Assimilatory nitrate utilization by bacteria on the West Florida Shelf as determined by stable isotope probing and functional microarray analysis

Dissolved inorganic nitrogen (DIN) uptake by marine heterotrophic bacteria has important implications for the global nitrogen (N) and carbon (C) cycles. Bacterial nitrate utilization is more prevalent in the marine environment than traditionally thought, but the taxonomic identity of bacteria that utilize nitrate is difficult to determine using traditional methodologies. 15N-based DNA stable isotope probing was applied to document direct use of nitrate by heterotrophic bacteria on the West Florida Shelf. Seawater was incubated in the presence of 2 mu M 15N ammonium or 15N nitrate. DNA was extracted, fractionated via CsCl ultracentrifugation, and each fraction was analyzed by terminal restriction fragment length polymorphism (TRFLP) analysis. TRFs that exhibited density shifts when compared to controls that had not received 15N amendments were identified by comparison with 16S rRNA gene sequence libraries. Relevant marine proteobacterial lineages, notably Thalassobacter and Alteromonadales, displayed evidence of 15N incorporation. RT-PCR and functional gene microarray analysis could not demonstrate the expression of the assimilatory nitrate reductase gene, nasA, but mRNA for dissimilatory pathways, i.e. nirS, nirK, narG, nosZ, napA, and nrfA was detected. These data directly implicate several bacterial populations in nitrate uptake, but suggest a more complex pattern for N flow than traditionally implied

Ocean Acidification Regulates the Activity, Community Structure, and Functional Potential of Heterotrophic Bacterioplankton in an Oligotrophic Gyre

Ocean acidification (OA), a consequence of increased global carbon dioxide (CO2) emissions, is considered a major threat to marine ecosystems. Its effects on bacterioplankton activity, diversity, and community composition have received considerable attention. However, the direct impact of OA on heterotrophic bacterioplankton is often masked by the significant response of phytoplankton due to the close coupling of heterotrophic bacterioplankton and autotrophs. Here we investigated the responses of a heterotrophic bacterioplankton assemblage to high pCO2 (790-ppm) treatment in warm tropical western Pacific waters by conducting a microcosm experiment in dark for 12 days. Heterotrophic bacterioplankton abundance and production were enhanced by OA over the first 6 days of incubation, while the diversity and species richness were negatively affected. Bacterioplankton community composition in the high pCO2 treatment changed faster than that in the control. The molecular ecological network analysis showed that the elevated CO2 changed the overall connections among the bacterial community and resulted in a simple network under high CO2 condition. Species-specific responses to OA were observed and could be attributed to the different life strategies and to the ability of a given species to adapt to environmental conditions. In addition, high-throughput functional gene array analysis revealed that genes related to carbon and nitrogen cycling were positively affected by acidification. Together, our findings suggest that OA has direct effects on heterotrophic bacterioplankton in a low-latitude warm ocean and may therefore affect global biogeochemical cycles

Soil microbial beta-diversity is linked with compositional variation in aboveground plant biomass in a semi-arid grassland

Background and aims: Exploring biodiversity linkages between aboveground and belowground biota is a core topic in ecology, and can have implications on our understanding of ecosystem process stability. Yet, this topic still remains underexplored. Here, we explored diversity linkages, in terms of both alpha- and beta- diversity, between plant and top soil microbial communities in a semi-arid grassland ecosystem. Methods: Soil microbial community structure was assessed based on both 16S rRNA and functional genes, and plant community composition was evaluated by traditional “species composition” and a newly-defined “biomass composition”, which includes the information on the biomass of each species. Results: The bacterial alpha-diversity, expressed as the richness and Shannon diversity of 16S rRNA genes, was significantly correlated with plant species richness and Shannon diversity, whereas the alpha-diversity of microbial functional genes showed marginal association with total plant biomass. Microbial beta-diversity, evaluated by 16S rRNA genes, showed close relationship with plant beta-diversity estimated by both “species composition” and “biomass composition”, while the microbial beta-diversity based on functional genes was only associated with the compositional variation in aboveground plant biomass. Conclusions: These results showed that the differences in metabolic potential of soil microbial communities, which is closely related with ecosystem functions, can be better predicted by the variation of plant-derived resources returned to soil, than merely by the species composition of the macro-organism communities

Tundra soil carbon is vulnerable to rapid microbial decomposition under climate warming

Microbial decomposition of soil carbon in high-latitude tundra underlain with permafrost is one of the most important, but poorly understood, potential positive feedbacks of greenhouse gas emissions from terrestrial ecosystems into the atmosphere in a warmer world. Using integrated metagenomic technologies, we showed that the microbial functional community structure in the active layer of tundra soil was significantly altered after only 1.5 years of warming, a rapid response demonstrating the high sensitivity of this ecosystem to warming. The abundances of microbial functional genes involved in both aerobic and anaerobic carbon decomposition were also markedly increased by this short-term warming. Consistent with this, ecosystem respiration (R eco) increased up to 38%. In addition, warming enhanced genes involved in nutrient cycling, which very likely contributed to an observed increase (30%) in gross primary productivity (GPP). However, the GPP increase did not offset the extra R eco, resulting in significantly more net carbon loss in warmed plots compared with control plots. Altogether, our results demonstrate the vulnerability of active-layer soil carbon in this permafrost-based tundra ecosystem to climate warming and the importance of microbial communities in mediating such vulnerability

Climate warming leads to divergent succession of grassland microbial communities

Accurate climate projections require an understanding of the effects of warming on ecological communities and the underlying mechanisms that drive them . However, little is known about the effects of climate warming on the succession of microbial communities . Here we examined the temporal succession of soil microbes in a long-term climate change experiment at a tall-grass prairie ecosystem. Experimental warming was found to significantly alter the community structure of bacteria and fungi. By determining the time-decay relationships and the paired differences of microbial communities under warming and ambient conditions, experimental warming was shown to lead to increasingly divergent succession of the soil microbial communities, with possibly higher impacts on fungi than bacteria. Variation partition- and null model-based analyses indicate that stochastic processes played larger roles than deterministic ones in explaining microbial community taxonomic and phylogenetic compositions. However, in warmed soils, the relative importance of stochastic processes decreased over time, indicating a potential deterministic environmental filtering elicited by warming. Although successional trajectories of microbial communities are difficult to predict under future climate change scenarios, their composition and structure are projected to be less variable due to warming-driven selection. 1–3 4,

Assessing the Microbial Community and Functional Genes in a Vertical Soil Profile with Long-Term Arsenic Contamination

Conceived and designed the experiments: GW. Performed the experiments: JX GL. Analyzed the data: JX JZ GW. Contributed reagents/materials/analysis tools: ST JZ GW. Wrote the paper: JX ZH JDVN JZ GW.Arsenic (As) contamination in soil and groundwater has become a serious problem to public health. To examine how microbial communities and functional genes respond to long-term arsenic contamination in vertical soil profile, soil samples were collected from the surface to the depth of 4 m (with an interval of 1 m) after 16-year arsenic downward infiltration. Integrating BioLog and functional gene microarray (GeoChip 3.0) technologies, we showed that microbial metabolic potential and diversity substantially decreased, and community structure was markedly distinct along the depth. Variations in microbial community functional genes, including genes responsible for As resistance, carbon and nitrogen cycling, phosphorus utilization and cytochrome c oxidases were detected. In particular, changes in community structures and activities were correlated with the biogeochemical features along the vertical soil profile when using the rbcL and nifH genes as biomarkers, evident for a gradual transition from aerobic to anaerobic lifestyles. The C/N showed marginally significant correlations with arsenic resistance (p = 0.069) and carbon cycling genes (p = 0.073), and significant correlation with nitrogen fixation genes (p = 0.024). The combination of C/N, NO3− and P showed the highest correlation (r = 0.779, p = 0.062) with the microbial community structure. Contradict to our hypotheses, a long-term arsenic downward infiltration was not the primary factor, while the spatial isolation and nutrient availability were the key forces in shaping the community structure. This study provides new insights about the heterogeneity of microbial community metabolic potential and future biodiversity preservation for arsenic bioremediation management.Yeshttp://www.plosone.org/static/editorial#pee

Proteomic and Physiological Responses of Kineococcus radiotolerans to Copper

Copper is a highly reactive, toxic metal; consequently, transport of this metal within the cell is tightly regulated. Intriguingly, the actinobacterium Kineococcus radiotolerans has been shown to not only accumulate soluble copper to high levels within the cytoplasm, but the phenotype also correlated with enhanced cell growth during chronic exposure to ionizing radiation. This study offers a first glimpse into the physiological and proteomic responses of K. radiotolerans to copper at increasing concentration and distinct growth phases. Aerobic growth rates and biomass yields were similar over a range of Cu(II) concentrations (0–1.5 mM) in complex medium. Copper uptake coincided with active cell growth and intracellular accumulation was positively correlated with Cu(II) concentration in the growth medium (R2 = 0.7). Approximately 40% of protein coding ORFs on the K. radiotolerans genome were differentially expressed in response to the copper treatments imposed. Copper accumulation coincided with increased abundance of proteins involved in oxidative stress and defense, DNA stabilization and repair, and protein turnover. Interestingly, the specific activity of superoxide dismutase was repressed by low to moderate concentrations of copper during exponential growth, and activity was unresponsive to perturbation with paraquot. The biochemical response pathways invoked by sub-lethal copper concentrations are exceptionally complex; though integral cellular functions are preserved, in part, through the coordination of defense enzymes, chaperones, antioxidants and protective osmolytes that likely help maintain cellular redox. This study extends our understanding of the ecology and physiology of this unique actinobacterium that could potentially inspire new biotechnologies in metal recovery and sequestration, and environmental restoration

Using hiCLIP to identify RNA duplexes that interact with a specific RNA-binding protein

The structure of RNA molecules has a critical role in regulating gene expression, largely through influencing their interactions with RNA-binding proteins (RBPs). RNA hybrid and individual-nucleotide resolution UV cross-linking and immunoprecipitation (hiCLIP) is a transcriptome-wide method of monitoring these interactions by identifying RNA duplexes bound by a specific RBP. The hiCLIP protocol consists of the following steps: in vivo cross-linking of RBPs to their bound RNAs; partial RNA digestion and purification of RNA duplexes interacting with the specific RBP using immunoprecipitation; ligation of the two arms of RNA duplexes via a linker; reverse transcription; cDNA library amplification; and finally high-throughput DNA sequencing. Mapping of the sequenced arms to a reference transcriptome identifies the exact locations of duplexes. hiCLIP data can directly identify all types of RNA duplexes bound by RBPs, including those that are challenging to predict computationally, such as intermolecular and long-range intramolecular duplexes. Moreover, the use of an adaptor that links the two arms of the RNA duplex permits hiCLIP to unambiguously identify the duplexes. Here we describe in detail the procedure for a hiCLIP experiment and the subsequent streamlined data analysis with an R package, 'hiclipr' (https://github.com/luslab/hiclipr/). Preparation of the library for high-throughput DNA sequencing takes ∼7 d and the basic bioinformatic pipeline takes 1 d