Image: The MiSeq next-generation sequencing system (Photo courtesy of the University of California Los Angeles).
The natural evolution of cystic fibrosis (CF) is a progressive decline in lung function caused by a vicious circle of inflammation and tissue destruction, which is triggered and maintained by the chronic bacterial colonization of the lower respiratory tract.
The introduction of culture-independent tools, particularly those based on next-generation sequencing (NGS), has allowed the identification of a more diverse and abundant lung microbiota that includes not only classical CF pathogens but also a wide community of bacterial taxa, most of which are uncultivable by routine methods.
Medical scientists at the Ramón y Cayal University Hospital (Madrid, Spain) and their colleagues analyzed 56 sputum samples from 15 cystic fibrosis patients at the hospital. Patients were classified by lung function impairment as mild (five patients), moderate (nine patients), or severe (one patient). Sputum samples were routinely aerobically cultured in accordance with the recommendations of the Spanish CF guidelines. All colonies, except for those of Burkholderia cepacia complex, were identified by matrix-assisted laser desorption ionization time of flight mass spectrometry.
The sputum samples were slowly defrosted at 4 °C for 24 hours to prevent DNA degradation and further thawed at room temperature. After complete vortex mixing of the sample, total DNA was obtained from an aliquot of ~0.5 mL of the supernatant with the QIAamp DNA minikit. DNA samples were sent for massive 16S rRNA gene V3-V4 amplicon sequencing on the MiSeq platform and for bioinformatic analysis.
The team found a wide range of 156 types of bacterial species in the samples, including Pseudomonas aeruginosa, Staphylococcus aureus, Burkholderia and Pandorea. Each patient had his or her own bacterial makeup that remained relatively steady over the study period. They also found two types of predator bacteria among the samples. Vampirovibrio, a bacterium that destroys cells by sucking out its contents, was found in 17 samples from 12 patients, while Bdellovibrio, which enters cells and feeds on its proteins, was found in six samples from three patients. The two types were found together in only one patient.
Rosa del Campo, MD, PhD, the senior author of the study, said, “Predator bacteria are ubiquitous and are usually found in environmental aquatic ecosystems. In humans, a recent study has found them in the intestinal microbiota of healthy individuals and in patients with cystic fibrosis. The findings indicate that the lung microbiota in cystic fibrosis patients is more complex than we believed. Our study suggests that predatory bacteria could be used as a therapeutic strategy to reduce the bacterial load of the lungs of these patients.” The study was published on September 26, 2017, in the journal mBIO.
Ramón y Cayal University Hospital