Early and specific targeted mass spectrometry-based identification of bacteria in endotracheal aspirates of patients suspected with ventilator-associated pneumonia

Rapid and reliable pathogen identification is compulsory to confirm ventilator-associated pneumonia (VAP) in order to initiate appropriate antibiotic treatment. In the present proof of concept, the effectiveness of rapid microorganism identification with a targeted bottom-up proteomics approach was investigated in endotracheal aspirate (ETA) samples of VAP patients. To do so, a prototype selected-reaction monitoring (SRM)-based assay was developed on a triple quadrupole mass spectrometer tracking proteotypic peptide surrogates of bacterial proteomes. Through the concurrent monitoring of 97 species-specific peptides, this preliminary assay was dimensioned to characterize the occurrence of six most frequent bacterial species responsible for over more than 65% of VAP. Assay performance was subsequently evaluated by analyzing early and regular 37 ETA samples collected from 15 patients. Twenty-five samples were above the significant threshold of 10 5 CFU/mL and five samples showed mixed infections (both pathogens ≥ 10 5 CFU/mL). The targeted proteomics assay showed 100% specificity for Acinetobacter baumannii , Escherichia coli , Haemophilus influenzae , Pseudomonas aeruginosa , Staphylococcus aureus , and Streptococcus pneumoniae . No false bacterial identification was reported and no interference was detected arising from the commensal flora. The overall species identification sensitivity was 19/25 (76%) and was higher at the patient level (84.6%). This successful proof of concept provides a rational to broaden the panel of bacteria for further clinical evaluation.