Interactions and Effects on Cysteine Synthase Activity of Aminooxyacetate and Boc-Aminooxyacetate on the Bioherbicides <i>Colletotrichum truncatum</i>and <i>Alternaria cassia</i>and Their Weed Hosts

Aminooxyacetate (AOA) is a pyridoxal phosphate antagonist that inhibits various plant enzymes (including transaminases) which require pyridoxal phosphate as a cofactor and it exhibits phytotoxic and herbicidal properties. We examined AOA and its analog, N-t-butoxycarbonyl-AOA (Boc-AOA) for phytotoxicity, interactions with weed pathogens (bioherbicides), and effects on an important pyridoxal requiring enzyme, cysteine synthase (CS, E.C. 4.2.99.8). Studies were performed on two weeds, i.e., hemp sesbania [Sesbania exaltata (Raf.) Rybd. Ex A.W. Hill] and sicklepod (Senna obtusifolia), and two pathogens, (Colletotrichum truncatum and Alternaria cassiae), that are bioherbicidal agents against hemp sesbania and sicklepod, respectively. Pathogenicity tests, and assays for extractable, and in vitro CS activities were utilized. Phytotoxicity bioassays indicated that the bulky t-butoxycarbonyl moiety substitution on the AOA molecule did not substantially hinder expression of biological activity of Boc-AOA in these tests. Generally, spray application of the compounds to young dark-grown seedlings caused little growth effects, but root-feeding of the chemicals reduced growth (stem elongation) in both weeds. Hemp sesbania was generally more tolerant than sicklepod to these compounds. The only apparent positive interaction of the chemicals with these pathogens was the Boc-AOA: C. truncatum combination treatment on hemp sesbania. Both compounds reduced extractable CS in the seedlings by 30%, 72 h after treatment. CS activity was reduced by 15% in hemp sesbania treated with C. truncatum but increased 20% above control levels after infection of sicklepod by A. cassiae. This latter effect suggests that CS may be involved in sicklepod defense mechanisms against this pathogen.

Do Aqueous Extracts from Buckwheat Compromise Seed Germination and Initial Development of Bidens pilosa and Euphorbia heterophylla?

Fagopyrum esculentum Moench (buckwheat) is a dicot species from the Polygonaceae family used as a cover crop in agricultural systems featured with a remarkable allelopathic potential for weed control, helping herbicide-resistance management and promoting substantial reductions in herbicide applications. The aim of this research was to examine the allelopathic potential of aqueous extracts from seeds and aerial part of buckwheat on seed germination and initial development of Bidens pilosa and Euphorbia heterophylla. Bioassay experiments were conducted under a completely randomized experimental design with four replications, containing 50 seeds each. Both weed seed species were harvested in a soybean field, and seed viability was previously assessed. Seeds were exposed to four concentrations (0, 25, 50, and 100%) from extracts of seeds (ES) and aerial part (EAP) of buckwheat. Germination speed index (GSI) in B. pilosa and E. heterophylla was daily evaluated throughout 14 and 16 days, respectively, whereas percentage of germination, abnormal seedlings, as well as non-germinated seeds, root (RL) and aerial part length (APL), and total dry matter (TDM) were rated at final germination test. EAP reduced the GSI, especially under the 100% concentration. Germination percentage was lower and abnormal seedlings increased for both weed species when seeds were exposed to EAP concentrations greater than 25%. However, ES did not impinge upon E. heterophylla germination. EAP and ES reduced the APL, RL, and TDM for concentrations greater than 50%, except for ES which did not affect E. heterophylla development. Both extracts from buckwheat have a high capacity to inhibit germination and compromise seedling development, culminating in such a potential alternative for B. pilosa and E. heterophylla management in agricultural systems.

应用生物除草剂防除高寒草地有害毒杂草的前景展望

本文简要介绍了国内外生物除草剂的研究概况和应用现状以及生物除草剂研究中需解决的主要问题 ,根据青海省草地主要有害毒杂草防治现状及存在的问题 。

Isolation and identification of Serratia marcescens Ha1 and herbicidal activity of Ha1 ‘pesta’ granular formulation

A total of 479 bacterial strains were isolated from brine （Bohai, Qinhuangdao City, Hebei Province, China）. Bioassay results indicated that 4 strains named Hal, Hal7, Ha38, and Ha384 had herbicidal activity. And strain Hal had the highest effective herbicidal activity. As a result, this study aims to JdentJfy strain Hal, characterize its physiological and biological activities, evaluate the herbicidal activity of its metabolites, and develop a ＇pesta＇ formulation and assess its effectiveness on Digitaria sanguinalis. Hal was identified as Serratia marcescens based on 16S rDNA sequencing. This strain has a flagellum, a diameter of 0.5 to 0.8 IJm, and a length of 0.9 to 2.0 IJm. The indole test shows positive results, and the catalase enzyme exhibits strong positive reactions. Results further showed that the inhibitory concentration （IC50） of the crude extracts to D. sanguinalis radicula and coleoptile were 3.332 and 2.828 mg mL-1, respectively. Both the suppression of D. sanguinalis and the cell viability of the Hal formulation in ＇pesta＇ were higher when stored at 4℃ than at （25＋2）℃. These results indi- cated that S. marcescens Hal can potentially be used as a biocontrol agent against D. sanguinalis.

Isolation, Identification, and Herbicidal Activity of Metabolites Produced by Pseudomonas aeruginosa CB-4

CB-4, a bacterial strain with highly effective herbicidal activity, was isolated from infected corn leaves. Through morphology, physiological and biochemical tests, and 16 S ribosomal DNA gene sequencing methods, CB-4 was identified as Pseudomonas aeruginosa. We conducted activity-evaluation experiments in the laboratory to assess the herbicidal potential of metabolites produced by strain CB-4. Crude extracts of strain CB-4 have high inhibition activity on Digitaria sanguinalis. In general, the root and shoot growth parameters of D. sanguinalis were significantly reduced by metabolites of strain CB-4. The IC50 of the culture filtrate extracts for the radicula and coleoptile of D. sanguinalis were 0.299 and 0.210 mg mL-1, respectively. Component 2 of the herbicidal activity of the crude toxin from strain CB-4 was successfully purified for the first time by using high-speed counter current chromatography with a two-phase solvent system composed of petroleum ether-ethyl acetate-methanol-water（4：5：4：5, v/v） and high-performance liquid chromatography. We concluded that the metabolites of strain CB-4 have the potential to be developed as a microbe-based herbicide.

The Allelopathic Effects of Sunflower and Wheat Root Exudates on Sinapis arvensis and Sinapis alba

In this study,we aimed to investigate the allelopathic effects of sunflower and wheat root exudates on the common weeds such as wild mustard and white mustard in our region.The root exudates which were obtained by soaking 8 weeks old sunflower and wheat seedlings(20 or 40 seedlings)in 100 mL of distilled water for 3 days were applied to the leaves of wild mustard and white mustard.In order to compare the allelopathic effect,the recommended dose(1 g.da^(-1))and twice the recommended dose(2 g.da^(-1))of Gromstor(Tribenuron-methyl),a herbicide preferred by farmers for the chemical control of these weeds was also applied.The allelopathy was performed for wild mustard and white mustard seedlings by the measurement of different physiological and biochemical parameters,such as chlorophyll a,chlorophyll b,total chlorophyll,carotenoid,proline,total protein amounts and superoxide dismutase enzyme activity.The amounts of total chl and carotenoid in wild mustard leaves decreased in all treatment groups compared to control.The highest decrease in total chl(50.93%)and carotenoid(46.69%)was oberved in the treatment of 40 wheat seedlings.100 mL^(-1) distilled water.In the white mustard leaves,the amount of total chl in all treatment groups except the treatment group of Gromstor 2 g.da^(-1) and carotenoid in all treatment groups increased compared to the control.The highest increases again were observed in 40 wheat seedlings.100 mL^(-1) distilled water treatment.The proline amounts in wild mustard and white mustard increased in all treatment groups.The highest increase was observed for the treatment of 20 wheat seedlings.100 mL^(-1) distilled water in wild mustard(459.69%)and 40 sunflower seedlings.100 mL^(-1) distilled water in white mustard plant(104.70%).In superoxide dismutase enzyme activities,treatments decreased activity except treatment of 40 sunflower seedling root exudate in wild mustard,while increased activity outside commercial herbicide treatment in white mustard.The results showed that sunflower and wheat root exudates have allelopathic effects on wild mustard and white mustard weeds.It is thought that the study will be a reference for new studies that will enable the use of plant root exudates as bioherbicides or foliar fertilizers and will contribute to the fight against weeds in organic agriculture.

Interaction of the Bioherbicide <i>Myrothecium verrucaria</i>with Technical-Grade Glyphosate on Glyphosate-Susceptible and -Resistant Palmer Amaranth

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.

Effects of <i>Myrothecium verrucaria</i>on Two Glyphosate-Resistant <i>Amaranthus palmeri</i>Biotypes Differing in Betacyanin Content

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.

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

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.

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

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.

Isolation and Characterization of a Phytotoxin from Xanthomonas campestris pv. retroflexus

A phytotoxin from Xanthomonas campestris pv. retroflexus was isolated using a chromatographer and HPLC, and the components were identified to be a mixture of minor molecular compounds including organic acids and cyclo-（proline-phenylalanine）. The greenhouse cultivation test was used to determine the influence of the isolated fractions on the growth of target weed redroot pigweed （Amaranthus retroflexus L）. The experimental results demonstrated that the cyclo-（Pro-Phe） had the weed inhibit activity obviously on dicotyledonous weed and the mixture with six organic acids showed stronger bioactivity. Further, greenhouse and field test were processed, and the test showed that the use of the toxin appeared to have the potential to be developed further as a bioherbicide system to control weedy grasses.

Hot Water Treatment Enhances the Bioherbicidal Efficacy of a Fungus

Hemp sesbania (Sesbania exaltata) plants (>30 cm tall) sprayed with hot water (45&deg;C – 95&deg;C), followed by spray applications of fungal spores of Colletotrichum truncatum (CT) at 1.0 × 107 spores/ml-1 and 22&deg;C – 25&deg;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&deg;C (5% mortality), and 60% mortality at 95&deg;C. Artificial dew treatments (25&deg;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&deg;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.

Biological Control Potential of <i>Colletotrichum gloeosporioides</i>for Coffee Senna (<i>Cassia occidentalis</i>)

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.

Interaction of Glufosinate and <i>Colletotrichum truncatum</i>on Ammonia Levels and Glutamine Synthetase Activity in Hemp Sesbania

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 interactions were found in the combinations of CT and glufosinate at the concentrations used, further insight on the biochemical action of CT and its interactions with this herbicide on hemp sesbania was achieved.

Interaction of the Bioherbicide Myrothecium verrucaria and Glyphosate for Kudzu Control

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.

Extending the Shelf-Life of <i>Myrothecium verrucaria</i>, a Bioherbicide

The shelf-life of a bioherbicide product is an important factor with regard to its commercial potential. The bioherbicidal efficacy of freshly fermented Myrothecium verrucaria (strain IMI 368023) (MV) mycelia formulations and MV mycelia preparations that had been freeze-dried and then stored at -20&deg;C for 8 years was compared. Two concentrations of each formulation (1.0x and 0.5x) were tested, utilizing bioassays on seedlings of the weed, hemp sesbania (Sesbania exaltata) under greenhouse conditions or in darkness utilizing hydroponically grown seedlings. Freeze drying of freshly prepared MV mycelium produced a light, brownish-colored powder. Efficacy tests of this reconstituted 8-year-old dried material showed that some bioherbicidal activity was lost during long-term storage, i.e., ~20% and ~60% seedling dry weight reduction at the 1.0x and 0.5x rate, respectively. Although plant mortality was greater in the fresh mycelial preparations treatments versus the freeze-dried and stored samples at all time points in the time-course, the stored material still caused >80% mortality, 15 days after treatment. Comparative disease progression ratings also showed a similar trend. Overall results show that freeze-drying MV is a useful method to reduce the bulk and cumbersomeness of storing heavy liquid fermentation product, while retaining bioherbicidal activity. These findings increase the utility of this bioherbicide and offer the potential to use the dried material in soil treatments or in a more concentrated form than attainable via the fermented product.

Effects of <i>Myrothecium verrucaria</i>on Ultrastructural Integrity of Kudzu (<i>Pueraria montana var. lobata</i>) and Phytotoxin Implications

The fungus Myrothecium verrucaria (Alb. & Schwein.) (MV), originally isolated from diseased sicklepod (Senna obtusifolia L.), has bioherbicial activity against kudzu and several other weeds when applied with low concentrations of the surfactant Silwet L-77. To more fully understand the initial events of MV infection or disease progression, and to improve knowledge related to its mechanism of action, the effects of MV and its product (roridin A) on kudzu seedlings were examined at the ultrastructural level. Ultrastructural analysis of MV effects on kudzu seedlings revealed a rapid (~1 h after treatment) detachment of the protoplast from the cell wall and plasmodesmata appeared to be broken off and retained in the wall. These symptoms occurred well in advance of the appearance of any fungal growth structures. Some fungal growth was observed after severe tissue degeneration (24 to 48 h after treatment), but this occurred primarily at the extra-cellular location with respect to the kudzu tissues. Kudzu seedlings treated with roridin A, a trichothecene produced by the fungus, exhibited some symptoms similar to those induced by the fungus applied in spore formulations with surfactant. The overall results are the first to report the ultrastructural effects of this bioherbicide on plants and suggest that penetration of a phytotoxic substance(s) in the fungal formulation was facilitated by the surfactant, and that roridin A exerts phytotoxicity toward kudzu.

Biological Control of the Weed Sesbania exaltata Using a Microsclerotia Formulation of the Bioherbicide <i>Colletotrichum truncatum</i>

Colletotrichum truncatum, grown on rice grain (3 to 4 weeks, 22°C to 24°C) produced a fungus-infested rice mixture of microsclerotia and conidia (spores) in a ratio of ~9:1, respectively. Greenhouse tests of this formulation (0.4 to 50 mg finely-ground fungus-rice product) which applied pre-emergence to 5 cm2 of soil surface, caused 22% to 96% hemp sesbania plant mortality, after 14 days. Post-emergence treatment (fungus-rice aqueous formulation;2.4 × 105 microsclerotia ml-1, 30% unrefined corn oil and 0.2% Silwet L-77 surfactant) of weeds surviving the pre-emergence application, resulted in 93% mortality, after 14 days. Based on greenhouse results, field tests were undertaken: 1) pre-emergence treatment (fungus-rice formulation at 2.4 × 105 microsclerotia cm-2), 2) post-emergence (fungus-rice product in 30% unrefined corn oil, 0.2% Silwet) only treatment, applied 15 days after planting and 3) pre-emergence treatment followed by post-emergence treatment (fungus-rice product in 30% unrefined corn oil, 0.2% Silwet) applied 15 days after planting to surviving weeds. Control treatments were: 1) autoclaved rice product sans fungus, 2) unrefined corn oil (30% unrefined corn oil, 0.2% Silwet in water) and 3) untreated plants. Planting dates were: early season (April-May), early-mid season (June-July), late-mid season (July-August), and late season (September-October). Weed mortality was recorded at 15 days for the pre-plus post-treatment, and at 30 days after planting for the pre-emergence only and the post-treatment only. The early season, pre-emergence treatment caused 67% hemp sesbania mortality (3-yr average) within 15 days and the post-emergence treatment caused 91% mortality of the surviving weeds. In the late-mid-season, pre-emergence treatment caused minimal (<5%) mortality at 15 days, but mortality in the post-emergence treatment was >80%. Results suggest that seasonal environmental conditions are important in the efficacy of this C. truncatum-rice product formulation when applied pre- or post-emergence to this onerous weed.

Interaction of a <i>Myrothecium verrucaria</i>Mycelial Preparation and a Glyphosate Product for Controlling Redvine (<i>Brunnichia ovata</i>) and Trumpet Creeper (<i>Campsis radicans</i>)

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.

Study on RNAi-based herbicide for Mikania micrantha

The invasive plant Mikania micrantha Kunth(M.micrantha)from South America poses a significant threat to the stability and biodiversity of ecosystems.However,an effective and economical method to control M.micrantha is still lacking.RNA interference(RNAi)has been widely studied and applied in agriculture for trait improvement.Spray-induced gene silencing(SIGS)can produce RNAi silencing effects without introducing heritable modifications to the plant genome and is becoming a novel nontransformation strategy for plant protection.In this study,the genes encoding chlorophyll a/b-binding proteins were selected as targets of RNAi,based on high-throughput sequencing of M.micrantha transcriptome and bioinformatic analyses of sequence specificity.Three types of RNAi molecules,double-stranded RNA,RNAi nanomicrosphere,and short hairpin RNA(shRNA),with their corresponding short interfering RNA sequences were designed and synthesized for SIGS vector construction,from which each RNAi molecule was transcribed and extracted to be sprayed on M.micrantha leaves.Whereas water-treated control leaves remained green,leaves treated with RNAi molecules turned yellow and eventually wilted.Quantitative real-time PCR showed that the expression levels of target genes were significantly reduced in the RNAi-treated groups compared with those of the control,suggesting that all three types of RNAi herbicides effectively silenced the endogenous target genes,which are essential for the growth of M.micrantha.We also found that shRNA showed better silencing efficiency than the other two molecules.Taken together,our study successfully designed three types of RNAi-based herbicides that specifically silenced endogenous target genes and controlled the growth of M.micrantha.Moreover,we identified a gene family encoding chlorophyll a/b-binding proteins that is important for the growth and development of M.micrantha and could serve as potential targets for controlling the spread of M.micrantha.