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Efficacy Improvement of a Bioherbicidal Fungus Using a Formulation-Based Approach

American Journal of Plant Sciences 2016年第16期7卷 2349-2358页

作者： Clyde D. Boyette Robert E. Hoagland kenneth C. stetina USDA-ARS Biological Control of Pests Research Unit Stoneville MS USA USDA-ARS Crop Production Systems Research Unit Stoneville MS USA

Greenhouse experiments were conducted to determine the effects of an invert (water-in-oil) emulsion (IE) on dew period duration and dew delay of Colletotrichum coccodes for biocontrol of the problematic weed, eastern black nightshade (Solanum ptycanthum). Dew periods of 4, 8, or 12 h provided 10%, 25%, and 40% control of eastern black nightshade plants, respectively, when C. coccodes (Strain NRRL 15,547) spores were applied in water + Tween 80 surfactant 12 days after inoculation, but a minimum of 16 h of dew was required to achieve ~95% plant mortality. In contrast, at these same intervals of dew, 95%, 100% and 100% mortality occurred, respectively, when fungal spores were formulated in the IE. Even in the absence of dew, 60% mortality and 70% dry weight reductions of plants was achieved with the fungus/IE formulation Delaying dew by 2 h after inoculation did not significantly reduce weed control or plant dry weight reductions when plants were inoculated with the fungus either in the aqueous or in the IE formulation. However, when dew was delayed for 4, 8, or 12 h, only 60%, 50%, and 25% mortality, respectively, of plants receiving the aqueous spore treatment occurred. In contrast, 95%, 90%, and 90% weed mortality occurred after the same dew delays of plants receiving the fungus/ IE treatments. These results demonstrate that formulating C. coccodes spores in an invert emulsion greatly improves the bioherbicidal potential of this fungus. Furthermore, results suggest that this formulation may render pathogens previously rejected for development as bioherbicides due to restrictive dew requirements more efficacious for use in controlling their target weeds.

关键词： Colletotrichum coccodes Solanum ptycanthum Bioherbicide Invert Emulsion Formulation

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Biological Control Potential of Colletotrichum gloeosporioidesfor Coffee Senna (Cassia occidentalis)

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American Journal of Plant Sciences 2012年第4期3卷 430-436页

作者： Clyde D. Boyette Robert E. Hoagland Mark A. Weaver kenneth stetina United States Department of Agriculture-Agricultural Research Service (USDA-ARS) Biological Control of Pests Research Unit United States Department of Agriculture-Agricultural Research Service (USDA-ARS) Biological Control of Pests Research Unit Stoneville USA USDA-ARS Crop Production Systems Research Unit Stoneville USA

A fungal pathogen, Colletotrichum gloeosporioides was isolated from a greenhouse-grown seedling of coffee senna (Cassia occidentalis) and evaluated as a mycoherbicide for that weed. Host range tests revealed that coffee senna, wild senna (C. marilandica), and sicklepod (C. obtusifolia) were also affected by this pathogen, but 35 other crop and weed species, representing 8 botanical families were not affected. The fungus sporulated prolifically on solid and liquid media with maximum spore germination and growth occurring at 20°C - 30°C. Optimal environmental conditions included at least 12 h of free moisture (dew) at 20°C - 30°C. Spray mixtures containing approximately 1.0 × 105 or more conidia·ml–1 gave maximum control when coffee senna seedlings were sprayed until runoff occurred. Coffee senna seedlings that were in the cotyledon to first-leaf growth stage were most susceptible to this pathogen. Weed control efficacy studies under field conditions demonstrated that control of coffee senna was directly proportional to the inoculum concentration applied. Results of these tests suggest that this fungus has potential as a mycoherbicide to control coffee senna, a serious weed in the southeastern U.S.

关键词： Bioherbicide Mycoherbicide Coffee Senna Cassia occidentalis Colletotrichum gloeosporioides

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Hot Water Treatment Enhances the Bioherbicidal Efficacy of a Fungus

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American Journal of Plant Sciences 2018年第10期9卷 2063-2076页

作者： C. Douglas Boyette Robert E. Hoagland kenneth C. stetina USDA-ARS Biological Control of Pests Research Unit Stoneville MS USA USDA-ARS Crop Production and Systems Research Unit Stoneville MS USA

Hemp sesbania (Sesbania exaltata) plants (>30 cm tall) sprayed with hot water (45°C – 95°C), followed by spray applications of fungal spores of Colletotrichum truncatum (CT) at 1.0 × 107 spores/ml-1 and 22°C – 25°C, suspended either in: 0.2% Silwet L-77 surfactant (SW);unrefined corn oil (CO)/distilled water (1:1, v:v);or 0.2% SW in CO were controlled by 80% - 95%, 12 days after treatment (DAT) under greenhouse conditions. These treatments also reduced dry weight accumulation of this weed. Plants treated with hot water without CT were also injured at temperatures ≥35°C (5% mortality), and 60% mortality at 95°C. Artificial dew treatments (25°C, 12 h), imposed on plants after the treatment protocols above, had little or no effect on weed mortality or dry weight reduction compared to treated plants without dew. Under field conditions, 85% control of hemp sesbania was achieved 12 - 15 DAT when a pre-treatment with hot water (65°C) was followed immediately with a CT application at the spore concentration as described above. Plants in field tests treated with CT without a hot water treatment were visually unaffected, with no mortality or plant biomass reductions recorded 15 DAT. These results suggest that use of hot water may be an important tool for improving the infectivity and bioherbicidal potential of some plant pathogens.

关键词： Bioherbicide Mycoherbicide Hemp Sesbania Sesbania exaltata Colletotrichum truncatum Hot Water Pre-Treatment Improved Bioherbicide Efficacy Weed Control

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Interaction of the Bioherbicide Myrothecium verrucaria and Glyphosate for Kudzu Control

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American Journal of Plant Sciences 2014年第26期5卷 3943-3956页

作者： Clyde Douglas Boyette Robert E. Hoagland Mark A. Weaver kenneth C. stetina United States Department of Agriculture-Agricultural Research Service (USDA-ARS) Biological Control of Pests Research Unit Stoneville MS USA United States Department of Agriculture-Agricultural Research Service (USDA-ARS) Crop Production Systems Research Unit Stoneville MS USA

Kudzu is an exotic invasive weed in the southeastern U.S. that is difficult to control with current commercial herbicides. Some success for its control has been achieved using a bioherbicidal agent, Myrothecium verrucaria (MV). Spore and mycelial formulations of MV were tested alone and in combination with glyphosate for control of kudzu (Pueraria lobata) under greenhouse and field conditions in naturally-infested areas. In greenhouse and field experiments, kudzu control increased as the concentration of spores or mycelia increased. Glyphosate alone provided 10%, 35%, 50% and 60% control in field experiments at 0.25, 0.50, 0.75 and 1.0X rates, respectively and MV alone spores provided 15%, 50%, 65% and 85% control at 0.25, 0.50, 0.75 and 1.0X rates, respectively. However, when MV spores were combined with glyphosate, significantly higher control occurred than that caused by either component alone. Similar levels of control were observed for MV mycelial formulations applied alone or with glyphosate at equivalent concentrations of the fungus. The rate of disease progression was more rapid and severe at all fungal spore or mycelial formulations and herbicide rates when these propagules were applied in combination with glyphosate. In field tests, 24 h after application, only 20% of kudzu plants were severely damaged by MV alone (0.25X), whereas 80% were severely diseased when MV spores and glyphosate were mixed and applied at 0.25X rates each. A similar trend occurred with the MV mycelial formulation applied at these rates. Synergist interactions on kudzu control were observed, especially when lower levels of MV (spores or mycelia) and glyphosate were combined and applied to kudzu in the greenhouse or in the field. These results suggest that it may be possible to incorporate glyphosate to improve the bioherbicidal control potential and reduce herbicide and inoculum requirements of M. verrucaria spores or mycelium for controlling kudzu.

关键词： Bioherbicide Glyphosate Kudzu Myrothecium verrucaria Pueraria lobata

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Effects of Myrothecium verrucariaon Two Glyphosate-Resistant Amaranthus palmeriBiotypes Differing in Betacyanin Content

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American Journal of Plant Sciences 2020年第2期11卷 214-225页

作者： Robert E. Hoagland Clyde Douglas Boyette Robin H. Jordan kenneth C. stetina Crop Production Systems Research Unit USDA-ARS Stoneville MS USA Biological Control of Pests Research Unit USDA-ARS Stoneville MS USA

Previously we found two biotypes of Amaranthus palmeri (Palmer amaranth) in a population of this economically important weed that were resistant to glyphosate but differed with respect to pigmentation. One biotype was typically red-pigmented (betacyanin) while the other was green, with no visual appearance of red hue on any plant part at any growth stage. We have also reported that a strain of Myrothecium verrucaria (MV) exhibited bioherbicidal activity against several important weeds including glyphosate-resistant Palmer amaranth. In greenhouse tests, MV was applied to these two biotypes (red and green) at two ages (3-week- and 6-week-old) and effects of this fungus monitored over a 5-day time course. Initial symptoms of MV (16 to 24 h after inoculation) were: epinastic curvature, wilting and development of lesions on leaves and stems. Generally, the younger plants tended to be more sensitive to MV than older plants. Bioherbicidal damage increased with time leading to necrosis and plant mortality and increasing disease progress. Severe loss of fresh weight occurred in both biotypes as compared to untreated plants. Results indicated that MV was effective on both biotypes, but effects on growth reduction and disease progression were more rapid and generally greater in the green biotype, suggesting that compounds responsible for red pigmentation may be more potent as defense against pathogen attack.

关键词： Betalain Bioherbicide Biological Weed Control Palmer Amaranth Pigweed

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Interaction of Glufosinate and Colletotrichum truncatumon Ammonia Levels and Glutamine Synthetase Activity in Hemp Sesbania

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American Journal of Plant Sciences 2018年第11期9卷 2320-2337页

作者： Robert E. Hoagland C. Douglas Boyette Robin H. Jordan kenneth C. stetina Crop Production Systems Research Unit USDA-ARS Stoneville MS USA Biological Control of Pests Research Unit USDA-ARS Stoneville MS USA

The use of microbes and microbial products as bioherbicides has been studied for several decades, and combinations of bioherbicides and herbicides have been examined to discover possible synergistic interactions to improve weed control efficacy. Bioassays were conducted to assess possible interactions of the herbicide glufosinate [2-amino-4-(hydroxymethylphosphinyl) butanoic acid] and Colletotrichum truncatum (CT), a fungal bioherbicide to control hemp sesbania (Sesbania exaltata)]. Glufosinate acts as a glutamine synthetase (GS) inhibitor that causes elevated ammonia levels, but the mode of action of CT is unknown. GS has also been implicated in plant defense in certain plant-pathogen interactions. The effects of spray applications of glufosinate (1.0 mM) orbioherbicide (8.0 × 104 conidia ml-1), applied alone or in combination were monitored (88 h time-course) on seedling growth, GS activity and ammonia levels in hypocotyl tissues under controlled environmental conditions. Growth (elongation and fresh weight) and extractable GS activity were inhibited in tissues by glufosinate and glufosinate plus CT treatments as early as 16 h, but CT treatment did not cause substantial growth reduction or GS inhibition until after ~40 h. Generally, ammonia levels in hemp sesbania tissues under these various treatments were inversely correlated with GS activity. Localization of hemp sesbania GS activity on electrophoretic gels indicated a lack of activity after 30 h in glufosinate and glufosinate plus CT-treated tissue. Untreated control tissues contained much lower ammonia levels at 24, 64, and 88 h after treatment than treatments with CT, glufosinate or their combination. CT alone caused elevated ammonia levels only after 64 - 88 h. Glufosinate incorporated in agar at 0.25 mM to 2.0 mM, caused a 10% - 45% reduction of CT colony radial growth, compared to fungal growth on agar without glufosinate, and the herbicide also inhibited sporulation of CT. Although no synergistic interac

关键词： Bioherbicide Hemp Sesbania Glutamine Synthetase Ammonia Glufosinate Colletotrichum truncatum

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Bioherbicidal Efficacy of a Myrothecium verrucaria-Sector on Several Plant Species

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American Journal of Plant Sciences 2016年第16期7卷 2376-2389页

作者： Robert E. Hoagland Clyde D. Boyette kenneth C. stetina Robin H. Jordan United States Department of Agriculture-Agricultural Research Service (USDA-ARS) Crop Production Systems Research Unit Stoneville MS USA United States Department of Agriculture-Agricultural Research Service (USDA-ARS) Biological Control of Pests Research Unit Stoneville MS USA

Comparative studies were conducted on mycelial preparations of the bioherbicide, Myrothecium verrucaria (MV) strain IMI 361690 and a recently discovered sector (MV-Sector BSH) of this fungus. The whitish sector was discovered, isolated, grown in pure culture on PDA and found to be a stable, non-spore producing mutant when cultured over several months under conditions that cause circadian sporulation during growth of its MV parent. Application of MV and MV-Sector BSH mycelial preparations to intact plants (hemp sesbania and sicklepod) and leaf discs (kudzu and glyphosate-resistant Palmer amaranth) showed that the sector efficacy was generally equal to, or slightly lower than MV. Bioassays of MV and this sector on seed germination and early growth of sicklepod and hemp sesbania seeds demonstrated that hemp sesbania seeds were slightly more sensitive to the fungus than sicklepod seeds and that the sector bioherbicidal activity was slightly less than that of MV. SDS-PAGE protein profiles of cellular extracts of MV and the sector and their respective culture supernatants showed several differences with respect to quantity and number of certain protein bands. Overall results showed that the isolate was a non-spore producing mutant with phytotoxicity to several weeds (including weeds tolerant or resistant to glyphosate), and that the phytotoxic effects were generally equivalent to those caused by MV treatment. Results of this first report of a non-sporulating MV mutant that suggest additional studies on protein analysis, and an extended weed host range under greenhouse and field conditions are needed in order to further evaluate its possible bioherbicidal potential.

关键词： Bioherbicide Glyphosate-Resistance Palmer Amaranth Kudzu Hemp Sesbania Sicklepod Myrothecium verrucaria Fungal Sector

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Interaction of the Bioherbicide Myrothecium verrucariawith Technical-Grade Glyphosate on Glyphosate-Susceptible and -Resistant Palmer Amaranth

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American Journal of Plant Sciences 2018年第11期9卷 2306-2319页

Previously we found that a strain of Myrothecium verrucaria (MV) exhibited bioherbicidal activity against several important weeds, and that some commercial formulations of glyphosate applied with MV resulted in synergistic interactions that improved weed control efficacy. We also found that MV had bioherbicidal activity against glyphosate-resistant Palmer amaranth. We have also reported that some commercial formulations are inhibitory to MV. Our objectives were to test the effect of unformulated glyphosate (high purity, technical-grade glyphosate) alone and in combination with MV for bioherbicidal activity on glyphosate-susceptible and -resistant Palmer amaranth biotypes under greenhouse conditions and to examine technical-grade glyphosate on the growth of this bioherbicide. High purity glyphosate (without adjuvants/surfactants) was not toxic to MV growth and sporulation at concentrations up to 2.0 mM when grown on agar supplemented with the herbicide. Both biotypes were injured by MV and MV plus glyphosate treatments as early as 19 h after application (3 h after a dew period of 16 h). These injury effects increased and were more evident through the 6-day time course, when after 120 h the MV plus glyphosate treatment had killed all glyphosate-susceptible and -resistant plants. The interaction of glyphosate plus MV was synergistic toward the control of Palmer amaranth. Data strongly suggest that the active ingredient is responsible for the synergy previously found when this bioherbicide was combined with some commercial formulations of glyphosate. Results demonstrated that MV can control both glyphosate-resistant and -susceptible Palmer amaranth seedlings and act synergistically with high-purity glyphosate to provide improved weed control.

关键词： Amaranthus palmeri Biocontrol Agent Bioherbicide Biological Weed Control Glyphosate-Resistance Myrothecium verrucaria Palmer Amaranth Pigweed

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Interaction of a Myrothecium verrucariaMycelial Preparation and a Glyphosate Product for Controlling Redvine (Brunnichia ovata) and Trumpet Creeper (Campsis radicans)

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American Journal of Plant Sciences 2020年第2期11卷 201-213页

作者： Clyde Douglas Boyette Robert E.Hoagland Charles T.Bryson Mark A.Weaver kenneth C.stetina USDA-ARS Biological Control of Pests Research UnitStonevilleMSUSA USDA-ARS Crop Production and Systems Research UnitStonevilleMSUSA

A mycelial formulation of the bioherbicidal fungus Myrothecium verrucaria (Alb. & Schwein.) Ditmar: Fr. (MV) was tested alone and in combination with a commercially available glyphosate [N-(phosphonomethyl)glycine] (GLY) product for controlling the invasive vines, redvine [Brunnichia ovata (Walt.) Shinners], and trumpet creeper [Campsis radicans (L.) Seem. ex Bureau] in field experiments conducted near Stoneville, MS. Several application timing regimens were evaluated (Fall, Spring, Fall + Spring, and Spring + Fall). We found that a Fall + Spring application of MV + GLY controlled redvine and trumpet creeper by 95%, 12 days after the second treatment, through a synergistic interaction of the fungus and glyphosate. Disease symptomatology was characterized by rapid necrosis of leaf and stem tissues, with mortality occurring within 72 h. Neither glyphosate alone, nor MV alone, effectively controlled either weed species under any application timing regimen. No visual disease or herbicide damage occurred on glyphosate-resistant soybean plants in the treated test plots. These results suggest that some formulations of glyphosate, mixed with the bioherbicide MV, can effectively control redvine and trumpet creeper, two of the most troublesome weeds in the row crops of the Mississippi Delta region in the mid-southern U.S.

关键词： Bioherbicide Interaction Mycelial Formulation Myrothecium verrucaria Campsis radicans Brunnichia ovata

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Host Range and Virulence of a Fungal Pathogen for Control of Giant Salvinia (Salvinia molesta)

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American Journal of Plant Sciences 2021年第3期12卷 444-454页

作者： Clyde D. Boyette Robert E. Hoagland Lawrence R. Higgenbotham H. Lynn Walker James A. Young kenneth C. stetina USDA-ARS Biological Control of Pests Research Unit Stoneville MS USA Crop Production Systems Research Unit Stoneville MS USA Louisiana Tech University Ruston LA USA

A teleomorph of the fungus Botryosphaeria rhodina (Berkeley et Curtis) von Arx, (Br) was evaluated as a bioherbicide for control of giant salvinia (Salvinia molesta D.S. Mitchell) under greenhouse conditions and in small-scale field trials. We found that fungal mycelium was highly infective and could be rapidly produced (48+ h) in soy flour-cornmeal liquid media contained in shake flasks or fermenters. A dew period was not required to achieve infection and mortality of inoculated plants. A surfactant (Silwet L-77, a polyalkyleneoxide modified heptamethyl-trisiloxane) incorporated in the fungal formulation was required for Br to infect and kill plants. Infection and mortality occurred rapidly (within 48 h after treatment), and re-growth of treated plants did not occur. In replicated field trials, Br controlled giant salvinia ~95%. Br also infected other plants,

关键词： Biological Control Botryosphaeria rhodina (Berkeley et Curtis) Von Arx Fungal Phytopathogen Salvinia molesta D.S. Mitchell Aquatic Weed

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