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 Aerobic methane synthesis and dynamics in a river water environment(Wiley, 2023-06) Alowaifeer, Abdullah M.; Wang, Qian; Bothner, Brian; Sibert, Ryan J.; Joye, Samantha B.; McDermott, Timothy R.Reports of aerobic biogenic methane (CH4) have generated new views about CH4 sources in nature. We examine this phenomenon in the free-flowing Yellowstone river wherein CH4 concentrations were tracked as a function of environmental conditions, phototrophic microorganisms (using chlorophyll a, Chl a, as proxy), as well as targeted methylated amines known to be associated with this process. CH4 was positively correlated with temperature and Chl a, although diurnal measurements showed CH4 concentrations were greatest during the night and lowest during maximal solar irradiation. CH4 efflux from the river surface was greater in quiescent edge waters (71–94 μmol m−2 d) than from open flowing current (~ 57 μmol m−2 d). Attempts to increase flux by disturbing the benthic environment in the quiescent water directly below (~ 1.0 m deep) or at varying distances (0–5 m) upstream of the flux chamber failed to increase surface flux. Glycine betaine (GB), dimethylamine and methylamine (MMA) were observed throughout the summer-long study, increasing during a period coinciding with a marked decline in Chl a, suggesting a lytic event led to their release; however, this did not correspond to increased CH4 concentrations. Spiking river water with GB or MMA yielded significantly greater CH4 than nonspiked controls, illustrating the metabolic potential of the river microbiome. In summary, this study provides evidence that: (1) phototrophic microorganisms are involved in CH4 synthesis in a river environment; (2) the river microbiome possesses the metabolic potential to convert methylated amines to CH4; and (3) river CH4 concentrations are dynamic diurnally as well as during the summer active months.Item Aerobic bacterial methane synthesis(Proceedings of the National Academy of Sciences, 2021-06) Wang, Qian; Alowaifeer, Abdullah; Kerner, Patricia; Balasubramanian, Narayanaganesh; Patterson, Angela; Christian, William; Tarver, Angela; Dore, John E.; Hatzenpichler, Roland; Bothner, Brian; McDermott, Timothy R.Reports of biogenic methane (CH4) synthesis associated with a range of organisms have steadily accumulated in the literature. This has not happened without controversy and in most cases the process is poorly understood at the gene and enzyme levels. In marine and freshwater environments, CH4 supersaturation of oxic surface waters has been termed the “methane paradox” because biological CH4 synthesis is viewed to be a strictly anaerobic process carried out by O2-sensitive methanogens. Interest in this phenomenon has surged within the past decade because of the importance of understanding sources and sinks of this potent greenhouse gas. In our work on Yellowstone Lake in Yellowstone National Park, we demonstrate microbiological conversion of methylamine to CH4 and isolate and characterize an Acidovorax sp. capable of this activity. Furthermore, we identify and clone a gene critical to this process (encodes pyridoxylamine phosphate-dependent aspartate aminotransferase) and demonstrate that this property can be transferred to Escherichia coli with this gene and will occur as a purified enzyme. This previously unrecognized process sheds light on environmental cycling of CH4, suggesting that O2-insensitive, ecologically relevant aerobic CH4 synthesis is likely of widespread distribution in the environment and should be considered in CH4 modeling efforts.Item Arsenate-Induced Changes in Bacterial Metabolite and Lipid Pools during Phosphate Stress(American Society for Microbiology, 2021-02) Zhuang, Weiping; Balasubramanian, Narayanaganesh; Wang, Lu; Wang, Qian; McDermott, Timothy R.; Copie, Valerie; Wang, Gejiao; Bothner, BrianArsenic is widespread in the environment and is one of the most ubiquitous environmental pollutants. Parodoxically, the growth of certain bacteria is enhanced by arsenic when phosphate is limited.Item Metabolic Responses to Arsenite Exposure Regulated through Histidine Kinases PhoR and AioS in Agrobacterium tumefaciens 5A(MDPI, 2020-09) Rawle, Rachel A.; Tokmina-Lukaszewska, Monika; Shi, Zunji; Kang, Yoon-Suk; Tripet, Brian P.; Dang, Fang; Wang, Gejiao; McDermott, Timothy R.; Copie, Valerie; Bothner, BrianArsenite (AsIII) oxidation is a microbially-catalyzed transformation that directly impacts arsenic toxicity, bioaccumulation, and bioavailability in environmental systems. The genes for AsIII oxidation (aio) encode a periplasmic AsIII sensor AioX, transmembrane histidine kinase AioS, and cognate regulatory partner AioR, which control expression of the AsIII oxidase AioBA. The aio genes are under ultimate control of the phosphate stress response via histidine kinase PhoR. To better understand the cell-wide impacts exerted by these key histidine kinases, we employed 1H nuclear magnetic resonance (1H NMR) and liquid chromatography-coupled mass spectrometry (LC-MS) metabolomics to characterize the metabolic profiles of ΔphoR and ΔaioS mutants of Agrobacterium tumefaciens 5A during AsIII oxidation. The data reveals a smaller group of metabolites impacted by the ΔaioS mutation, including hypoxanthine and various maltose derivatives, while a larger impact is observed for the ΔphoR mutation, influencing betaine, glutamate, and different sugars. The metabolomics data were integrated with previously published transcriptomics analyses to detail pathways perturbed during AsIII oxidation and those modulated by PhoR and/or AioS. The results highlight considerable disruptions in central carbon metabolism in the ΔphoR mutant. These data provide a detailed map of the metabolic impacts of AsIII, PhoR, and/or AioS, and inform current paradigms concerning arsenic–microbe interactions and nutrient cycling in contaminated environments.Item A Single Microbiome Gene Alters Murine Susceptibility to Acute Arsenic Exposure(2021-05) Wang, Qian; McDermott, Timothy R.; Walk, Seth T.Environmental toxicant exposure contributes to morbidity and mortality of many human diseases. With respect to arsenic, microbially driven chemical transformations dictate its toxicity and mobility in virtually every environment yet studied, so a general hypothesis is that the human gut microbiome determines disease outcome following exposure. However, the complex nature of the gut microbiome and the myriad of potential interactions with human cells/tissues make it challenging to quantify the influence of specific arsenic-active functions—a requisite step in developing effective disease prevention and/or clinical intervention strategies. To control both mammalian and microbial function during toxicant exposure, we genetically defined the gut microbiome of mice using only Escherichia coli strain, AW3110 (▵arsRBC), or the same strain carrying a single genome copy of the Fucus vesiculosus metallothionein gene (AW3110::fmt); a cysteine-rich peptide that complexes with arsenite, facilitating bioaccumulation and reducing its toxic effects. AW3110::fmt bioaccumulated significantly more arsenic and gnotobiotic mice colonized by this strain excreted significantly more arsenic in stool and accumulated significantly less arsenic in organs. Moreover, AW3110::fmt gnotobiotic mice were protected from acute toxicity exposure (20 ppm AsIII) relative to controls. This study demonstrates—in a highly controlled fashion—that a single microbiome function (arsenic bioaccumulation) encoded by a single gene in a single human gut microbiome bacterium significantly alters mammalian host arsenic exposure. The experimental model described herein allows for a highly controlled and directed assessment of microbiome functions, and is useful to quantify the influence of specific microbiome-arsenic interactions that help mitigate human disease.Item The gut microbiome is required for full protection against acute arsenic toxicity in mouse models(2018-12) Coryell, Michael; McAlpine, Mark; Pinkham, Nicholas V.; McDermott, Timothy R.; Walk, Seth T.Arsenic poisons an estimated 200 million people worldwide through contaminated food and drinking water. Confusingly, the gut microbiome has been suggested to both mitigate and exacerbate arsenic toxicity. Here, we show that the microbiome protects mice from arsenic-induced mortality. Both antibiotic-treated and germ-free mice excrete less arsenic in stool and accumulate more arsenic in organs compared to control mice. Mice lacking the primary arsenic detoxification enzyme (As3mt) are hypersensitive to arsenic after antibiotic treatment or when derived germ-free, compared to wild-type and/or conventional counterparts. Human microbiome (stool) transplants protect germ-free As3mt-KO mice from arsenic-induced mortality, but protection depends on microbiome stability and the presence of specific bacteria, including Faecalibacterium. Our results demonstrate that both a functional As3mt and specific microbiome members are required for protection against acute arsenic toxicity in mouse models. We anticipate that the gut microbiome will become an important explanatory factor of disease (arsenicosis) penetrance in humans, and a novel target for prevention and treatment strategies.Item Phosphate starvation response controls genes required to synthesize the phosphate analog arsenate(2018-05) Wang, Qian; Kang, Yoon-Suk; Alowaifeer, Abdullah; Shi, Kaixiang; Fan, Xia; Wang, Lu; Jetter, Jonathan; Bothner, Brian; Wang, Gejiao; McDermott, Timothy R.Environmental arsenic poisoning affects roughly 200 million people worldwide. The toxicity and mobility of arsenic in the environment is significantly influenced by microbial redox reactions, with arsenite (AsIII ) being more toxic than arsenate (AsV ). Microbial oxidation of AsIII to AsV is known to be regulated by the AioXSR signal transduction system and viewed to function for detoxification or energy generation. Here, we show that AsIII oxidation is ultimately regulated by the phosphate starvation response (PSR), requiring the sensor kinase PhoR for expression of the AsIII oxidase structural genes aioBA. The PhoRB and AioSR signal transduction systems are capable of transphosphorylation cross-talk, closely integrating AsIII oxidation with the PSR. Further, under PSR conditions, AsV significantly extends bacterial growth and accumulates in the lipid fraction to the apparent exclusion of phosphorus. This could spare phosphorus for nucleic acid synthesis or triphosphate metabolism wherein unstable arsenic esters are not tolerated, thereby enhancing cell survival potential. We conclude that AsIII oxidation is logically part of the bacterial PSR, enabling the synthesis of the phosphate analog AsV to replace phosphorus in specific biomolecules or to synthesize other molecules capable of a similar function, although not for total replacement of cellular phosphate.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.Item A periplasmic arsenite-binding protein involved in regulating arsenite oxidation(2011-12) Liu, Guanghui; Liu, Mengyao; Kim, Eun-Hae; Maaty, Walid S.; Bothner, Brian; Lei, Benfang; Rensing, Christopher; Wang, Gejiao; McDermott, Timothy R.Arsenic (As) is the most common toxic element in the environment, ranking first on the Superfund List of Hazardous Substances. Microbial redox transformations are the principal drivers of As chemical speciation, which in turn dictates As mobility and toxicity. Consequently, in order to manage or remediate environmental As, land managers need to understand how and why microorganisms react to As. Studies have demonstrated a two-component signal transduction system comprised of AioS (sensor kinase) and AioR (response regulator) is involved in regulating microbial AsIII oxidation, with the AsIII oxidase structural genes aioB and aioA being upregulated by AsIII. However, it is not known whether AsIII is first detected directly by AioS or by an intermediate. Herein we demonstrate the essential role of a periplasmic AsIII-binding protein encoded by aioX, which is upregulated by AsIII. An ΔaioX mutant is defective for upregulation of the aioBA genes and consequently AsIII oxidation. Purified AioX expressed without its TAT-type signal peptide behaves as a monomer (MW 32 kDa), and Western blots show AioX to be exclusively associated with the cytoplasmic membrane. AioX binds AsIII with a KD of 2.4 µM AsIII; however, mutating a conserved Cys108 to either alanine or serine resulted in lack of AsIII binding, lack of aioBA induction, and correlated with a negative AsIII oxidation phenotype. The discovery and characterization of AioX illustrates a novel AsIII sensing mechanism that appears to be used in a range of bacteria and also provides one of the first examples of a bacterial signal anchor protein.