Background

Airway infections are difficult to distinguish from acute rejection in lung transplant recipients. Traditional culture techniques take time that may delay treatment. We hypothesized that a rapid multiplex molecular assay could improve time to diagnosis and appropriate clinical decision making.

Methods

In a prospective observational study of recipients undergoing bronchoscopy, we assessed the BioFire^® FilmArray^® Pneumonia Panel (BFPP) in parallel to standard of care (SOC) diagnostics. Research clinicians performed shadow (research only) clinical decision making in real time. Time to report and interpretation were reported as median and interquartile ranges and compared by Wilcoxon signed-ranked test. Agreement was defined based on detection of any species targeted in the molecular assay.

Results

For the 150 enrolled subjects, BFPP results were available 3.8 hours (IQR 2.8-5.1) following bronchoscopy, compared to 13 hours for viral SOC (IQR 10-34, P < .001) results and 48 hours for bacterial SOC (IQR 46-70, P < .001) results. Positive BFPP results were interpreted in 9 hours (IQR 5-20) following bronchoscopy, compared to 74 hours for SOC (IQR 37-110, P < .001). Assays agreed for 138 (92%) of the 150 subjects. Of 22 BFPP diagnoses, five (23%) resulted in a shadow antibiotic recommendation. Notable BFPP deficiencies included fungal species and H parainfluenzae, accounting for 15 (27%) and 13 (23%) of the 56 actionable SOC results, respectively.

Conclusions

This molecular diagnostic including bacterial targets has the potential to shorten time to diagnosis and augment current clinical decision making but cannot replace SOC culture methods.

Pulmonary ischemia-reperfusion injury (IRI) is a clinical syndrome of acute lung injury that occurs after lung transplantation or remote organ ischemia. IRI causes early mortality and has no effective therapies. While NK cells are innate lymphocytes capable of recognizing injured cells, their roles in acute lung injury are incompletely understood. Here, we demonstrated that NK cells were increased in frequency and cytotoxicity in 2 different IRI mouse models. We showed that NK cells trafficked to the lung tissue from peripheral reservoirs and were more mature within lung tissue. Acute lung ischemia-reperfusion injury was blunted in a NK cell-deficient mouse strain but restored with adoptive transfer of NK cells. Mechanistically, NK cell NKG2D receptor ligands were induced on lung endothelial and epithelial cells following IRI, and antibody-mediated NK cell depletion or NKG2D stress receptor blockade abrogated acute lung injury. In human lung tissue, NK cells were increased at sites of ischemia-reperfusion injury and activated NK cells were increased in prospectively collected human bronchoalveolar lavage in subjects with severe IRI. These data support a causal role for recipient peripheral NK cells in pulmonary IRI via NK cell NKG2D receptor ligation. Therapies targeting NK cells may hold promise in acute lung injury.

Clinical outcomes after lung transplantation, a life-saving therapy for patients with end-stage lung diseases, are limited by primary graft dysfunction (PGD). PGD is an early form of acute lung injury with no specific pharmacologic therapies. Here, we present a large multicenter study of plasma and bronchoalveolar lavage (BAL) samples collected on the first posttransplant day, a critical time for investigations of immune pathways related to PGD. We demonstrated that ligands for NKG2D receptors were increased in the BAL from participants who developed severe PGD and were associated with increased time to extubation, prolonged intensive care unit length of stay, and poor peak lung function. Neutrophil extracellular traps (NETs) were increased in PGD and correlated with BAL TNF-α and IFN-γ cytokines. Mechanistically, we found that airway epithelial cell NKG2D ligands were increased following hypoxic challenge. NK cell killing of hypoxic airway epithelial cells was abrogated with NKG2D receptor blockade, and TNF-α and IFN-γ provoked neutrophils to release NETs in culture. These data support an aberrant NK cell/neutrophil axis in human PGD pathogenesis. Early measurement of stress ligands and blockade of the NKG2D receptor hold promise for risk stratification and management of PGD.