College of Agriculture
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As the foundation of the land grant mission at Montana State University, the College of Agriculture and the Montana Agricultural Experiment Station provide instruction in traditional and innovative degree programs and conduct research on old and new challenges for Montana’s agricultural community. This integration creates opportunities for students and faculty to excel through hands-on learning, to serve through campus and community engagement, to explore unique solutions to distinct and interesting questions and to connect Montanans with the global community through research discoveries and outreach.
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Item Sulfur cycling and host-virus interactions in Aquificales-dominated biofilms from Yellowstone’s hottest ecosystems(Springer Nature, 2021-10) McKay, Luke J.; Nigro, Olivia D.; Dlakić, Mensur; Luttrell, Karen M.; Rusch, Douglas B.; Fields, Matthew W.; Inskeep, William P.Modern linkages among magmatic, geochemical, and geobiological processes provide clues about the importance of thermophiles in the origin of biogeochemical cycles. The aim of this study was to identify the primary chemoautotrophs and host–virus interactions involved in microbial colonization and biogeochemical cycling at sublacustrine, vapor-dominated vents that represent the hottest measured ecosystems in Yellowstone National Park (~140 °C). Filamentous microbial communities exposed to extreme thermal and geochemical gradients were sampled using a remotely operated vehicle and subjected to random metagenome sequencing and microscopic analyses. Sulfurihydrogenibium (phylum Aquificae) was the predominant lineage (up to 84% relative abundance) detected at vents that discharged high levels of dissolved H2, H2S, and CO2. Metabolic analyses indicated carbon fixation by Sulfurihydrogenibium spp. was powered by the oxidation of reduced sulfur and H2, which provides organic carbon for heterotrophic community members. Highly variable Sulfurihydrogenibium genomes suggested the importance of intra-population diversity under extreme environmental and viral pressures. Numerous lytic viruses (primarily unclassified taxa) were associated with diverse archaea and bacteria in the vent community. Five circular dsDNA uncultivated virus genomes (UViGs) of ~40 kbp length were linked to the Sulfurihydrogenibium metagenome-assembled genome (MAG) by CRISPR spacer matches. Four UViGs contained consistent genome architecture and formed a monophyletic cluster with the recently proposed Pyrovirus genus within the Caudovirales. Sulfurihydrogenibium spp. also contained CRISPR arrays linked to plasmid DNA with genes for a novel type IV filament system and a highly expressed β-barrel porin. A diverse suite of transcribed secretion systems was consistent with direct microscopic analyses, which revealed an extensive extracellular matrix likely critical to community structure and function. We hypothesize these attributes are fundamental to the establishment and survival of microbial communities in highly turbulent, extreme-gradient environments.Item Roadmap for naming uncultivated Archaea and Bacteria(2020-08) Murray, Alison E.; Freudenstein, John; Gribaldo, Simonetta; Hatzenpichler, Roland; Hugenholtz, Philip; Kampfer, Peter; Konstantinidis, Konstantinos T.; Lane, Christopher E.; Papke, R. Thane; Parks, Donovan H.; Rossello-Mora, Ramon; Stott, Matthew B.; Sutcliffe, Iain C.; Thrash, J. Cameron; Venter, Stephanus N.; Whitman, William B.; Acinas, Silvia G.; Amann, Rudolf I.; Anantharaman, Karthik; Armengaud, Jean; Baker, Brett J.; Barco, Roman A.; Bode, Helge B.; Boyd, Eric S.; Brady, Carrie L.; Carini, Paul; Chain, Patrick S. G.; Colman, Daniel R.; DeAngelis, Kristen M.; Asuncion de los Rios, Maria; Estrada-de los Santos, Paulina; Dunlap, Christopher A.; Eisen, Jonathan A.; Emerson, David; Ettema, Thisjs J. G.; Eveillard, Damien R.; Girguis, Peter R.; Hentschel, Ute; Hollibaugh, James T.; Hug, Laura A.; Inskeep, William P.; Ivanova, Elena P.; Klenk, Hans-Peter; Li, Wen-Jun; Lloyd, Karen G.; Loffler, Frank E.; Makhalanyane, Thulani P.; Moser, Duane P.; Nunoura, Takuro; Palmer, Marike; Parro, Victor; Pedros-Alio, Carlos; Probst, Alexander J.; Smits, Theo H. M.; Steen, Andrew D.; Steenkamp, Emma T.; Spang, Anja; Stewart, Frank J.; Tiedje, James M.; Vandamme, Peter; Wagner, Michael; Wang, Feng-Ping; Yarza, Pablo; Hedlund, Brian P.; Reysenbach, Anna-LouiseThe assembly of single-amplified genomes (SAGs) and metagenome-assembled genomes (MAGs) has led to a surge in genome-based discoveries of members affiliated with Archaea and Bacteria, bringing with it a need to develop guidelines for nomenclature of uncultivated microorganisms. The International Code of Nomenclature of Prokaryotes (ICNP) only recognizes cultures as ‘type material’, thereby preventing the naming of uncultivated organisms. In this Consensus Statement, we propose two potential paths to solve this nomenclatural conundrum. One option is the adoption of previously proposed modifications to the ICNP to recognize DNA sequences as acceptable type material; the other option creates a nomenclatural code for uncultivated Archaea and Bacteria that could eventually be merged with the ICNP in the future. Regardless of the path taken, we believe that action is needed now within the scientific community to develop consistent rules for nomenclature of uncultivated taxa in order to provide clarity and stability, and to effectively communicate microbial diversity.Item Physiology, Metabolism, and Fossilization of Hot-Spring Filamentous Microbial Mats(2019-04) Dong, Yiran; Sanford, Robert A.; Inskeep, William P.; Srivastava, Vaibhav; Bulone, Vincent; Fields, Christopher J.; Yau, Peter M.; Sivaguru, Mayandi; Ahren, Dag; Fouke, Kyle W.; Weber, Joseph; Werth, Charles R.; Cann, Isaac K.; Keating, Kathleen M.; Khetani, Radhika S.; Hernandez, Alvaro G.; Wright, Chris; Band, Mark; Imai, Brian S.; Fried, Glenn A.; Fouke, Bruce W.The evolutionarily ancient Aquificales bacterium Sulfurihydrogenibium spp. dominates filamentous microbial mat communities in shallow, fast-flowing, and dysoxic hot-spring drainage systems around the world. In the present study, field observations of these fettuccini-like microbial mats at Mammoth Hot Springs in Yellowstone National Park are integrated with geology, geochemistry, hydrology, microscopy, and multi-omic molecular biology analyses. Strategic sampling of living filamentous mats along with the hot-spring CaCO3 (travertine) in which they are actively being entombed and fossilized has permitted the first direct linkage of Sulfurihydrogenibium spp. physiology and metabolism with the formation of distinct travertine streamer microbial biomarkers. Results indicate that, during chemoautotrophy and CO2 carbon fixation, the 87-98% Sulfurihydrogenibium-dominated mats utilize chaperons to facilitate enzyme stability and function. High-abundance transcripts and proteins for type IV pili and extracellular polymeric substances (EPSs) are consistent with their strong mucus-rich filaments tens of centimeters long that withstand hydrodynamic shear as they become encrusted by more than 5mm of travertine per day. Their primary energy source is the oxidation of reduced sulfur (e.g., sulfide, sulfur, or thiosulfate) and the simultaneous uptake of extremely low concentrations of dissolved O2 facilitated by bd-type cytochromes. The formation of elevated travertine ridges permits the Sulfurihydrogenibium-dominated mats to create a shallow platform from which to access low levels of dissolved oxygen at the virtual exclusion of other microorganisms. These ridged travertine streamer microbial biomarkers are well preserved and create a robust fossil record of microbial physiological and metabolic activities in modern and ancient hot-spring ecosystems.Item Metagenomes from high-temperature chemotrophic systems reveal geochemical controls on microbial community structure and function(2010-03) Inskeep, William P.; Rusch, Douglas B.; Jay, Zackary J.; Herrgard, Markus J.; Kozubal, Mark A.; Richardson, Toby H.; Macur, Richard E.; Hamamura, Natsuko; Jennings, Ryan deM.; Fouke, Bruce W.; Reysenbach, Anna-Louise; Roberto, Frank; Young, Mark J.; Schwartz, Ariel; Boyd, Eric S.; Badger, Jonathan H.; Mathur, Eric J.; Ortmann, Alice C.; Bateson, Mary M.; Geesey, Gill G.; Frazier, MarvinThe Yellowstone caldera contains the most numerous and diverse geothermal systems on Earth, yielding an extensive array of unique high-temperature environments that host a variety of deeply-rooted and understudied Archaea, Bacteria and Eukarya. The combination of extreme temperature and chemical conditions encountered in geothermal environments often results in considerably less microbial diversity than other terrestrial habitats and offers a tremendous opportunity for studying the structure and function of indigenous microbial communities and for establishing linkages between putative metabolisms and element cycling. Metagenome sequence (14–15,000 Sanger reads per site) was obtained for five high-temperature (>65°C) chemotrophic microbial communities sampled from geothermal springs (or pools) in Yellowstone National Park (YNP) that exhibit a wide range in geochemistry including pH, dissolved sulfide, dissolved oxygen and ferrous iron. Metagenome data revealed significant differences in the predominant phyla associated with each of these geochemical environments. Novel members of the Sulfolobales are dominant in low pH environments, while other Crenarchaeota including distantly-related Thermoproteales and Desulfurococcales populations dominate in suboxic sulfidic sediments. Several novel archaeal groups are well represented in an acidic (pH 3) Fe-oxyhydroxide mat, where a higher O2 influx is accompanied with an increase in archaeal diversity. The presence or absence of genes and pathways important in S oxidation-reduction, H2-oxidation, and aerobic respiration (terminal oxidation) provide insight regarding the metabolic strategies of indigenous organisms present in geothermal systems. Multiple-pathway and protein-specific functional analysis of metagenome sequence data corroborated results from phylogenetic analyses and clearly demonstrate major differences in metabolic potential across sites. The distribution of functional genes involved in electron transport is consistent with the hypothesis that geochemical parameters (e.g., pH, sulfide, Fe, O2) control microbial community structure and function in YNP geothermal springs.Item Multiscale analysis of autotroph-heterotroph interactions in a high-temperature microbial community(2018-09) Hunt, Kristopher A.; Jennings, Ryan deM.; Inskeep, William P.; Carlson, Ross P.Interactions among microbial community members can lead to emergent properties, such as enhanced productivity, stability, and robustness. Iron-oxide mats in acidic (pH 2–4), high-temperature (> 65 °C) springs of Yellowstone National Park contain relatively simple microbial communities and are well-characterized geochemically. Consequently, these communities are excellent model systems for studying the metabolic activity of individual populations and key microbial interactions. The primary goals of the current study were to integrate data collected in situ with in silico calculations across process-scales encompassing enzymatic activity, cellular metabolism, community interactions, and ecosystem biogeochemistry, as well as to predict and quantify the functional limits of autotroph-heterotroph interactions. Metagenomic and transcriptomic data were used to reconstruct carbon and energy metabolisms of an important autotroph (Metallosphaera yellowstonensis) and heterotroph (Geoarchaeum sp. OSPB) from the studied Fe(III)-oxide mat communities. Standard and hybrid elementary flux mode and flux balance analyses of metabolic models predicted cellular- and community-level metabolic acclimations to simulated environmental stresses, respectively. In situ geochemical analyses, including oxygen depth-profiles, Fe(III)-oxide deposition rates, stable carbon isotopes and mat biomass concentrations, were combined with cellular models to explore autotroph-heterotroph interactions important to community structure-function. Integration of metabolic modeling with in situ measurements, including the relative population abundance of autotrophs to heterotrophs, demonstrated that Fe(III)-oxide mat communities operate at their maximum total community growth rate (i.e. sum of autotroph and heterotroph growth rates), as opposed to net community growth rate (i.e. total community growth rate subtracting autotroph consumed by heterotroph), as predicted from the maximum power principle. Integration of multiscale data with ecological theory provides a basis for predicting autotroph-heterotroph interactions and community-level cellular organization.Item Life in High-Temperature Environments: Modern-Day Analogs of Early Earth Still Relevant Today(2017-09) Inskeep, William P.The discovery of new single-celled organism lineages has been remarkable since the adoption of molecular genetics and the discovery of the domain Archaea (Woese et al., 1990). Prior to molecular techniques that initially emphasized the sequences of ribosomal genes (e.g., 16S rRNA), the discipline of microbiology relied nearly entirely on cultivation and the ability to grow a specific microorganism in pure culture under defined conditions. This meant that only microorganisms that grew easily under laboratory conditions were cultivated, and, in many cases, these often rapidly growing organisms do not correspond to the more numerous and relevant microbes that actually inhabit different microbiomes. We now appreciate that many of the microorganisms easily grown under laboratory conditions are often related to their more abundant and important relatives found in situ, but they generally do not exhibit the same functional attributes as numerically relevant microorganisms.Item Marsarchaeota are an aerobic archaeal lineage abundant in geothermal iron oxide microbial mats(2018-05) Jay, Zackary J.; Beam, Jacob P.; Dlakic, Mensur; Rusch, Douglas B.; Kozubal, Mark A.; Inskeep, William P.The discovery of archaeal lineages is critical to our understanding of the universal tree of life and evolutionary history of the Earth. Geochemically diverse thermal environments in Yellowstone National Park provide unprecedented opportunities for studying archaea in habitats that may represent analogues of early Earth. Here, we report the discovery and characterization of a phylum-level archaeal lineage proposed and herein referred to as the \'Marsarchaeota\', after the red planet. The Marsarchaeota contains at least two major subgroups prevalent in acidic, microaerobic geothermal Fe(III) oxide microbial mats across a temperature range from similar to 50-80 degrees C. Metagenomics, single-cell sequencing, enrichment culturing and in situ transcriptional analyses reveal their biogeochemical role as facultative aerobic chemoorganotrophs that may also mediate the reduction of Fe(III). Phylogenomic analyses of replicate assemblies corresponding to two groups of Marsarchaeota indicate that they branch between the Crenarchaeota and all other major archaeal lineages. Transcriptomic analyses of several Fe(III) oxide mat communities reveal that these organisms were actively transcribing two different terminal oxidase complexes in situ and genes comprising an F-420-dependent butanal catabolism. The broad distribution of Marsarchaeota in geothermal, microaerobic Fe(III) oxide mats suggests that similar habitat types probably played an important role in the evolution of archaea.Item Integration of metagenomic and stable carbon isotope evidence reveals the extent and mechanisms of carbon dioxide fixation in high-temperature microbial communities.(2017-02) Jennings, Ryan deM.; Moran, James J.; Jay, Zackary J.; Beam, Jacob P.; Whitmore, Laura M.; Kozubal, Mark A.; Kreuzer, Helen W.; Inskeep, William P.Although the biological fixation of CO2 by chemolithoautotrophs provides a diverse suite of organic compounds utilized by chemoorganoheterotrophs as a carbon and energy source, the relative amounts of autotrophic C in chemotrophic microbial communities are not well-established. The extent and mechanisms of CO2 fixation were evaluated across a comprehensive set of high-temperature, chemotrophic microbial communities in Yellowstone National Park by combining metagenomic and stable 13C isotope analyses. Fifteen geothermal sites representing three distinct habitat types (iron-oxide mats, anoxic sulfur sediments, and filamentous “streamer” communities) were investigated. Genes of the 3-hydroxypropionate/4-hydroxybutyrate, dicarboxylate/4-hydroxybutyrate, and reverse tricarboxylic acid CO2 fixation pathways were identified in assembled genome sequence corresponding to the predominant Crenarchaeota and Aquificales observed across this habitat range. Stable 13C analyses of dissolved inorganic and organic C (DIC, DOC), and possible landscape C sources were used to interpret the 13C content of microbial community samples. Isotope mixing models showed that the minimum fractions of autotrophic C in microbial biomass were >50% in the majority of communities analyzed. The significance of CO2 as a C source in these communities provides a foundation for understanding community assembly and succession, and metabolic linkages among early-branching thermophilic autotrophs and heterotrophs.Item Formaldehyde as a carbon and electron shuttle between autotroph and heterotroph populations in acidic hydrothermal vents of Norris Geyser Basin, Yellowstone National Park(2016-05) Moran, James J.; Whitmore, Laura M.; Isern, Nancy G.; Romine, Margaret F.; Riha, Krystin M.; Inskeep, William P.; Kreuzer, Helen W.The Norris Geyser Basin in Yellowstone National Park contains a large number of hydrothermal systems, which host microbial populations supported by primary productivity associated with a suite of chemolithotrophic metabolisms. We demonstrate that Metallosphaera yellowstonensis MK1, a facultative autotrophic archaeon isolated from a hyperthermal acidic hydrous ferric oxide (HFO) spring in Norris Geyser Basin, excretes formaldehyde during autotrophic growth. To determine the fate of formaldehyde in this low organic carbon environment, we incubated native microbial mat (containing M. yellowstonensis) from a HFO spring with 13C-formaldehyde. Isotopic analysis of incubation-derived CO2 and biomass showed that formaldehyde was both oxidized and assimilated by members of the community. Autotrophy, formaldehyde oxidation, and formaldehyde assimilation displayed different sensitivities to chemical inhibitors, suggesting that distinct sub-populations in the mat selectively perform these functions. Our results demonstrate that electrons originally resulting from iron oxidation can energetically fuel autotrophic carbon fixation and associated formaldehyde excretion, and that formaldehyde is both oxidized and assimilated by different organisms within the native microbial community. Thus, formaldehyde can effectively act as a carbon and electron shuttle connecting the autotrophic, iron oxidizing members with associated heterotrophic members in the HFO community.Item Geochemistry and Mixing Drive the Spatial Distribution of Free-Living Archaea and Bacteria in Yellowstone Lake(2016-02) Kan, Jinjun; Clingenpeel, Scott; Dow, Charles L.; McDermott, Timothy R.; Macur, Richard E.; Inskeep, William P.; Nealson, Kenneth H.Yellowstone Lake, the largest subalpine lake in the United States, harbors great novelty and diversity of Bacteria and Archaea. Size-fractionated water samples (0.1–0.8, 0.8–3.0, and 3.0–20 μm) were collected from surface photic zone, deep mixing zone, and vent fluids at different locations in the lake by using a remotely operated vehicle (ROV). Quantification with real-time PCR indicated that Bacteria dominated free-living microorganisms with Bacteria/Archaea ratios ranging from 4037:1 (surface water) to 25:1 (vent water). Microbial population structures (both Bacteria and Archaea) were assessed using 454-FLX sequencing with a total of 662,302 pyrosequencing reads for V1 and V2 regions of 16S rRNA genes. Non-metric multidimensional scaling (NMDS) analyses indicated that strong spatial distribution patterns existed from surface to deep vents for free-living Archaea and Bacteria in the lake. Along with pH, major vent-associated geochemical constituents including CH4, CO2, H2, DIC (dissolved inorganic carbon), DOC (dissolved organic carbon), SO42-, O2 and metals were likely the major drivers for microbial population structures, however, mixing events occurring in the lake also impacted the distribution patterns. Distinct Bacteria and Archaea were present among size fractions, and bigger size fractions included particle-associated microbes (> 3 μm) and contained higher predicted operational taxonomic unit richness and microbial diversities (genus level) than free-living ones (<0.8 μm). Our study represents the first attempt at addressing the spatial distribution of Bacteria and Archaea in Yellowstone Lake, and our results highlight the variable contribution of Archaea and Bacteria to the hydrogeochemical-relevant metabolism of hydrogen, carbon, nitrogen, and sulfur.