Microbial signatures and host immune responses associated with the development of ventilator-associated pneumonia among patients with neurological injuries

ABSTRACT Ventilator-associated pneumonia (VAP) remains a leading complication in mechanically ventilated patients, yet the contribution of the respiratory microbiota remains poorly understood. The PULMIVAP study is a multicenter, longitudinal cohort investigation of respiratory microbiota composition and host immune responses in critically ill adults intubated for non-pulmonary conditions. A total of 146 intubated adult patients were enrolled across eight Italian ICUs, forming matched groups of 73 with VAP and 73 without. Oropharyngeal swabs and endotracheal aspirates were collected at intubation and either at VAP diagnosis or at a matched point in controls for a total of 584 biological samples. Metataxonomic analyses revealed substantial temporal shifts in microbial communities across both upper and lower respiratory compartments, with a trend toward reduced microbial richness in patients who developed VAP. Several genera, such as Corynebacterium, were more abundant in no-VAP patients, whereas Escherichia-Shigella and Peptoniphilus were enriched in VAP samples. Cytokine-microbiota correlation analysis suggested a pro-inflammatory signature in VAP patients, with Citrobacter positively associated with IFN-γ and TNF-α, while several commensal genera were inversely correlated with inflammatory mediators. Additionally, taxa associated with VAP correlated with lower PaO 2 /FiO 2 ratios, implicating them in disease severity. Consistently, several bacteria, such as Corynebacterium , appeared to be linked to better respiratory outcomes, suggesting protective or risk-associated microbial profiles. Overall, these findings highlight the complex interplay between microbial communities and mucosal immunity in the pathogenesis of VAP. The identification of condition-associated microbial and immunological signatures may inform future strategies for risk stratification and targeted prevention. IMPORTANCE Ventilator-associated pneumonia (VAP) remains a major complication of mechanical ventilation, yet most microbiome studies have focused on late-stage infection or single airway compartments, limiting insight into early microbial dynamics associated with VAP risk. By longitudinally characterizing upper and lower airway microbiota before and during VAP development, this study provides new insights into microbial and immune patterns associated with susceptibility and disease severity in humans. These findings contribute to the current understanding of VAP pathogenesis by suggesting a role for early airway dysbiosis and local immune responses alongside clinical factors. Remarkably, the identification of taxa associated with risk or protection supports the potential for microbiota-informed monitoring and future risk stratification strategies during mechanical ventilation. CLINICAL TRIALS This study is registered with ClinicalTrials.gov as NCT04849039 .