Fungal and Bacterial Diversity of Airway Microbiota in Adults with Cystic Fibrosis: Concordance Between Conventional Methods and Ultra-Deep Sequencing, and Their Practical use in the Clinical Laboratory

Autor: Benoit Wallaert, Franziska A. Stressmann, Françoise Botterel, Kenneth D. Bruce, Cécile Angebault, J. M. Costa, Odile Cabaret, Laurence Delhaes, Frédéric Wallet
Rok vydání: 2017
Předmět:
Adult
DNA
Bacterial
Microbiological Techniques
0301 basic medicine
medicine.medical_specialty
Cystic Fibrosis
Veterinary (miscellaneous)
Respiratory System
030106 microbiology
Plant Science
DNA
Ribosomal
Applied Microbiology and Biotechnology
Microbiology
Cystic fibrosis
Young Adult
03 medical and health sciences
Medical microbiology
RNA
Ribosomal
16S
DNA
Ribosomal Spacer
RNA
Ribosomal
18S
medicine
Cluster Analysis
Humans
Internal transcribed spacer
DNA
Fungal
Phylogeny
Bacteria
biology
Microbiota
Fungi
Sputum
High-Throughput Nucleotide Sequencing
Sequence Analysis
DNA
Ribosomal RNA
medicine.disease
16S ribosomal RNA
biology.organism_classification
Agricultural and Biological Sciences (miscellaneous)
Terminal restriction fragment length polymorphism
030104 developmental biology
Female
medicine.symptom
Agronomy and Crop Science
Polymorphism
Restriction Fragment Length
Follow-Up Studies
Zdroj: Mycopathologia. 183:171-183
ISSN: 1573-0832
0301-486X
Popis: Given the complexity of the airway microbiota in the respiratory tract of cystic fibrosis (CF) patients, it seems crucial to compile the most exhaustive and exact list of the microbial communities inhabiting CF airways. The aim of the present study was to compare the bacterial and fungal diversity of sputa from adult CF patients during non-exacerbation period by culture-based and molecular methods, and ultra-deep-sequencing (UDS). Sputum samples from four CF patients were cultured and analysed by DNA extractions followed by terminal restriction fragment length polymorphism analysis through resolution of bacterial ribosomal gene (rDNA) fragments, and cloning plus sequencing of part of fungal rRNA genes. These approaches were compared with UDS method targeting 16S rDNA gene and the internal transcribed spacer (ITS) 2 region of rDNA. A total of 27 bacterial and 18 fungal genera were detected from the four patients. Five (18%) and 3 (16%) genera were detected by culture for bacteria and fungi, respectively, 9 (33%) and 3 (16%) by first generation sequencing (FGS) methods, and 26 (96%) and 18 (100%) by UDS. The mean number of genera detected by UDS per patient was statistically higher than by culture or FGS methods. Patients with severe airway disease as assessed by standard spirometry exhibited a reduced fungal and bacterial diversity. UDS approach evaluates more extensively the diversity of fungal and bacterial flora compared with cultures. However, it currently remains difficult to routinely use UDS mainly because of the lack of standardization, and the current cost of this method.
Databáze: OpenAIRE
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