Recovering glycoside hydrolase genes from active tundra cellulolytic bacteria

• Published in: Canadian Journal of Microbiology
• Main Authors: Pinnell, Lee J., Dunford, Eric, Ronan, Patrick, Hausner, Martina, Neufeld, Josh D.
• Format: Article in Journal/Newspaper
• Language: English
• Published: Canadian Science Publishing 2014
• Subjects: Tundra
• Online Access: http://dx.doi.org/10.1139/cjm-2014-0193; http://www.nrcresearchpress.com/doi/full-xml/10.1139/cjm-2014-0193; http://www.nrcresearchpress.com/doi/pdf/10.1139/cjm-2014-0193

Bacteria responsible for cellulose hydrolysis in situ are poorly understood, largely because of the relatively recent development of cultivation-independent methods for their detection and characterization. This study combined DNA stable-isotope probing (DNA-SIP) and metagenomics for identifying active bacterial communities that assimilated carbon from glucose and cellulose in Arctic tundra microcosms. Following DNA-SIP, bacterial fingerprint analysis of gradient fractions confirmed isotopic enrichment. Sequenced fingerprint bands and clone library analysis of 16S rRNA genes identified active bacterial taxa associated with cellulose-associated labelled DNA, including Bacteroidetes (Sphingobacteriales), Betaproteobacteria (Burkholderiales), Alphaproteobacteria (Caulobacteraceae), and Chloroflexi (Anaerolineaceae). We also compared glycoside hydrolase metagenomic profiles from bulk soil and heavy DNA recovered from DNA-SIP incubations. Active populations consuming [ 13 C]glucose and [ 13 C]cellulose were distinct, based on ordinations of light and heavy DNA. Metagenomic analysis demonstrated a ∼3-fold increase in the relative abundance of glycoside hydrolases in DNA-SIP libraries over bulk-soil libraries. The data also indicate that multiple displacement amplification introduced bias into the resulting metagenomic analysis. This research identified DNA-SIP incubation conditions for glucose and cellulose that were suitable for Arctic tundra soil and confirmed that DNA-SIP enrichment can increase target gene frequencies in metagenomic libraries.