Abstract
Microbiome research of host-associated communities has been advanced recently through improvements in sequencing technologies and bioinformatic methods. Traditional microbiological culture, when combined with molecular techniques, can provide a sensitive platform to comprehensively study the airway microbiota. Here we describe the culture methods necessary to capture a large proportion of the airway microbiota and molecular methods for profiling bacterial communities through the 16S rRNA gene, which, when combined, offer a more complete picture of the diversity of airway microbial communities than either method alone.
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References
Man WH, de Steenhuijsen Piters WAA, Bogaert D (2017) The microbiota of the respiratory tract: gatekeeper to respiratory health. Nat Rev Microbiol 15(5):259–270
Stearns JC, Davidson CJ, McKeon S, Whelan FJ, Fontes ME, Schryvers AB et al (2015) Culture and molecular-based profiles show shifts in bacterial communities of the upper respiratory tract that occur with age. ISMEJ 9:1246–1259
Charlson ES, Bittinger K, Haas AR, Fitzgerald AS, Frank I, Yadav A et al (2011 Oct) Topographical continuity of bacterial populations in the healthy human respiratory tract. Am J Respir Crit Care Med 184(8):957–963
Bassis CM, Tang AL, Young VB, Pynnonen MA (2014) The nasal cavity microbiota of healthy adults. Microbiome 2:27
Bogaert D, Keijser B, Huse S, Rossen J, Veenhoven R, van Gils E et al (2011) Variability and diversity of nasopharyngeal microbiota in children: a metagenomic analysis. Semple M, editor. PLoS One 6(2):e17035
Whelan FJ, Verschoor CP, Stearns JC, Rossi L, Johnstone J, Surette MG et al (2014) The loss of topography in the microbial communities of the upper respiratory tract in the elderly. Ann Am Thorac Soc 11(4):513–521
Chotirmall SH, Burke CM (2015) Aging and the microbiome: implications for asthma in the elderly? Expert Rev Respir Med 9(2):125–128
Dickson RP, Erb-Downward JR, Freeman CM, McCloskey L, Beck JM, Huffnagle GB et al (2015) Spatial variation in the healthy human lung microbiome and the adapted island model of lung biogeography. Ann Am Thorac Soc 12(6):821–830
Segal LN, Alekseyenko AV, Clemente JC, Kulkarni R, Wu B, Gao Z et al (2013) Enrichment of lung microbiome with supraglottic taxa is associated with increased pulmonary inflammation. Microbiome 1(1):19
Dickson RP, Erb-Downward JR, Freeman CM, McCloskey L, Falkowski NR, Huffnagle GB et al (2017) Bacterial topography of the healthy human lower respiratory tract. MBio 8(1):e02287-16. https://doi.org/10.1128/mBio.02287-16
Lau JT, Whelan FJ, Herath I, Lee CH, Collins SM, Bercik P et al (2016) Capturing the diversity of the human gut microbiota through culture-enriched molecular profiling. Genome Med 8(1):72
Sibley CD, Grinwis ME, Field TR, Eshaghurshan CS, Faria MM, Dowd SE et al (2011) Culture enriched molecular profiling of the cystic fibrosis airway microbiome. PLoS One 6(7):11
Browne HP, Forster SC, Anonye BO, Kumar N, Neville BA, Stares MD et al (2016) Culturing of “unculturable”human microbiota reveals novel taxa and extensive sporulation. Nature 533(7604):543–546
Gevers D, Knight R, Petrosino JF, Huang K, McGuire AL, Birren BW et al (2012) The human microbiome project: a community resource for the healthy human microbiome. PLoS Biol 10(8):e1001377
Human Microbiome Project Consortium (2012) Structure, function and diversity of the healthy human microbiome. Nature 486(7402):207–214
Stearns JC, Lynch MDL, Senadheera DB, Tenenbaum HC, Goldberg MB, Cvitkovitch DG et al (2011) Bacterial biogeography of the human digestive tract. Sci Rep 1:170
Bartram AK, Lynch MDJ, Stearns JC, Moreno-Hagelsieb G, Neufeld JD (2011) Generation of multimillion-sequence 16S rRNA gene libraries from complex microbial communities by assembling paired-end illumina reads. Appl Environ Microbiol 77(11):3846–3852
Bentley DR, Balasubramanian S, Swerdlow HP, Smith GP, Milton J, Brown CG et al (2008) Accurate whole human genome sequencing using reversible terminator chemistry. Nature 456(7218):53–59
Caporaso GJ, Kuczynski J, Stombaugh J, Bittinger K, Bushman FD, Costello EK et al (2010) QIIME allows analysis of high-throughput community sequencing data. Nat Methods 7:335–336
Schloss PD, Westcott SL, Ryabin T, Hall JR, Hartmann M, Hollister EB et al (2009) Introducing mothur: open-source, platform-independent, community-supported software for describing and comparing microbial communities. Appl Environ Microbiol 75(23):7537–7541
McMurdie PJ, Holmes S (2013) Phyloseq: an R package for reproducible interactive analysis and graphics of microbiome census data. PLoS One 8(4):e61217
Kuczynski J, Lauber CL, Walters WA, Parfrey LW, Clemente JC, Gevers D et al (2011) Experimental and analytical tools for studying the human microbiome. Nat Rev Genet 13(1):47–58
Acknowledgments
Thank you to the members of the Surette laboratory as well as the clinicians and patients who have helped us improve the culturing conditions and techniques described within over the years.
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Whelan, F.J., Rossi, L., Stearns, J.C., Surette, M.G. (2018). Culture and Molecular Profiling of the Respiratory Tract Microbiota. In: Beiko, R., Hsiao, W., Parkinson, J. (eds) Microbiome Analysis. Methods in Molecular Biology, vol 1849. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8728-3_4
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DOI: https://doi.org/10.1007/978-1-4939-8728-3_4
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