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Invasive plant species: Inventory, mapping, and monitoring - A national strategy

America is under siege by invasive species of plants and animals, and by diseases. The current environmental, economic, and health-related costs of invasive species could exceed $138 billion per year-more than all other natural disasters combined. Notorious examples include West Nile virus, Dutch elm disease, chestnut blight, and purple loose- strife in the Northeast; kudzu, Brazilian peppertree, water hyacinth, nutria, and fire ants in the Southeast; zebra mussels, leafy spurge, and Asian long-horn beetles in the Midwest; salt cedar, Russian olive, and Africanized bees in the Southwest; yellow star thistle, European wild oats, oak wilt disease, Asian clams, and white pine blister rust in California; cheatgrass, various knapweeds, and thistles in the Great Basin; whirling disease of salmonids in the Northwest; hundreds of invasive species from microbes to mammals in Hawaii; and the brown tree snake in Guam. Thousands of species from other countries are introduced intentionally or accidentally into the United States each year. Based on past experience, 10-15 percent can be expected to establish free-living populations and about 1 percent can be expected to cause significant impacts to ecosystems, native species, economic productivity, and (or) human health.

Information and Technology Report

Assessing the effects of microbial metabolism and metabolities on reservoir pore structure

The effect of microbial treatment on pore structure of sandstone and carbonatereservoirs was determined. Understanding how different bacterial strains and their metabolic bioproducts affect reservoir pore structure will permit the prudent application of microorganisms for enhanced oil recovery. The microbial strains tested included Clostridium acetobutylicum, a polymer-producing Bacillus strain, and an unidentified halophilic anaerobe that mainly produced acids and gases. Electrical conductivity, absolute permeability, porosity and centrifuge capillary pressure were used to examine rock pore structures. Modifications of the pore structure observed in the laboratory cores included pore enlargement due to acid dissolution of carbonates and poare throat reduction due to biomass plugging. This paper shows that careful selection of microbes based on proper understanding of the reservoir petrophysical characteristics is necessary for applications of microbially enhanced oil recovery. These methods and results can be useful to field operators and laboratory researchers involved in design and screening of reservoirs for MEOR. The methods are also applicable in evaluation of formation damage caused by drilling, injection or completion fluids or stimulation caused by acids.

Conference Paper

Ills in the pipeline: Emerging infectious diseases and wildlife

In the recent film Contagion, a medical thriller released in fall 2011, the fictitious MEV-1 virus—passed from bat to pig to humans—spreads across the globe as easily as the common cold, killing millions of humans and causing mass hysteria as medical researchers race to find a cure. Though it's Hollywood hyperbole, the film holds a kernel of truth: Researchers believe that the close proximity of Malaysian hog farms to forested areas—the natural habitat for fruit bats—allowed the previously unknown Nipah virus to spill from bats into pigs and subsequently into people, resulting in more than 100 human deaths (Epstein et al. 2006). There is no doubt that in recent times we have seen an unprecedented number of emerging infectious diseases, defined by the Institute for Medicine as new, reemerging, or drug-resistant infections whose incidence has increased or whose incidence threatens to increase in the near future. Many of these have a wildlife origin (Taylor et al. 2001). While this jump may be due, in part, to increased vigilance and reporting, there is a general consensus that current global conditions are creating a situation that is very favorable to the transmission of microbes that cause diseases. (For reviews, see Daszak et al. 2001 and Keesing et al. 2010). Likewise, it's increasingly important that wildlife professionals become aware of how and why new infectious diseases spread and what, if anything, can be done to minimize impacts on wildlife.

The Wildlife Professional

Far from superficial: microbial diversity associated with the skin and mucus of fish

During horizontal or water-borne infection involving an obligate pathogen (e.g. – Aeromonas salmonicida, cause of furunculosis), the pathogen interacted with and influenced the microbial diversity of the dermal mucus of fish. Prior to infection, the prevalent bacterial flora cultured from juvenile Atlantic salmon (Salmo salar) included Pseudomonas fluorescens, Comomonas terrigenia, Acinetobacter sp., Moraxella sp., Pseudomonas dimunita, Alcaligenes denitrificans, Pseudomonas pseudoalcaligenes, and Pseudomonas alcaligenes, Serratia liquefaciens, Aeromonas hydrophila, other motile Aeromonas spp., and Corynebacterium aquaticum. After A. salmonicida was initially detected in this population as an external mucus infection, Acinetobacter sp., Moraxella sp., C. terrigenia, P. fluorescens, and P. dimunita, Staphylococcus sp., and A. hydrophila, were also present in appreciable numbers. Within several weeks, however, the A. salmonicida infection amplified and composed 78% of the total flora in the mucus. Only P. dimunita (4%). P. fluorescens (2%), and C. terrigenia (1%) were cultured at that time and more than a third of these fish showed evidence of a systemic A. salmonicida infection within their kidneys. Eight weeks after oral oxytetracycline treatments, A. salmonicida was no longer isolated from the mucus or kidneys of any fish and glucose inert or other oxidative microbes (e.g., P. fluorescens, C. terrigenia, Acinetobacter sp., Moraxella sp.) were beginning to repopulate the external surface of the salmon in increasing frequency. Still present and composing fairly large percentages of the total flora were A. hydrophila, as well as Enterobacter sp., and P. putrefaciens. A normal microbial diversity was re-established as the fish recovered. In another investigation, reduced biological diversity was noted in the dermal mucus among smallmouth bass that were sampled from the Jackson River (Covington, VA). In these fish, A. hydrophila and P. putrefaciens were the two predominant microorganisms composing 49.5% and 31.2% of the total bacterial flora, despite the absence of systemic infection or any other clinical signs of disease. In another instance, P. fluorescens was the sole bacterium associated with the surface of Atlantic salmon eggs regardless of their viability at the eyed stage of development. Collectively, these results indicate that the kinetics and distributions of the surface bacterial flora on aquatic organisms is affected by numerous factors including pathogen invasion, environmental conditions, and fish culture practices.

Conference Paper

Sampling from living organisms

Living organisms, unlike inanimate surfaces, seem to exert some control over their surface microbiota, in many cases maintaining conserved, species-specific microbial communities. Microbial ecologists seek to characterize and identify these microbes to understand the roles they are playing in the larger organism's biology.

Book chapter

What's weathering? Mineralogy and field leach studies in mine waste, Leadville and Montezuma mining districts, Colorado

Weathering is important in the development of rock fabrics that control porosity in mine-waste materials, and in turn, porosity affects metal transport through and from mine-waste piles into watersheds. Mine-waste piles are dynamic physical and chemical systems as evidenced by remnant Fe-oxide boxwork structures after sulfide minerals, development of alteration rinds and etch pits on grains, and precipitation of secondary minerals under low temperature conditions. These microscale changes in the mine-waste materials are the result of partial to total dissolution of sulfide and other minerals. Mine-waste materials from the Dinero, Lower Chatauqua, and Saints John sites, Leadville and Montezuma mining districts, Colorado, exhibit rock fabrics that indicate that weathering products, e.g., Fe oxyhydroxides, jarosite, and clays, have been transported in suspension through the waste piles and deposited in voids and as coatings on rock fragments. Microscale characterization of weathered, partially dissolved minerals lends insight into the source of leachable metals in these mine-waste sites. Mineralogic studies show that galena in the Lower Chatauqua waste is enriched in Ag. Qualitative and semiquantitative microanalysis of weathered, altered galena grains from all three sites show that the Ag-bearing galena is more susceptible to dissolution. It is not surprising, then, that solutions experimentally leached from Lower Chatauqua waste are higher in Pb (2310 ppb) compared to leachates from the Dinero (31 ppb) and Saints John (1360 ppb) wastes. The mobility of metals is increased at acidic pH. Using the USGS Field Leach Test protocol, leachate derived from the Dinero waste has a pH of 3 and high concentrations of Al (443 ppb), Fe (441 ppb), and Zn (7970 ppb). Leachate from Sts. John tailings has a pH about 4 and high concentrations of Mn (1520 ppb), Zn (2240 ppb), and Pb (1360 ppb). Leachate from the Lower Chatauqua waste has an intermediate pH of 5, but in addition to the high Pb level already mentioned, it contains high levels of K (1.9 ppm), Mn (6720 ppb), and Zn (1550 ppb). The high concentration of metals, despite the intermediate pH of the leachate, may be explained by acidic microenvironments that exist at the surfaces of sulfide minerals, where sulfur-and iron-oxidizing microbes may flourish. It is at the reactive mineral-oxygen-water interface where metals are released and low-pH sulfate precipitates such as jarosite-beudantite form.

Colorado

Evidence and biogeochemical implications for glacially-derived sediments in an active margin cold seep

Delineating sediment organic matter origins and sediment accumulation rates at gas hydratebearing and hydrocarbon seeps is complicated by the microbial transfer of 13C-depleted and 14Cdepleted methane carbon into sedimentary pools. Sediment 13C and 14C measurements from four cores recovered at Bullseye vent on the northern Cascadia margin are used to identify methane carbon assimilation into different carbon pools. While the total organic carbon (TOC) is mostly unaltered and primarily terrigenous in origin, planktonic foraminifera and the bulk carbonate display evidence of methane overprinting. Mass balance models are applied to determine the extent to which methane overprinting increased the radiocarbon ages of the biogenic foraminifera. The corrected and calibrated foraminifera ages between sediment depths of 70 and 573 cm are from 14.9 to 15.9 ka BP, which coincides with the retreat of the late Quaternary Cordilleran Ice Sheet from Vancouver Island. Uniform TOC _13C values of -24.5 &plusmn; 0.5&permil; from the upper 8 meters of sediment at Bullseye vent suggest all cored material is Pleistocene-derived glacimarine material deposited as the ice edge retreated landward. Bullseye vent is located within an uplifted sediment block isolated from turbidite deposition and has been a site of non-deposition since the ice sheet retreated from the shelf. Biogeochemical implications of seep sediments being dominated by aged, organic-poor (<0.4 wt% TOC) material are that methane is the primary energy source, and microbes directly and indirectly associated with the anaerobic oxidation of methane (AOM) will dominate the seep microbial community.

Conference Paper

Subsurface fate and transport of sulfamethoxazole, 4-nonylphenol, and 17β-estradiol

Subsurface fate and transport of the antibiotic sulfamethoxazole (SX), the non-ionic surfactant degradation product 4-nonylphenol (NP), and the sex hormone 17&beta;-estradiol (E2) were evaluated in a plume of contaminated groundwater at Cape Cod, Massachusetts, USA. The plume is the result of 60 years of wastewater treatment plant effluent disposal into rapid infiltration beds. Natural-gradient, in situ tracer experiments were used to evaluate subsurface transport of SX, NP, and E2 (injected at 300, 530, and 0.55 &micro;g/L, respectively) relative to the conservative tracer bromide. Two geochemical zones were evaluated: (1) uncontaminated groundwater overlying the plume, and (2) contaminated groundwater within the plume that has recently become oxic after decades of anoxic conditions. The uncontaminated groundwater is characterized by a microbial community unacclimated to treated wastewater, whereas the contaminated groundwater is characterized by microbes acclimated to wastewater contaminants. Results from the tracer tests in both zones showed that the antibiotic SX was co-transported with the conservative tracer bromide, with little retardation or mass removal. In contrast, NP and E2, which are more hydrophobic and biodegradable, showed sorption (relative retardation factors ranged up to 5.9) and mass loss at both the uncontaminated and contaminated sites.

Conference Paper

Fate(s) of injected CO2 in a coal-bearing formation, Louisiana, Gulf Coast Basin: Chemical and isotopic tracers of microbial-brine-rock-CO2 interactions

Coal beds are one of the most promising reservoirs for geologic carbon dioxide (CO₂) sequestration, as CO₂ can strongly adsorb onto organic matter and displace methane; however, little is known about the long-term fate of CO₂ sequestered in coal beds. The "2800' sand" of the Olla oil field is a coal-bearing, oil and gas-producing reservoir of the Paleocene–Eocene Wilcox Group in north-central Louisiana. In the 1980s, this field, specifically the 2800' sand, was flooded with CO₂ in an enhanced oil recovery (EOR) project, with 9.0×10⁷m³ of CO₂ remaining in the 2800' sand after injection ceased. This study utilized isotopic and geochemical tracers from co-produced natural gas, oil and brine from reservoirs located stratigraphically above, below and within the 2800' sand to determine the fate of the remaining EOR-CO₂, examining the possibilities of CO₂ migration, dissolution, mineral trapping, gas-phase trapping, and sorption to coal beds, while also testing a previous hypothesis that EOR-CO₂ may have been converted by microbes (CO₂-reducing methanogens) into methane, creating a microbial "hotspot". Reservoirs stratigraphically-comparable to the 2800' sand, but located in adjacent oil fields across a 90-km transect were sampled to investigate regional trends in gas composition, brine chemistry and microbial activity. The source field for the EOR-CO₂, the Black Lake Field, was also sampled to establish the δ¹³C-CO₂ value of the injected gas (0.9‰ +/- 0.9‰). Four samples collected from the Olla 2800' sand produced CO₂-rich gas with δ¹³C-CO₂ values (average 9.9‰) much lower than average (pre-injection) conditions (+15.9‰, average of sands located stratigraphically below the 2800' sand in the Olla Field) and at much higher CO₂ concentrations (24.9 mole %) than average (7.6 mole %, average of sands located stratigraphically below the 2800' sand in the Olla Field), suggesting the presence of EOR-CO₂ and gas-phase trapping as a major storage mechanism. Using δ¹³C values of CO₂ and dissolved organic carbon (DIC), CO₂ dissolution was also shown to be a major storage mechanism for 3 of the 4 samples from the Olla 2800' sand. Minor storage mechanisms were shown to be migration, which only affected 2 samples (from 1 well), and some EOR-CO₂ conversion to microbial methane for 3 of the 4 Olla 2800' sand samples. Since methanogenesis was not shown to be a major storage mechanism for the EOR-CO₂ in the Olla Field (CO₂ injection did not stimulate methanogenesis), samples were examined from adjacent oil fields to determine the cause of the Olla microbial "hot-spot". Microbial methane was found in all oil fields sampled, but indicators of methanogenesis (e.g. alkalinity, high δ¹³C-DIC values) were the greatest in the Olla Field, and the environmental conditions (salinity, pH, temperature) were most ideal for microbial CO₂ reduction in the Olla field, compared to adjacent fields.

Thesis

Effect of N fertilization and tillage on nitrous oxide (N2O) loss from soil under wheat production

Nitrous oxide (N2O-N) is one of the most important gases in the atmosphere because it is 300 times more powerful than carbon dioxide in its ability to trap heat, and is a key chemical agent of ozone depletion. The amount of N2O-N emitted from agricultural fields can be quite high, depending on the complex interplay between N fertility and residue management, plant N uptake, microbial processes, environmental conditions, and wet-up and dry-down events. High N fertilizer rates generally increase yields, but may disproportionately increase N2O-N losses due to prolonged residence time in soil when not used by the crop, and incomplete decomposition of excess N-compounds by microbes. Tillage could also affect N2O-N losses through changes in soil moisture content. Though nitrogen monoxide (NO) is one form of N lost from the soil, especially under conventional tillage, this study objective was to quantify N2O loss in wheat fields from applied urea on soil under no-till (NT) versus incorporated urea under conventional till (CT).

Report

Ground water to surface water: Chemistry of thermal outflows in Yellowstone National Park

Geothermal waters in the earth’s subsurface boil with steam separation and may mix with dilute ground waters (that may or may not contain sulfuric acid from sulfur oxidation), resulting in a wide range of compositions when they discharge and emerge at the surface. As they discharge onto the ground surface they undergo evaporative cooling, degassing, oxidation, and mineral precipitation. Within this aquatic environment of rapidly changing physical and chemical parameters, numerous microbial communities develop—some of which affect oxidation and mineral precipitation. Microbes are responsible for rapid oxidation of iron and arsenic in thermal outflows, and for catalyzing the production of sulfuric acid from the oxidation of elemental sulfur. The attractive visual display of colors observed in Yellowstone’s geothermal waters reflects this interplay of physical, chemical, and biological phenomena. Oxidation of dissolved sulfide to thiosulfate occurs abiotically, and thiosulfate can be found in many of Yellowstone’s thermal waters—at any pH, temperature, and composition. Polythionates, on the other hand, are rarely found in Yellowstone waters but are associated with sulfur hydrolysis in Cinder Pool. Oxidation rates of iron and arsenic in overflows have been estimated at 1-3 mM/h and 0.04-0.1 mM/h, respectively—orders of magnitude faster than the abiotic rate. The abiotic production of thiosulfate from oxidation of dissolved sulfi de at Angel Terrace and Ojo Caliente is about 3-30 µM/min, faster by 2-3 orders of magnitude than the laboratory rate at 25°C. The partitioning of dissolved sulfide between that volatilized to the air and that oxidized to thiosulfate has been estimated at Angel Terrace and at Ojo Caliente. For the pH range of 6-8 and the temperature range of 50-93°C, 67-86% of the dissolved sulfide is lost to the atmosphere and 10-33% is oxidized to thiosulfate. Only a very small percentage, if any, forms elemental sulfur under these conditions.

Wyoming

Application of cross-borehole radar to monitor fieldscale vegetable old injection experiments for biostimulation

Cross-borehole radar methods were used to monitor a field-scale biostimulation pilot project at the Anoka County Riverfront Park (ACP), located downgradient of the Naval Industrial Reserve Ordnance Plant, in Fridley, Minnesota. The goal of the pilot project is to evaluate biostimulation using emulsified vegetable oil to treat ground water contaminated with chlorinated hydrocarbons. Vegetable oil is intended to serve as substrate to naturally occurring microbes, which ultimately break down chlorinated hydrocarbons into chloride, carbon dioxide, and water through oxidation-reduction reactions. In support of this effort, cross-borehole radar data were acquired by the U.S Geological Survey in five site visits over 1.5 years. This paper presents level-run (zero-offset profile) and time-lapse radar tomography data collected in multiple planes. Comparison of pre- and post-injection data sets provides valuable insights into the spatial and temporal distribution of both emulsified vegetable oil and also the extent of ground water with altered chemistry resulting from injections — information important for understanding microbial degradation of chlorinated hydrocarbons at the site. In order to facilitate data interpretation and test the effectiveness of radar for monitoring oil-emulsion placement and movement, three injection mixtures with different radar signatures were used: (1) vegetable oil emulsion, (2) vegetable oil emulsion with a colloidal iron tracer, and (3) vegetable oil emulsion with a magnetite tracer. Based on petrophysical modeling, mixture (1) is expected to increase radar velocity and decrease radar attenuation relative to background — a water-saturated porous medium; mixtures (2) and (3) are expected to increase radar velocity and also increase radar attenuation due to their greater electrical conductivity compared to native ground water. Radar slowness (inverse radar velocity) tomograms and level-run profiles show decreases in slowness in the vicinity of injection wells. Slowness anomalies are observed only in planes connected to injection wells, indicating that the emplaced emulsified vegetable oil does not migrate far after injection. In contrast to the localization of slowness anomalies, attenuation anomalies are observed in all level-run profiles, particularly those downgradient of the injection wells. Despite the expected signatures of different tracers, increases in attenuation are observed downgradient of all three injections; thus, we infer that the attenuation changes do not result from the iron tracers. One viable explanation for the observed attenuation changes is that products of oil-enhanced biodegradation (for example, ferrous iron) increase electrical conductivity of ground water and thus radar attenuation. Application of radar methods to data from the ACP demonstrated the utility of radar for monitoring biostimulation. Results of level-run and tomographic surveys identified (1) the distribution of emulsified vegetable oil, and (2) the distribution of ground water with oil-affected chemistry. Ongoing research efforts include simultaneous tomographic inversion of radar data from multiple planes, petrophysical modeling, geostatistical interpolation, and development of an integrated interpretation considering conventional borehole logs and surface-to-borehole radar data.

Minnesota

People need freshwater biodiversity: Nine reasons freshwater biodiversity is important for humans

Freshwater biodiversity, from fish to frogs and microbes to macrophytes, provides a vast array of services to people. These services are under a growing threat as human-related pressure threatens freshwater biodiversity on multiple fronts. Here, Abigail Lynch and colleagues discuss nine fundamental ecosystem services that freshwater biodiversity provide to people. Given the importance of freshwater biodiversity to our very existence, they make a plea for a redoubled effort to better conserve this precious resource.

Global Water Forum

Detection of spring viraemia of carp virus in imported amphibians reveals an unanticipated foreign animal disease threat

Global translocation of plants and animals is a well-recognized mechanism for introduction of pathogens into new regions. To mitigate this risk, various tools such as preshipment health certificates, quarantines, screening for specific disease agents and outright bans have been implemented. However, such measures only target known infectious agents and their hosts and may fail to prevent translocation of even well-recognized pathogens if they are carried by novel host species. In a recent example, we screened an imported shipment of Chinese firebelly newts ( Cynops orientalis ) for Batrachochytrium salamandrivorans , an emergent fungal pathogen of salamanders. All animals tested negative for the fungus. However, a virus was cultured from internal organs from 7 of the 11 individual dead salamanders and from two pools of tissues from four additional dead animals. Sequencing of a portion of the glycoprotein gene from all viral isolates indicated 100 % identity and that they were most closely related to spring viraemia of carp virus (SVCV). Subsequently, SVCV-specific PCR testing indicated the presence of virus in internal organs from each of the four animals previously pooled, and whole-genome sequencing of one of the viral isolates confirmed genomic arrangement characteristic of SVCV. SVCV is a rhabdovirus pathogen of cyprinid fish that is listed as notifiable to the Office International des Epizooties. This discovery reveals a novel route for potential spillover of this economically important pathogen as rhabdovirus has not previously been documented in amphibians.

Emerging Microbes & Infections

Surveillance for highly pathogenic influenza A viruses in California during 2014–2015 provides insights into viral evolutionary pathways and the spatiotemporal extent of viruses in the Pacific Americas Flyway

We used surveillance data collected in California before, concurrent with, and subsequent to an outbreak of highly pathogenic (HP) clade 2.3.4.4 influenza A viruses (IAVs) in 2014–2015 to (i) evaluate IAV prevalence in waterfowl, (ii) assess the evidence for spill-over infections in marine mammals and (iii) genetically characterize low-pathogenic (LP) and HP IAVs to refine inference on the spatiotemporal extent of HP genome constellations and to evaluate possible evolutionary pathways. We screened samples from 1496 waterfowl and 1142 marine mammals collected from April 2014 to August 2015 and detected IAV RNA in 159 samples collected from birds ( n = 157) and pinnipeds ( n = 2). HP IAV RNA was identified in three samples originating from American wigeon ( Anas americana ). Genetic sequence data were generated for a clade 2.3.4.4 HP IAV-positive diagnostic sample and 57 LP IAV isolates. Phylogenetic analyses revealed that the HP IAV was a reassortant H5N8 virus with gene segments closely related to LP IAVs detected in mallards ( Anas platyrhynchos ) sampled in California and other IAVs detected in wild birds sampled within the Pacific Americas Flyway. In addition, our analysis provided support for common ancestry between LP IAVs recovered from waterfowl sampled in California and gene segments of reassortant HP H5N1 IAVs detected in British Columbia, Canada and Washington, USA. Our investigation provides evidence that waterfowl are likely to have played a role in the evolution of reassortant HP IAVs in the Pacific Americas Flyway during 2014–2015, whereas we did not find support for spill-over infections in potential pinniped hosts.

Emerging Microbes & Infections

Genomic characterization of highly pathogenic H5 avian influenza viruses from Alaska during 2022 provides evidence for genotype-specific trends of spatiotemporal and interspecies dissemination

The ongoing panzootic of highly pathogenic H5 clade 2.3.4.4b avian influenza (HPAI) spread to North America in late 2021, with detections of HPAI viruses in Alaska beginning in April 2022. HPAI viruses have since spread across the state, affecting many species of wild birds as well as domestic poultry and wild mammals. To better understand the dissemination of HPAI viruses spatiotemporally and among hosts in Alaska and adjacent regions, we compared the genomes of 177 confirmed HPAI viruses detected in Alaska during April – December 2022. Results suggest multiple viral introductions into Alaska between November 2021 and August or September 2022, as well as dissemination to areas within and outside of the state. Viral genotypes differed in their spatiotemporal spread, likely influenced by timing of introductions relative to population immunity. We found evidence for dissemination of HPAI viruses between wild bird species, wild birds and domestic poultry, as well as wild birds and wild mammals. Continued monitoring for and genomic characterization of HPAI viruses in Alaska can improve our understanding of the evolution and dispersal of these economically costly and ecologically relevant pathogens.

Emerging Microbes & Infections

Conversion of the pathogenic fungus Colletotrichum magna to a nonpathogenic, endophytic mutualist by gene disruption

Hygromycin-resistant transformants of the cucurbit pathogen Colletotrichum magna (teleomorph: Glomerella magna) were generated by restriction enzyme-mediated integration (REMI) transformation. A rapid pathogenicity assay involving watermelon (Citrullus lanatus) seedlings was developed and 14,400 REMI transformants were screened and assessed for their ability to cause disease, colonize plant tissues, and confer disease resistance against wild-type C. magna. A total of 176 nonpathogenic REMI mutants capable of colonizing cucurbit plants were isolated and assigned to three groups based on their ability to confer disease resistance: phenotype A, 80 to 100% disease protection; phenotype B, 10 to 65% disease protection; and phenotype C, 0 to 4% disease protection. Molecular and genetic analyses of one REMI mutant (R1) indicated that the nonpathogenic phenotype A resulted from a single-site integration. R1 showed a 1:1 segregation of hygromycin resistance and nonpathogenicity and all hygromycin-resistant progeny were nonpathogenic. The integrated vector and 5.5 kb of flanking fungal genomic DNA were isolated from R1 and designated pGMR1. To verify that pGMR1 contained pathogenicity gene sequences, a wild-type isolate of C. magna was transformed with pGMR1 to induce gene disruptions by homologous integration. Approximately 47% of the pGMR1 transformants expressed phenotype A, indicating homologous integration and gene disruption.

Molecular Plant-Microbe Interactions

Fungal biology and agriculture: revisiting the field

Plant pathology has made significant progress over the years, a process that involved overcoming a variety of conceptual and technological hurdles. Descriptive mycology and the advent of chemical plant-disease management have been followed by biochemical and physiological studies of fungi and their hosts. The later establishment of biochemical genetics along with the introduction of DNA-mediated transformation have set the stage for dissection of gene function and advances in our understanding of fungal cell biology and plant-fungus interactions. Currently, with the advent of high-throughput technologies, we have the capacity to acquire vast data sets that have direct relevance to the numerous subdisciplines within fungal biology and pathology. These data provide unique opportunities for basic research and for engineering solutions to important agricultural problems. However, we also are faced with the challenge of data organization and mining to analyze the relationships between fungal and plant genomes and to elucidate the physiological function of pertinent DNA sequences. We present our perspective of fungal biology and agriculture, including administrative and political challenges to plant protection research.

Molecular Plant-Microbe Interactions