Berkeley Lab

New CRISPR–Cas systems from uncultivated microbes

a) Lineages with and without isolated representatives, and the scale of little investigated biology. b) Locus organization of the discovered systems

Scientific Achievement

Novel genomes reconstructed from metagenomes were shown to encode previously unknown CRISPR-Cas systems with potential for genome editing.

Significance and Impact

The new genome-editing systems are among the most compact yet identified. The systems are currently being developed as alternatives to the Cas9-based system.

Research Details
  • An extensive set of genomes reconstructed largely in DOE funded research was searched for novel gene clusters adjacent to CRISPR arrays.
  • Two candidate systems, CasX and CasY, were tested in interference assays
  • Novel archaeal genomes were shown to contain the first Cas9-based loci from this domain of life
Citation

Burstein, D.; Harrington, L. B.; Strutt, S. C.; Probst, A. J.; Anantharaman, K.; Thomas, B. C.; Doudna, J. A.; Banfield, J. F. (2017), New CRISPR–Cas systems from uncultivated microbes, Nature, 542(7640), 237-241 DOI: 10.1038/nature21059.

Diverse Microbial Metabolism in Aquifer BGC Hot Spot

Key-metabolic-pathways

Summary of key metabolic pathways expressed by a prominent bacterium (Hydrogenophaga b174) in an NRZ biogeochemical hot spot in the Rifle aquifer. Surprisingly, this bacterium actively catalyzed both heterotrophic and chemolithoautotrophic processes and influenced biogeochemical cycling of several elements, including C, N, and S. Unexpectedly, denitrification played an important role in this metabolism.

Organic matter deposits in alluvial aquifers have been shown to result in the formation of NRZs, which can modulate aquifer redox status and influence the speciation and mobility of metals, significantly affecting groundwater geochemistry. This study (Jewell et al., Frontiers in Microbiology) sought to better understand how natural organic matter fuels microbial communities within anoxic biogeochemical hot spots (NRZs) in a shallow alluvial aquifer at the Rifle (CO) site. Overall, the results highlighted the complex nature of organic matter transformation in NRZs and the microbial metabolic pathways that interact to mediate redox status and elemental cycling.

Summary

The authors used an anaerobic microcosm experiment in which NRZ sediments served as the sole source of electron donors and microorganisms. Biogeochemical data indicated that the decomposition of native organic matter occurred in different phases, beginning with mineralization of dissolved organic matter (DOM) to CO2 during the first week of incubation, followed by a pulse of acetogenesis that dominated carbon flux after two weeks. The depletion of DOM over time was strongly correlated with increases in expression of many genes associated with heterotrophy (e.g., amino acid, fatty acid, and carbohydrate metabolism) belonging to a Hydrogenophaga strain that accounted for a relatively large percentage (~8%) of the metatranscriptome. This Hydrogenophaga strain also expressed genes indicative of chemolithoautotrophy, including CO2 fixation, H2 oxidation, S-compound oxidation, and denitrification. The pulse of acetogenesis appears to have been collectively catalyzed by a number of different organisms and metabolisms, most prominently pyruvate:ferredoxin oxidoreductase. Unexpected genes were identified among the most highly expressed (>98th percentile) transcripts, including acetone carboxylase and cell-wall-associated hydrolases with unknown substrates. Many of the most highly expressed hydrolases belonged to a Ca. Bathyarchaeota strain and may have been associated with recycling of bacterial biomass.

Modelling the Evolution of Complex Conductivity During Calcite Precipitation on Glass Beads

A: Complex conductivity model of the porous medium with calcite precipitation.
Imaginary conductivity of calcite as a function of time in days before (A) and after (B) the pore clogging

Scientific Achievement

Complex conductivity signals of calcite provides an in-situ monitoring approach for its precipitation, yet the polarization mechanism is not clear. We developed a mechanistic model considering the electrochemical polarization of the Stern and diffuse layers surrounding calcite particles.

Significance and Impact

The model provides a mechanistic understanding of complex conductivity signals from in-situ calcite precipitation that often occurs during subsurface reactional processes. This is key to develop geophysical monitoring for dynamic monitoring of subsurface biogeochemistry.

Research Details
  • The experimental data sets are provided by Wu et al. (2010)
  • Numerical modeling is based on the stern layer electrochemical polarization model at the calcite – fluid interface
  • Comparison between the model simulation and experimental results provides validation of the model.
Citation

Leroy, P., Li, S., Revil, A., Wu, Y., 2017, Modelling the Evolution of Complex Conductivity During Calcite Precipitation on Glass Beads, Geophys. J. Int. 209,
123–140, DOI: 10.1093/gji/ggx001

Asgard archaea illuminate the origin of eukaryotic cellular complexity

Phylogenetic tree showing the placement of Eukaryotes within Archaea

Scientific Achievement

Regarding eukaryote origin, we describe a superphylum of uncultivated archaea whose genomes are enriched for proteins formerly considered eukaryote specific, indicating the archaeal host cell already contained many key components that govern eukaryotic cellular complexity.

Significance and Impact

The findings support the phylogenetic inference that eukaryotes arose from an archaeal lineage, which potentially shifts the tree of life back to two domains, Results elucidate archaeal innovations that may have laid the foundation for the evolution of complex, multi-cellular life.

Research Details
  • Archaeal genomes that group together phylogenetically were collected from a variety of different ecosystems
  • Together, these genomes were used to define the new archaeal superphylum referred to as Asgard
  • Genomes were investigated regarding their metabolic capacities
  • Normally eukaryotic pathways were identified.
Citation

Zaremba-Niedzwiedzka, K.; Caceres, E. F.; Saw, J. H.; Bäckström, D.; Juzokaite, L.;et al..; Banfield, J. F.; Schramm, A.; Baker, B. J.; Spang, A.; Ettema, T. J. G. (2017) Asgard archaea illuminate the origin of eukaryotic cellular complexity, Nature, 541(7637), 353-358 DOI: 10.1038/nature21031.

Unusual respiratory capacity and nitrogen metabolism in a Parcubacterium (OD1) of the Candidate Phyla Radiation

An exceedingly novel bacterium has very unusual capacities encoded by highly divergent genes

Scientific Achievement

Parcunitrobacter nitroensis, a Candidate Phylum Radiation (CPR) bacterium, has versatile energy pathways and the first complete electron transport chain described in a member of this major subdivision within Domain Bacteria

Significance and Impact

The CPR is a major group of environmentally-relevant bacteria elucidated our in prior DOE-funded research. Unlike virtually all CPR, we report an exceedingly novel organism that challenges understanding of CPR evolution.

Research Details
  • We sampled microbial communities from an aquifer adjacent to the Colorado River in Rifle, Colorado
  • Whole community DNA was sequenced
  • A near-complete genome was reconstructed
  • Metabolic predictions uncover unexpected capacities encoded by highly divergent genes
Citation

Castelle, C. J.; Brown, C. T.; Thomas, B. C.; Williams, K. H.; Banfield, J. F. (2017), Unusual respiratory capacity and nitrogen metabolism in a Parcubacterium (OD1) of the Candidate Phyla Radiation, Scientific Reports, 7, 40101 DOI: 10.1038/srep40101

Modeling Microbial Energetics and Community Dynamics

Mathematical framework of microbial models
can depend on the representative interacting unit. (Courtesy of Song, H.-S. et al., Processes, 2(4), 711, 2014,
Figure 2.)

Scientific Achievement

This chapter summarizes traditional thermodynamic- based microbial modeling approaches as well as next generation population-based methods that are aimed at improving our understanding of function, dynamics, and emergence in microbial ecosystems.

Significance and Impact

Modelers seeking to incorporate microbially mediated reactions into their codes will find a description of traditional and emergent approaches, example applications, and current limitations.

Research Details
  • Provided a description of traditional thermodynamic-based models and two emergent approaches, functional trait-based models and dynamic energy partitioning models, including example applications, for describing microbial community dynamics within the subsurface environment.
  • Presented an overview of current obstacles to implementation of these microbial population-based models and suggested opportunities for the integration of culture-independent (meta-omic) and culture-dependent experiment to structure and parameterize these models.
Citation

Arora, B., Cheng, Y., King, E., Bouskill, N. and Brodie, E., 2017. 27 Modeling Microbial Energetics. Handbook of Metal-Microbe Interactions and Bioremediation, p.445.

Impact of intra-meander hyporheic flow on nitrogen cycling

Fig. 1. (A) Lower East River catchment characterized by mountainous to rolling topography with multiple meanders; (B) DEM with 10 m resolution (solid yellow line) failed to capture meanders; however DEM and NHD dataset (solid red line) were able to mark exact locations of meanders.
Fig. 2. Nitrogen transformation in the hyporheic zone of two meanders of the lower East River.
Fig. 3. The extent of the hyporheic zone (on the right) is smaller when the high resolution NHD dataset is not used.

Scientific Achievement

We investigated the effect of redox gradients on nitrogen transformation in the subsurface by integrating a complex reaction network into PFLOTRAN, which is an open source, massively parallel, three-dimensional, reactive flow and transport code.

Significance and Impact

We evaluated how hyporheic exchanges and hydro-geomorphological characteristics together show varying levels of denitrification potential within two active meanders of the lower East River catchment in southwestern Colorado.

Research Details
  • The meander driven hyporheic exchanges increase denitrification rates because of relatively longer residence times in the organic carbon-rich sediments.
  • High-resolution data such as NHD provide an efficient strategy to correctly delineate meanders with computational tractability.
Citation

Dwivedi, D., C.I. Steefel, B. Arora, and G. Bisht (2017), Impact of intra-meander hyporheic flow on nitrogen cycling, Procedia Earth and Planetary Science, 17, 404-407, doi: 10.1016/j.proeps.2016.12.102.

Measurement of bacterial replication rates in microbial communities

Higher coverage near the origin than near the terminus, especially in fast (top) compared to slow growing populations (bottom).

Scientific Achievement

We developed an algorithm, iRep, that uses draft-quality genome sequences and single time-point metagenome sequencing to infer microbial population replication rates.

Significance and Impact

This broadly applicable method is now widely used to profile bacterial activity in situ and to track organism responses to varying conditions, information that finds application in modeling studies.

Research Details
  • A method that leverages information related to variation in genome coverage was developed to investigate in situ growth rates.
  • The method was tested using published experimental data and applied to study groundwater bacteria
Citation

Brown, C. T.; Olm, M. R.; Thomas, B. C.; Banfield, J. F. (2016) Measurement of bacterial replication rates in microbial communities, Nat Biotech, 34(12), 1256-1263 DOI: 10.1038/nbt.3704.

Seasonal hyporheic dynamics control coupled microbiology and geochemistry in Colorado River sediments

(Left) Map of hydraulic head in the floodplain aquifer across three sampling seasons. Samples for this study were collected in the river in the bottom left of the image. The black dots indicate groundwater monitoring well locations used for hydrologic calculations. (Upper right) River stage hydrograph for the Colorado River with sampling times indicated by blue lines. (Lower right) Magnitude (black line) and direction (red arrows) of Darcy flux through the floodplain aquifer near the sampling location based upon local hydraulic head calculations. Groundwater flow is generally south toward the river.

Scientific Achievement

Seasonal variations in snowmelt-driven river stage on the Colorado River drives strong redox fluctuations across a ~80-cm vertical portion of the riverbed, and results in the assembly of a unique ‘hyporheic’ microbiome

Significance and Impact

Biogeochemical fluctuations in the hyporheic zone are impacted by changes in snowmelt. An understanding of these dynamics is critical for predicting fluxes of metals, carbon, and other nutrients from this reactive hotspot

Research Details
  • Strong changes in snowmelt-driven river stage altered patterns of hyporheic mixing in the Colorado River, adjacent to the Rifle site.
  • This mixing modulated redox fronts in the riverbed, with greater dissolved iron concentrations at shallower depths during periods of low river discharge
  • Dynamic mixing and geochemical conditions in a ~80-cm vertical section of the riverbed resulted in a microbial community that was distinct from those in river water and groundwater
Citation

Danczak RE, AH Sawyer, KH Williams, JC Stegen, C Hobson, and MJ Wilkins. 2016. Seasonal hyporheic dynamics control coupled microbiology and geochemistry in Colorado River sediments. Journal of Geophysical Research – Biogeosciences. 121: 2976-2987, DOI: 10.1002/2016JG003527

A new wavelet-entropy approach reveals dominant controls on and distribution of geochemical hot moments at a contaminated floodplain site

Wavelet spectrum of chloride in i) contaminated aquifer and ii) naturally reduced zone of the Rifle site.

Scientific Achievement

A novel wavelet-entropy approach was used to identify the governing transport and biogeochemical factors causing geochemical hot moments and their distribution along different transects of a contaminated river floodplain.

Significance and Impact
  • Geochemical hot moments were found to be primarily transport-related/hydrologically-driven at the site except within a naturally reduced zone, where hot moments were dominated by lithologic characteristics.
  • This study presents a transferrable approach for water quality managers in identifying hot moments of contaminants in other settings.
Research Details
  • We interrogated complex, multivariate geochemical datasets to identify geochemical hot moments in a floodplain environment.
  • Geochemical hot moments were associated with seasonal hydrologic variability (∼4 months) in the contaminated aquifer and with annual hydrologic cycle and residence times (∼12 months) in the seep zone.
  • Hot moments associated with a naturally reduced zone within the aquifer were found to be biogeochemically-driven, with a different dominant frequency (∼3 months), in contrast to what was observed in other regions of the floodplain.
Citation

Arora, B., Dwivedi, D., Hubbard, S.S., Steefel, C.I. and Williams, K.H., 2016. Identifying geochemical hot moments and their controls on a contaminated river floodplain system using wavelet and entropy approaches. Environmental Modelling & Software, 85, pp.27-41. DOI: 10.1016/j.envsoft.2016.08.005