Revealing the dynamics of polymicrobial infections: update on the qPCR as a promising tool for the quantification of bacterial jungles

Abstract

P. aeruginosa and S. aureus are important opportunistic human pathogens in many polymicrobial infections. Interactions between them change the infection dynamics, resulting in increased potential for disease development. Determining the relative bacterial abundance through culture-dependent approaches is hindered by the presence of cells in a viable but poorly-cultivable state, or underestimated by the presence of cell-aggregates. The monitoring of bacterial pathogens by the use of culture-independent tools has led to fresh insights into the complex relationships between host and microbes, but often key experimental controls are lacking. This study aimed at examining changes in microbial composition in P. aeruginosa (PA) and S. aureus (SA) communities by quantitative PCR (qPCR). Total RNA was extracted and normalized against the amount of an exogenous RNA control-molecule. The Cq value for each gene of interest was transformed into relative quantities taking the differences between the target and the calibrator exogenous mRNA.