Isolation and cultivation of lung microbiota with culturomics

Abstract:

[Objective] To understand the composition and characteristics of the human lung microbiota, and build a respiratory tract microbial bank which provides strains for further study, we isolated lung microbiota with culturomics.[Methods] Six clinical alveolar lavage fluid samples were collected, and blood culture bottles supplemented with different additives were used to pre-enrich the samples. The strains in the blood bottles were isolated, cultured, and preserved at different time points of the pre-enrichment. Isolated strains were identified with matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry or 16S rRNA gene sequencing.[Results] A total of 101 known bacterial species, 6 potential new bacterial species, and 2 fungal species were obtained. The bacteria belonged to 45 genera, 35 families, 24 orders, 14 classes of 5 phyla. More isolates were obtained from the early stage of pre-enrichment, and there were more isolates in the anaerobic environment than at aerobic conditions. The additional supplement of sheep blood and rumen fluid during pre-enrichment were beneficial to strain isolation. The lung bacteria isolated in this experiment overlapped to a large extent with those from other parts (especially the intestine) of the human body.[Conclusion] Culturomics realizes the isolation of lung microbiota, and it is essential to continue to optimize the methods for isolating more bacteria from lungs.