics against A. fumigatus.
*
Corresponding author. Mailing address: Department of
Human and Animal Infectious Disease Research, Merck Research
Laboratories, RY80Y-230, P.O. Box 2000, Rahway, NJ 07065-0900. Phone:
(732) 594-5646. Fax: (732) 594-1399. E-mail:
cameron_douglas{at}merck.com.
This article has been cited by other articles:
| Clin. Vaccine Immunol. | Clin. Microbiol. Rev. |
|---|---|
| J. Clin. Microbiol. | ALL ASM JOURNALS |