The prevalence and microbiology of concomitant respiratory bacterial infections in patients with SARS-CoV-2 infection are not yet fully understood. In this retrospective study, we assessed respiratory bacterial co-infections in lower respiratory tract samples taken from intensive care unit-hospitalized COVID-19 patients, by comparing the conventional culture approach to an innovative molecular diagnostic technology. A total of 230 lower respiratory tract samples (i.e., bronchial aspirates or bronchoalveolar lavages) were taken from 178 critically ill COVID-19 patients. Each sample was processed by a semi-quantitative culture and by a multiplex PCR panel (FilmArray Pneumonia Plus panel), allowing rapid detection of a wide range of clinically relevant pathogens and a limited number of antimicrobial resistance markers. More than 30% of samples showed a positive bacterial culture, with Pseudomonas aeruginosa , Klebsiella pneumoniae and Staphylococcus aureus the most detected pathogens. FilmArray showed an overall sensitivity and specificity of 89.6% and 98.3%, respectively, with a negative predictive value of 99.7%. The molecular test significantly reduced the turn-around-time (TAT) and increased the rates of microbial detection. Most cases missed by culture were characterized by low bacterial loads (10 4 –10 5 copies/mL). FilmArray missed a list of pathogens not included in the molecular panel, especially Stenotrophomonas maltophilia (8 cases). FilmArray can be useful to detect bacterial pathogens in lower respiratory tract specimens of COVID-19 patients, with a significant decrease of TAT. The test is particularly useful to rule out bacterial co-infections and avoid the inappropriate prescription of antibiotics.

Introduction. Rapid identification of the causative agent of sepsis is crucial for patient outcomes. Aim. The Sepsityper sample preparation method enables direct microbial identification of positive blood culture samples via matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF MS). Hypothesis/Gap statement. The implementation of the Sepsityper method in the routine practice could represent a fundamental tool to achieve a prompt identification of the causative agent of bloodstream infections, and therefore accelerate the adoption of the proper antibiotic treatment. Methodology. In this study, the novel rapid workflow of the MALDI Biotypr Sepsityper kit (Bruker Daltonik GmbH, Germany) was evaluated using routine samples from a 2-year period (n=6918), and dedicated optimized protocols for the microbial groups that were more difficult to identify were developed. Moreover, the use of the residual bacterial pellet to perform susceptibility testing using different methods (commercial broth microdilution, disc diffusion, gradient diffusion) was investigated. Results. The rapid Sepsityper protocol allowed the identification of 5470/6338 (86.3 %) monomicrobial samples at species level, with very good performance for all of the clinically most significant pathogens (2510/2592 enterobacteria, 631/669 Staphylococcus aureus and 223/246 enterococci were identified). Streptococcus pneumoniae , Bacteroides fragilis and yeasts were the most troublesome to identify, but the application of specific optimized protocols significantly improved their rate of identification (from 14.7–71.5 %, 47.8–89.7 % and 37.1–89.5 %, respectively). Specificity was 100 % (no identification was made for the false-positive samples). Further, the residual pellet proved to be suitable to investigate susceptibility to antimicrobials, enabling us to simplify the workflow and shorten the time to report. Conclusion. The Rapid Sepsityper workflow proved to be a reliable sample preparation method for identification and susceptibility testing directly from positive blood cultures, providing novel approaches for accelerated diagnostics of bloodstream infections.