Novel Quantitative PCR for <i>Rhodococcus equi</i> and Macrolide Resistance Detection in Equine Respiratory Samples

Abstract

<i>R. equi</i> is an important veterinary pathogen that takes the lives of many foals every year. With the emergence and spread of MDR <i>R. equi</i> to current antimicrobial treatment, new tools that can provide a fast and accurate diagnosis of the disease and antimicrobial resistance profile are needed. Here, we have developed and analytically validated a multiplex qPCR for the simultaneous detection of <i>R. equi</i> and related macrolide resistance genes in equine respiratory samples. The three sets of oligos designed in this study to identify <i>R. equi</i> housekeeping gene <i>choE</i> and macrolide resistance genes <i>erm</i>(46) and <i>erm</i>(51) showed high analytic sensitivity with a limit of detection (LOD) individually and in combination below 12 complete genome copies per PCR reaction, and an amplification efficiency between 90% and 147%. Additionally, our multiplex qPCR shows high specificity in in-silico analysis. Furthermore, it did not present any cross-reaction with normal flora from the equine respiratory tract, nor commonly encountered respiratory pathogens in horses or other genetically close organisms. Our new quantitative PCR is a trustable tool that will improve the speed of <i>R. equi</i> infection diagnosis, as well as helping in treatment selection.