Inter-laboratory testing of the effect of DNA blocking reagent G2 on DNA extraction from low-biomass clay samples

Here we show that a commercial blocking reagent (G2) based on modifed eukaryotic DNA signifcantly improved DNA extraction efciency. We subjected G2 to an inter-laboratory testing, where DNA was extracted from the same clay subsoil using the same batch of kits. The inter-laboratory extraction campaign revealed large variation among the participating laboratories, but the reagent increased the number of PCR-amplifed16S rRNA genes recovered from biomass naturally present in the soils by one log unit. An extensive sequencing approach demonstrated that the blocking reagent was free of contaminating DNA, and may therefore also be used in metagenomics studies that require direct sequencing

Emissions of biogenic volatile organic compounds from arctic shrub litter are coupled with changes in the bacterial community composition

Emissions of biogenic volatile organic compounds (BVOCs) from natural ecosystems impact atmospheric chemistry as well as biological interactions and even soil biogeochemical processes. Plant litter emits substantial amounts of BVOCs. These emissions may contribute to total ecosystem emissions especially in the Arctic where the living plant biomass is low and the amount of litter is expected to increase as the deciduous shrubs expand in response to a warmer climate. Here, we incubated in the laboratory litter from the evergreen Cassiope tetragona and deciduous Salix spp. from a high arctic and a low arctic location. The 8-week-long incubation was conducted with temperature increasing from 5 °C to 26 °C, mimicking the transition from winter to summer. BVOC emissions from the decomposing litter were sampled weekly in adsorbent cartridges and analyzed using gas chromatography–mass spectrometry, and the bacterial community composition was investigated by sequencing of PCR amplified 16S rRNA gene fragments. Our results showed that litter from C. tetragona, which is a terpenoid storing species, had higher BVOC emission rates (mainly terpenoids) than the Salix litter, which does not have specialized BVOC storing compartments. The C. tetragona litter emissions were higher in the high arctic than the low arctic samples. The emission rates from the C. tetragona litter increased during the incubation period, whereas emission rates from the Salix litter decreased, suggesting that the emissions originated from different sources and/or processes. The bacterial community composition in the Salix litter, but not in the C. tetragona litter, changed in parallel with the changes in the BVOC emissions during the incubation period. Therefore, we suggest that bacteria may be more important for the BVOC emissions from decomposing Salix litter than C. tetragona litter

Culturing of "Unculturable" Subsurface Microbes: Natural Organic Carbon Source Fuels the Growth of Diverse and Distinct Bacteria From Groundwater.

Recovery and cultivation of diverse environmentally-relevant microorganisms from the terrestrial subsurface remain a challenge despite recent advances in modern molecular technology. Here, we applied complex carbon (C) sources, i.e., sediment dissolved organic matter (DOM) and bacterial cell lysate, to enrich groundwater microbial communities for 30 days. As comparisons, we also included enrichments amended with simple C sources including glucose, acetate, benzoate, oleic acid, cellulose, and mixed vitamins. Our results demonstrate that complex C is far more effective in enriching diverse and distinct microorganisms from groundwater than simple C. Simple C enrichments yield significantly lower biodiversity, and are dominated by few phyla (e.g., Proteobacteria and Bacteroidetes), while microcosms enriched with complex C demonstrate significantly higher biodiversity including phyla that are poorly represented in published culture collections (e.g., Verrucomicrobia, Planctomycetes, and Armatimonadetes). Subsequent isolation from complex C enrichments yielded 228 bacterial isolates representing five phyla, 17 orders, and 56 distinct species, including candidate novel, rarely cultivated, and undescribed organisms. Results from this study will substantially advance cultivation and isolation strategies for recovering diverse and novel subsurface microorganisms. Obtaining axenic representatives of "once-unculturable" microorganisms will enhance our understanding of microbial physiology and function in different biogeochemical niches of terrestrial subsurface ecosystems

Culturing of “Unculturable” Subsurface Microbes: Natural Organic Carbon Source Fuels the Growth of Diverse and Distinct Bacteria From Groundwater

© Copyright © 2020 Wu, Spencer, Gushgari-Doyle, Yee, Voriskova, Li, Alm and Chakraborty. Recovery and cultivation of diverse environmentally-relevant microorganisms from the terrestrial subsurface remain a challenge despite recent advances in modern molecular technology. Here, we applied complex carbon (C) sources, i.e., sediment dissolved organic matter (DOM) and bacterial cell lysate, to enrich groundwater microbial communities for 30 days. As comparisons, we also included enrichments amended with simple C sources including glucose, acetate, benzoate, oleic acid, cellulose, and mixed vitamins. Our results demonstrate that complex C is far more effective in enriching diverse and distinct microorganisms from groundwater than simple C. Simple C enrichments yield significantly lower biodiversity, and are dominated by few phyla (e.g., Proteobacteria and Bacteroidetes), while microcosms enriched with complex C demonstrate significantly higher biodiversity including phyla that are poorly represented in published culture collections (e.g., Verrucomicrobia, Planctomycetes, and Armatimonadetes). Subsequent isolation from complex C enrichments yielded 228 bacterial isolates representing five phyla, 17 orders, and 56 distinct species, including candidate novel, rarely cultivated, and undescribed organisms. Results from this study will substantially advance cultivation and isolation strategies for recovering diverse and novel subsurface microorganisms. Obtaining axenic representatives of “once-unculturable” microorganisms will enhance our understanding of microbial physiology and function in different biogeochemical niches of terrestrial subsurface ecosystems

Inter-laboratory testing of the effect of DNA blocking reagent G2 on DNA extraction from low-biomass clay samples

Here we show that a commercial blocking reagent (G2) based on modified eukaryotic DNA significantly improved DNA extraction efficiency. We subjected G2 to an inter-laboratory testing, where DNA was extracted from the same clay subsoil using the same batch of kits. The inter-laboratory extraction campaign revealed large variation among the participating laboratories, but the reagent increased the number of PCR-amplified16S rRNA genes recovered from biomass naturally present in the soils by one log unit. An extensive sequencing approach demonstrated that the blocking reagent was free of contaminating DNA, and may therefore also be used in metagenomics studies that require direct sequencing