Quantitation of pulmonary fungal burden in Paracoccidioides brasiliensis-infected mice by real-time PCR

Abstract

ABSTRACT Although colony-forming unit (CFU) counting is widely used to quantify fungal load in tissue from animal experimentally infected with Paracoccidioides brasiliensis, several technical disadvantages have been described. Here we developed highly accurate quantitative PCR (qPCR) assays to determine the relative P brasiliensis load in lungs from infected mice. SYBR Green- and TaqMan-based assays using primers and probe for the 43-kDa glycoprotein (gp43) gene detected as little as 270 gene copies (about 2 fg of DNA) per reaction. Although qPCR assays cannot distinguish between living and dead yeasts, we found a highly positive linear correlation between CFU and qPCR.

Keywords

• Paracoccidioides brasiliensis
• Paracoccidioidomycosis
• Quantitative PCR
• gp43 gene
• Experimental infection
• Real-Time PCR