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Open Access Publications from the University of California

Scholarly Works (10 results)

  • Thesis
  • Peer Reviewed
UC Berkeley Electronic Theses and Dissertations (2024)

Recent advances in metagenomics have made it possible to resolve genomes of individual microorganisms from metagenomic data. This is known as genome-resolved metagenomics (GRM), and it has provided new insights into the ecological roles of individual microbial species within complex communities. GRM has enabled the discovery of novel microbial groups, including the Candidate Phyla Radiation (CPR) bacteria and the Asgard archaea. CPR bacteria are widespread and often live in symbiotic relationships with other microorganisms, while Asgard archaea are thought to be related to the ancestor of eukaryotic cells. My dissertation focuses on these newly resolved microbial groups, including consideration of their lifestyles, interactions, and evolution. Specific focus includes the close and specific inter-organism interactions (such as CPR bacteria and their host microorganisms) and evolutionary processes in archaea that may have laid the foundations for the development of multicellularity within the Asgard archaea. A central aspect of this research is the consideration of Mobile Genetic Elements (MGEs) that associate with these microbial groups and the biochemical mechanisms that regulate interactions between MGEs and their microbial hosts.

The first chapter of my dissertation focuses on Candidate Phyla Radiation (CPR) bacteria, which are commonly found in microbial communities but their role in biogeochemical cycles is not well understood. I studied biofilms that grow in sulfide-rich springs and found that these host diverse CPR bacteria. Ultra-small cells attached to the surfaces of chemolithotrophic sulfur-oxidizing filamentous bacteria are inferred to be CPR bacterial episymbionts (the first reported association of CPR with Proteobacteria). Some CPR bacteria have a novel electron bifurcating group 3b [NiFe]-hydrogenase, and other proteins potentially linked to sulfur and hydrogen metabolism. Thus, these CPR have the potential to directly impact biogeochemistry in groundwater systems. The second chapter focuses on hydrogenases—enzymes crucial for hydrogen cycling in anaerobic ecosystems. Through a combination of metagenomics, biochemistry, and genomics, this work reveals previously unknown [FeFe]-hydrogenases in novel archaeal lineages. This discovery not only expands our understanding of these vital enzymes but also highlights the metabolic versatility and evolutionary innovation within uncultivable archaeal phyla.

The third and fourth chapters focus on Asgard archaeal metabolism and mobile genetic elements. Through metagenomic analysis of wetland soil samples, I report two complete genomes for Atabeyarchaeota (after Atabey, a supreme goddess of freshwater and fertility in the Taíno religion from Puerto Rico), a new Asgard archaeal lineage, and the first complete genome of a Freyarchaeota, providing insights into their metabolic potential, evolutionary history, and ecological roles. I also investigated their diverse mobile genetic elements, which contribute to microbial adaptation and evolution. This is the first focused study of Asgard MGEs and thus of the processes that drive genome divergence and population dynamics in the modern lineages. The fifth chapter discusses the challenges and opportunities in archaeal genomics, emphasizing the limitations of our current knowledge due to the scarcity of cultivated strains and complete genomes. With the advent of long-read sequencing technologies, such as PacBio and Nanopore, I describe my efforts in sequencing wetland soil samples, which led to the recovery of hundreds of circular genomes. This endeavor not only highlights the diversity and metabolic potential of archaea but also underscores the importance of advanced sequencing technologies in uncovering the hidden diversity of microbial life.

In summary, these studies provide new insights into the diversity and evolution of CPR bacteria and Asgard archaea, as well as the role of mobile genetic elements in shaping microbial genomes. The application of genome-resolved metagenomics has enabled a more comprehensive understanding of microbial communities and their functions in complex environments, which has important implications for fields such as microbial ecology, evolution, biotechnology, and population dynamics.

    Cover page: Microbial Tapestry: Interwoven activities of novel uncultivated microorganisms, their genome structures, and mobile genetic elements in subsurface ecosystems
    • Article
    • Peer Reviewed
    UC Berkeley Previously Published Works (2023)
    Carbon fixation is the process by which CO2 is converted from a gas into biomass. The Calvin-Benson-Bassham cycle (CBB) is the dominant carbon-consuming pathway on Earth, driving >99.5% of the ∼120 billion tons of carbon that are converted to sugar by plants, algae, and cyanobacteria. The carboxylase enzyme in the CBB, ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco), fixes one CO2 molecule per turn of the cycle into bioavailable sugars. Despite being critical to the assimilation of carbon, rubisco's kinetic rate is not very fast, limiting flux through the pathway. This bottleneck presents a paradox: Why has rubisco not evolved to be a better catalyst? Many hypothesize that the catalytic mechanism of rubisco is subject to one or more trade-offs and that rubisco variants have been optimized for their native physiological environment. Here, we review the evolution and biochemistry of rubisco through the lens of structure and mechanism in order to understand what trade-offs limit its improvement. We also review the many attempts to improve rubisco itself and thereby promote plant growth.
      • Article
      • Peer Reviewed
      UC Berkeley Previously Published Works (2021)
      Prokaryotic nanocompartments, also known as encapsulins, are a recently discovered proteinaceous organelle-like compartment in prokaryotes that compartmentalize cargo enzymes. While initial studies have begun to elucidate the structure and physiological roles of encapsulins, bioinformatic evidence suggests that a great diversity of encapsulin nanocompartments remains unexplored. Here, we describe a novel encapsulin in the freshwater cyanobacterium Synechococcus elongatus PCC 7942. This nanocompartment is upregulated upon sulfate starvation and encapsulates a cysteine desulfurase enzyme via an N-terminal targeting sequence. Using cryo-electron microscopy, we have determined the structure of the nanocompartment complex to 2.2 Å resolution. Lastly, biochemical characterization of the complex demonstrated that the activity of the cysteine desulfurase is enhanced upon encapsulation. Taken together, our discovery, structural analysis, and enzymatic characterization of this prokaryotic nanocompartment provide a foundation for future studies seeking to understand the physiological role of this encapsulin in various bacteria.
        Cover page: Discovery and characterization of a novel family of prokaryotic nanocompartments involved in sulfur metabolismCreative Commons 'BY' version 4.0 license
        • Article
        • Peer Reviewed
        UC Berkeley Previously Published Works (2020)
        Despite very low concentrations of cobalt in marine waters, cyanobacteria in the genus Prochlorococcus retain the genetic machinery for the synthesis and use of cobalt-bearing cofactors (cobalamins) in their genomes. We explore cobalt metabolism in a Prochlorococcus isolate from the equatorial Pacific Ocean (strain MIT9215) through a series of growth experiments under iron- and cobalt-limiting conditions. Metal uptake rates, quantitative proteomic measurements of cobalamin-dependent enzymes, and theoretical calculations all indicate that Prochlorococcus MIT9215 can sustain growth with less than 50 cobalt atoms per cell, ∼100-fold lower than minimum iron requirements for these cells (∼5,100 atoms per cell). Quantitative descriptions of Prochlorococcus cobalt limitation are used to interpret the cobalt distribution in the equatorial Pacific Ocean, where surface concentrations are among the lowest measured globally but Prochlorococcus biomass is high. A low minimum cobalt quota ensures that other nutrients, notably iron, will be exhausted before cobalt can be fully depleted, helping to explain the persistence of cobalt-dependent metabolism in marine cyanobacteria.
          Cover page: Minimal cobalt metabolism in the marine cyanobacterium Prochlorococcus
          • Article
          • Peer Reviewed
          UC Berkeley Previously Published Works (2024)
          The roles of Asgard archaea in eukaryogenesis and marine biogeochemical cycles are well studied, yet their contributions in soil ecosystems remain unknown. Of particular interest are Asgard archaeal contributions to methane cycling in wetland soils. To investigate this, we reconstructed two complete genomes for soil-associated Atabeyarchaeia, a new Asgard lineage, and a complete genome of Freyarchaeia, and predicted their metabolism in situ. Metatranscriptomics reveals expression of genes for [NiFe]-hydrogenases, pyruvate oxidation and carbon fixation via the Wood-Ljungdahl pathway. Also expressed are genes encoding enzymes for amino acid metabolism, anaerobic aldehyde oxidation, hydrogen peroxide detoxification and carbohydrate breakdown to acetate and formate. Overall, soil-associated Asgard archaea are predicted to include non-methanogenic acetogens, highlighting their potential role in carbon cycling in terrestrial environments.
            Cover page: Asgard archaea modulate potential methanogenesis substrates in wetland soil.Creative Commons 'BY' version 4.0 license
            • Article
            • Peer Reviewed
            UC Berkeley Previously Published Works (2021)
            Candidate Phyla Radiation (CPR) bacteria are small, likely episymbiotic organisms found across Earth's ecosystems. Despite their prevalence, the distribution of CPR lineages across habitats and the genomic signatures of transitions among these habitats remain unclear. Here, we expand the genome inventory for Absconditabacteria (SR1), Gracilibacteria, and Saccharibacteria (TM7), CPR bacteria known to occur in both animal-associated and environmental microbiomes, and investigate variation in gene content with habitat of origin. By overlaying phylogeny with habitat information, we show that bacteria from these three lineages have undergone multiple transitions from environmental habitats into animal microbiomes. Based on co-occurrence analyses of hundreds of metagenomes, we extend the prior suggestion that certain Saccharibacteria have broad bacterial host ranges and constrain possible host relationships for Absconditabacteria and Gracilibacteria. Full-proteome analyses show that animal-associated Saccharibacteria have smaller gene repertoires than their environmental counterparts and are enriched in numerous protein families, including those likely functioning in amino acid metabolism, phage defense, and detoxification of peroxide. In contrast, some freshwater Saccharibacteria encode a putative rhodopsin. For protein families exhibiting the clearest patterns of differential habitat distribution, we compared protein and species phylogenies to estimate the incidence of lateral gene transfer and genomic loss occurring over the species tree. These analyses suggest that habitat transitions were likely not accompanied by large transfer or loss events but rather were associated with continuous proteome remodeling. Thus, we speculate that CPR habitat transitions were driven largely by availability of suitable host taxa and were reinforced by acquisition and loss of some capacities. IMPORTANCE Studying the genetic differences between related microorganisms from different environment types can indicate factors associated with their movement among habitats. This is particularly interesting for bacteria from the Candidate Phyla Radiation because their minimal metabolic capabilities require associations with microbial hosts. We found that shifts of Absconditabacteria, Gracilibacteria, and Saccharibacteria between environmental ecosystems and mammalian mouths/guts probably did not involve major episodes of gene gain and loss; rather, gradual genomic change likely followed habitat migration. The results inform our understanding of how little-known microorganisms establish in the human microbiota where they may ultimately impact health.
              Cover page: Patterns of Gene Content and Co-occurrence Constrain the Evolutionary Path toward Animal Association in Candidate Phyla Radiation BacteriaCreative Commons 'BY' version 4.0 license
              • Article
              • Peer Reviewed
              UC Berkeley Previously Published Works (2024)
              BACKGROUND: Underground research laboratories (URLs) provide a window on the deep biosphere and enable investigation of potential microbial impacts on nuclear waste, CO2 and H2 stored in the subsurface. We carried out the first multi-year study of groundwater microbiomes sampled from defined intervals between 140 and 400 m below the surface of the Horonobe and Mizunami URLs, Japan. RESULTS: We reconstructed draft genomes for > 90% of all organisms detected over a four year period. The Horonobe and Mizunami microbiomes are dissimilar, likely because the Mizunami URL is hosted in granitic rock and the Horonobe URL in sedimentary rock. Despite this, hydrogen metabolism, rubisco-based CO2 fixation, reduction of nitrogen compounds and sulfate reduction are well represented functions in microbiomes from both URLs, although methane metabolism is more prevalent at the organic- and CO2-rich Horonobe URL. High fluid flow zones and proximity to subsurface tunnels select for candidate phyla radiation bacteria in the Mizunami URL. We detected near-identical genotypes for approximately one third of all genomically defined organisms at multiple depths within the Horonobe URL. This cannot be explained by inactivity, as in situ growth was detected for some bacteria, albeit at slow rates. Given the current low hydraulic conductivity and groundwater compositional heterogeneity, ongoing inter-site strain dispersal seems unlikely. Alternatively, the Horonobe URL microbiome homogeneity may be explained by higher groundwater mobility during the last glacial period. Genotypically-defined species closely related to those detected in the URLs were identified in three other subsurface environments in the USA. Thus, dispersal rates between widely separated underground sites may be fast enough relative to mutation rates to have precluded substantial divergence in species composition. Species overlaps between subsurface locations on different continents constrain expectations regarding the scale of global subsurface biodiversity. CONCLUSIONS: Our analyses reveal microbiome stability in the sedimentary rocks and surprising microbial community compositional and genotypic overlap over sites separated by hundreds of meters of rock, potentially explained by dispersal via slow groundwater flow or during a prior hydrological regime. Overall, microbiome and geochemical stability over the study period has important implications for underground storage applications.
                Cover page: Diverse microbiome functions, limited temporal variation and substantial genomic conservation within sedimentary and granite rock deep underground research laboratories.
                • Article
                • Peer Reviewed
                UC Berkeley Previously Published Works (2022)
                Anaerobic methane oxidation exerts a key control on greenhouse gas emissions1, yet factors that modulate the activity of microorganisms performing this function remain poorly understood. Here we discovered extraordinarily large, diverse DNA sequences that primarily encode hypothetical proteins through studying groundwater, sediments and wetland soil where methane production and oxidation occur. Four curated, complete genomes are linear, up to approximately 1 Mb in length and share genome organization, including replichore structure, long inverted terminal repeats and genome-wide unique perfect tandem direct repeats that are intergenic or generate amino acid repeats. We infer that these are highly divergent archaeal extrachromosomal elements with a distinct evolutionary origin. Gene sequence similarity, phylogeny and local divergence of sequence composition indicate that many of their genes were assimilated from methane-oxidizing Methanoperedens archaea. We refer to these elements as 'Borgs'. We identified at least 19 different Borg types coexisting with Methanoperedens spp. in four distinct ecosystems. Borgs provide methane-oxidizing Methanoperedens archaea access to genes encoding proteins involved in redox reactions and energy conservation (for example, clusters of multihaem cytochromes and methyl coenzyme M reductase). These data suggest that Borgs might have previously unrecognized roles in the metabolism of this group of archaea, which are known to modulate greenhouse gas emissions, but further studies are now needed to establish their functional relevance.
                  Cover page: Borgs are giant genetic elements with potential to expand metabolic capacityCreative Commons 'BY' version 4.0 license
                  • Article
                  • Peer Reviewed
                  UC Berkeley Previously Published Works (2024)

                  Background

                  Biofilms in sulfide-rich springs present intricate microbial communities that play pivotal roles in biogeochemical cycling. We studied chemoautotrophically based biofilms that host diverse CPR bacteria and grow in sulfide-rich springs to investigate microbial controls on biogeochemical cycling.

                  Results

                  Sulfide springs biofilms were investigated using bulk geochemical analysis, genome-resolved metagenomics, and scanning transmission X-ray microscopy (STXM) at room temperature and 87 K. Chemolithotrophic sulfur-oxidizing bacteria, including Thiothrix and Beggiatoa, dominate the biofilms, which also contain CPR Gracilibacteria, Absconditabacteria, Saccharibacteria, Peregrinibacteria, Berkelbacteria, Microgenomates, and Parcubacteria. STXM imaging revealed ultra-small cells near the surfaces of filamentous bacteria that may be CPR bacterial episymbionts. STXM and NEXAFS spectroscopy at carbon K and sulfur L2,3 edges show that filamentous bacteria contain protein-encapsulated spherical elemental sulfur granules, indicating that they are sulfur oxidizers, likely Thiothrix. Berkelbacteria and Moranbacteria in the same biofilm sample are predicted to have a novel electron bifurcating group 3b [NiFe]-hydrogenase, putatively a sulfhydrogenase, potentially linked to sulfur metabolism via redox cofactors. This complex could potentially contribute to symbioses, for example, with sulfur-oxidizing bacteria such as Thiothrix that is based on cryptic sulfur cycling. One Doudnabacteria genome encodes adjacent sulfur dioxygenase and rhodanese genes that may convert thiosulfate to sulfite. We find similar conserved genomic architecture associated with CPR bacteria from other sulfur-rich subsurface ecosystems.

                  Conclusions

                  Our combined metagenomic, geochemical, spectromicroscopic, and structural bioinformatics analyses of biofilms growing in sulfide-rich springs revealed consortia that contain CPR bacteria and sulfur-oxidizing Proteobacteria, including Thiothrix, and bacteria from a new family within Beggiatoales. We infer roles for CPR bacteria in sulfur and hydrogen cycling. Video Abstract.
                    Cover page: Autotrophic biofilms sustained by deeply sourced groundwater host diverse bacteria implicated in sulfur and hydrogen metabolismCreative Commons 'BY' version 4.0 license
                    • Article
                    • Peer Reviewed
                    UC Berkeley Previously Published Works (2021)
                    Regular surveillance testing of asymptomatic individuals for SARS-CoV-2 has been center to SARS-CoV-2 outbreak prevention on college and university campuses. Here we describe the voluntary saliva testing program instituted at the University of California, Berkeley during an early period of the SARS-CoV-2 pandemic in 2020. The program was administered as a research study ahead of clinical implementation, enabling us to launch surveillance testing while continuing to optimize the assay. Results of both the testing protocol itself and the study participants' experience show how the program succeeded in providing routine, robust testing capable of contributing to outbreak prevention within a campus community and offer strategies for encouraging participation and a sense of civic responsibility.
                      Cover page: Launching a saliva-based SARS-CoV-2 surveillance testing program on a university campus
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