Reduction of turnaround time for non-tuberculous mycobacteria detection in heater–cooler units by propidium monoazide–real-time polymerase chain reaction

Abstract

Invasive non-tuberculous mycobacteria (NTM) infections are emerging worldwide in patients undergoing open-chest cardiac bypass surgery exposed to contaminated heater-cooler units (HCUs). Although this outbreak has been investigated by culturing bacteria isolated from HCU aerosol and water samples, these conventional methods have low-analytic sensitivity, high rates of sample contamination, and long turnaround time. To develop a simple and effective method to detect NTM in HCUs by real-time polymerase chain reaction (PCR), with a short laboratory turnaround time and reliable culture results. A total of 281 water samples collected from various HCUs at seven Italian hospitals were simultaneously screened for NTM by a propidium monoazide (PMA)-PCR assay and by conventional culture testing. The results were analysed with culture testing as the reference method. (i) The agreement between culture testing and PMA-PCR was 85.0% with a cycle threshold (CT) cut-off value of <38 vs 80.0% with a CT of <43, with a moderate Cohen's κ-coefficient; (ii) the CT cut-off value of <42 was deemed more suitable for predicting positive specimens; (iii) given the low concentration of target DNA in water samples, the minimum volume to be tested was 1 L. The use of PMA-PCR for fast detection of NTM from environmental samples is highly recommended in order to ascertain whether HCUs may represent a potential source of human exposure to NTM. This reliable and simple method reduces laboratory turnaround time compared to conventional methods (one to two days vs eight weeks, respectively), thereby improving control strategies and effective management of HCUs.