Upper Tract Microbiome Modifications After Lung Transplantation and Its Impact in CLAD

Purpose Survival after lung transplantation is limited in large part due to the high incidence of chronic lung allograft dysfunction (CLAD). Infection is a recognized risk factor for the development of CLAD, and both acute infection and chronic lung allograft colonization with microorganisms increase the risk for CLAD. The aim of our study was to analyze nasopharyngeal (NP) microbiome composition and modifications following lung transplantation, with a focus on its relationship with CLAD. Methods This was a longitudinal study in which we analyzed bacterial microbiome in NP swab samples in 68 lung transplanted (LT) patients the day of the transplant, when patients were discharged, at 2-5 months and 12 months after lung transplantation. We also analysed NP swab samples from 10 healthy subjects. CLAD was assessed after 2 years of follow-up. DNA was extracted from the NP swabs. To create the amplicon library, the hyper-variable region (V4) of the 16S ribosomal RNA gene was amplified by standard PCR. Amplicons were purified and sequenced in barcoded pools using Illumina MiSeq technology. Raw sequence reads were demultiplexed by using idemp. The resulting single-end reads were processed using the Dada2 pipeline obtaining an amplicon sequence variant table to which taxonomy was assigned. Alpha diversity, including Shannon and Observed indices, and beta diversity metrics were calculated. We compared these metrics and particular taxa relative abundances between samples of LT patients at different time points and samples of healthy subjects. Results From the 68 studied patients, 13 developed CLAD during the two years of follow-up. We observed a significant decrease in the Observed index at the day of the discharge and 2-5 months after transplant when compared with healthy subjects. One year after transplant, CLAD patients had higher Shannon diversity than LT patients who did not develop the dysfunction. On the other hand, a lower beta diversity was observed between healthy subjects and patients with no CLAD. Besides, we detected an increased genus in NP swab samples associated with no CLAD also observed in NP samples from healthy subjects. Conclusion Nasopharyngeal overall microbiome composition, as well as relative abundances of particular taxa of non-CLAD patients, was closest to the one from healthy subjects suggesting a better outcome of the transplant if better microbiome normalization. Survival after lung transplantation is limited in large part due to the high incidence of chronic lung allograft dysfunction (CLAD). Infection is a recognized risk factor for the development of CLAD, and both acute infection and chronic lung allograft colonization with microorganisms increase the risk for CLAD. The aim of our study was to analyze nasopharyngeal (NP) microbiome composition and modifications following lung transplantation, with a focus on its relationship with CLAD. This was a longitudinal study in which we analyzed bacterial microbiome in NP swab samples in 68 lung transplanted (LT) patients the day of the transplant, when patients were discharged, at 2-5 months and 12 months after lung transplantation. We also analysed NP swab samples from 10 healthy subjects. CLAD was assessed after 2 years of follow-up. DNA was extracted from the NP swabs. To create the amplicon library, the hyper-variable region (V4) of the 16S ribosomal RNA gene was amplified by standard PCR. Amplicons were purified and sequenced in barcoded pools using Illumina MiSeq technology. Raw sequence reads were demultiplexed by using idemp. The resulting single-end reads were processed using the Dada2 pipeline obtaining an amplicon sequence variant table to which taxonomy was assigned. Alpha diversity, including Shannon and Observed indices, and beta diversity metrics were calculated. We compared these metrics and particular taxa relative abundances between samples of LT patients at different time points and samples of healthy subjects. From the 68 studied patients, 13 developed CLAD during the two years of follow-up. We observed a significant decrease in the Observed index at the day of the discharge and 2-5 months after transplant when compared with healthy subjects. One year after transplant, CLAD patients had higher Shannon diversity than LT patients who did not develop the dysfunction. On the other hand, a lower beta diversity was observed between healthy subjects and patients with no CLAD. Besides, we detected an increased genus in NP swab samples associated with no CLAD also observed in NP samples from healthy subjects. Nasopharyngeal overall microbiome composition, as well as relative abundances of particular taxa of non-CLAD patients, was closest to the one from healthy subjects suggesting a better outcome of the transplant if better microbiome normalization.