Rapid quantification of bacteria associated to ventilator associated pneumonia in endotracheal aspirate specimens

Ventilator-Associated Pneumonia (VAP) is one of the most serious infection in Intensive Care Unit. Today, its diagnosis relies mainly on conventional microbiological reports that are provided to the clinicians 24-72h after the sampling. Because an earlier diagnosis may reduce patient mortality and abusive use of antibiotics, we optimized a molecular biology-based method to quantify the DNA of bacteria frequently involved in VAP, directly from EndoTracheal Aspirates (ETA), in less than 3h30. First, a Sample Processing Control is spiked into each 400 μL specimen. Second, the sample is submitted to a liquefaction process and an enzymatic lysis, and DNA is extracted using the easyMAG® system. Then, qPCR are run and DNA is quantified using standard curves. Dynamic ranges for quantification were between ∼10^3 and ∼10^7 CFU/mL, depending on the bacterial target. The method was tested using ETA collected daily from mechanically ventilated patients (see patients 1 and 2, Figure 1) and we could follow the colonization with S. aureus over time. Indeed, the same trends were observed between the semi-quantitative culture and the ETA method. Further assessment is now needed to evaluate the medical validity of this method for the early diagnosis of VAP, on larger populations.