Robert E. Hoagland

Effect of saponins on the growth and activity of rhizosphere bacteria

Saponins are a group of steroid or triterpenoid glycosides and related chemicals (sapogenins: non-glycosylated) found in roots, shoots, seeds, and flowers of many plant species. Saponins are of agronomic interest because of allelopathic interference with plant growth (Oleszek and Jurzysta, 1987; Waller et al., 1993). Saponins can be released into the soil by secretion from roots and/or leaching from living or decaying plant material (Mishustin and Naumova, 1955; Oleszek and Jurzysta, 1987). Conservation management practices designed to maintain plant residues on the soil surface such as the use of cover crops and reduced tillage are rapidly being adopted by growers; thus the impact of these compounds and other natural products on crop productivity and soil and rhizosphere microbial ecology warrants study.

Resistance mechanism of propanil-resistant barnyardgrass : II. In-vivo metabolism of the propanil molecule

Propanil-resistant barnyardgrass populations, previously verified in Arkansas rice fields and in greenhouse tests, were examined in the laboratory to ascertain if the resistance mechanism in this weed biotype was herbicide metabolism. Propanil-resistant barnyardgrass was controlled >95% in the greenhouse when carbaryl (an aryl acylamidase inhibitor) was applied two days prior to propanil. Laboratory studies with 14 C-radiolabelled propanil indicated that the herbicide was hydrolysed in propanil-resistant barnyardgrass and rice to form 3,4-dichloroaniline, but no detectable hydrolysis occurred in susceptible barnyardgrass. Two additional polar metabolites were detected in propanil-resistant barnyardgrass and rice and tentatively identified by thin layer chromatography. Overall, metabolites in the resistant barnyardgrass had R f values similar to those in rice, indicating similar metabolism for both species. These data, coupled with data from a previous report on the resistant biotype showing no differential absorption/translocation or molecular modification of the herbicide binding site in the resistant biotype, indicate that the resistance mechanism is metabolic degradation of propanil.

Glyphosate and bioherbicide interaction for controlling kudzu (Pueraria lobata), redvine (Brunnichia ovata), and trumpetcreeper (Campsis radicans)

Abstract In controlled environment experiments, the bioherbicidal fungus Myrothecium verrucaria (Alb. & Schwein.) Ditmar:Fr. was tested alone, in combination with, prior to, and following treatment with glyphosate [N-(phosphonomethyl)glycine] for control of kudzu [Pueraria lobata (Willd.) Ohwi], redvine [Brunnichia ovata (Walt.) Shinners], and trumpetcreeper [Campsis radicans (L.) Seem. ex Bureau] at temperatures of 20, 30, and 40°C. At all temperatures, kudzu was most adversely affected by the fungus, followed by trumpetcreeper and redvine, as indicated by greater mortality and dry weight reductions. Trumpetcreeper and redvine mortalities and dry weight reductions significantly increased when the fungus was applied 2 days after the glyphosate treatment. Application of the fungus combined with or prior to glyphosate treatment resulted in reduced weed control. Although pathogenesis and mortality also occurred at 20°C, disease development was favored by higher temperatures (30 and 40°C). Infected weeds of each species exhibited similar disease symptomatology within 12 h following treatment at incubation temperatures of 30 and 40°C. Disease symptomatology was characterized by necrotic flecking on leaves that coalesced into large lesions. Symptoms progressed, initially infecting cotyledons and leaves, and later (within 48 h) producing stem lesions. The fungus sporulated profusely on infected tissue and was easily reisolated. These results suggest that timing of glyphosate application in relation to combined treatment with the bioherbicide M. verrucaria can improve the control of kudzu, redvine, and trumpetcreeper.

BIOHERBICIDES: RESEARCH AND RISKS

Many microbes have bioherbicidal activity, and several phytopathogenic fungi and bacteria have been patented as weed-control agents. The phytotoxic components of most agents have not been elucidated, but some phytotoxins and other secondary compounds produced by such microbes may be toxic to mammalian systems. Furthermore, few rigorous assessments have addressed uptake, translocation, metabolism, and persistence of these phytotoxins (some of which have not been identified), or the environmental effects of repeated augmentative applications of these microorganisms on long-term impact, environmental fate, or interactions with other microbial communities. Generally, there is a lack of definitive research on the overall toxicological risk of bioherbicidal microorganisms to the degree achieved or required for synthetic herbicides. This article presents a brief overview of bioherbicides (microorganisms and/or their phytotoxins), with emphasis on the toxicity of certain bioherbicides, and general considerations of risks associated with bioherbicidal use. A subsequent article presents some of our research results and future research directions on efforts to develop the bioherbicidal fungus Myrothecium verrucaria as a safe and efficacious bioherbicide have been published elsewhere.

Evaluation of the bioherbicide Myrothecium verrucaria for weed control in tomato (Lycopersicon esculentum)

Abstract An isolate of the fungus Myrothecium verrucaria was evaluated for its biocontrol potential against common purslane, horse purslane, spotted spurge, and prostrate spurge, all serious weed pests in commercial tomato fields in the southeastern US. In greenhouse and field tests, M. verrucaria was highly virulent against these weeds when applied as conidial sprays formulated in 0.2% Silwet L-77 surfactant, even in the absence of dew. In field test plots naturally infested with these weeds, seedlings in the two-to-three leaf growth stage treated with M. verrucaria at 2×107 conidia mL−1 in 0.2% Silwet, exhibited leaf and stem necrosis within 24 h following inoculation, with mortality occurring within 96 h. After 7 days, M. verrucaria had killed 90–95% of both purslane species and 85–95% of both spurge species. Tomatoes that were transplanted into plots treated with M. verrucaria remained healthy and vigorous throughout the growing season. Since M. verrucaria effectively controlled several common weeds under field conditions, this fungus appears to have potential as an effective bioherbicide for pre-plant weed control in production systems with transplanted tomato.

Myrothecium verrucaria isolates and formulations as bioherbicide agents for kudzu

Abstract The fungus Myrothecium verrucaria (MV) has previously been shown to have potential as a bioherbicide for kudzu (Pueraria lobata) control. It has also been shown that MV wild-type (MV-wt) often forms sectors, when grown on various nutrient media. Experiments compared MV-wt and MV sector efficacy when grown on agar or on rice grains. In greenhouse evaluations of sectors, applied as foliar sprays in water or in other formulations (corn oil, surfactant, and corn oil plus surfactant) for efficacy against kudzu seedlings, some sectors possessed bioherbicidal activity equal that of MV-wt, but others exhibited lower activity. Without a dew period, aqueous formulations of MV-wt, a yellow sector, and a white sector provided zero control, but all three isolates were active without a dew period when formulated in corn oil, Silwet L-77 surfactant, and in surfactant plus corn oil. Generally, the yellow sector was less effective than the other two isolates in any formulations, and the MV-wt and white sector provided approximately 100% mortality of the test plants. Dew (10 h) increased weed control to 100, 33, and 65%, respectively, for MV-wt, the yellow sector and the white sector. All isolates provided nearly 100% control in the oil and surfactant formulations with a dew period compared to treatments receiving no dew. Soil incorporation studies were also performed to compare MV-wt efficacy of preparations grown on agar versus growth on rice grains. Higher efficacies (1.75–3.3-fold increase) were obtained from rice grain preparations compared to preparations grown on agar, when preparations were incorporated at several rates into soil prior to planting. Cell-free extracts of the MV-rice cultures were also phytotoxic to kudzu seedlings up to the eight- to 10-leaf growth stage. Thus, formulation, growth media, and the application method are important determinants in the efficacy of MV and MV sectors on kudzu seedlings.

Studies of the Phytotoxicity of Saponins on Weed and Crop Plants

Several saponins or sapogenins including β-escin, betulin, β-glycyrrhetinic acid, hecogenin, oleandrin, and oleanolic acid were tested in the laboratory, growth chamber, and greenhouse on various weed and crop species. Seed germination, root and shoot growth after root, foliar, or soil application, electrolyte leakage from leaf discs, and greening of etiolated plant tissues were monitored. Esterase activity using fluorescein diacetate (FDA) and p-nitrophenyl butyrate (PNPB) was also assayed. The compounds had differential effects on these parameters, depending on the species. The effects of these compounds on electrolyte leakage ranged from no effect to a 10-fold increase above untreated tissue levels after 72 h. Escin increased FDA activity up to 35% above untreated tissue, but other compounds caused no effect or reduced FDA activity. PNPB activity was generally not affected. In greening studies of excised tissue, escin reduced chlorophyll production by 90–100% in several species, with other compounds giving intermediate or no effects. Foliar application (1.0 mM) in the greenhouse had no substantial effect (visible or shoot biomass) on 10 species. However, in short-term (8 to 13 day) tests, β-escin, applied to soil at 88 and 350 μmol/kg soil, drastically reduced emergence in barnyardgrass (Echinochloa crus-galli L. Beauv.), hemp sesbania [Sesbania exaltata (Raf.) Rydb. ex A.W. Hill], wheat (Triticum aestivum L.), and soybean [Glycine max (L.) Merr.]. β-Escin also reduced growth in all species but soybean, and barnyardgrass was the most sensitive species tested. Results are discussed in relation to the role of these compounds as plant growth-regulating natural products.

Hemp sesbania (Sesbania exaltata) control in rice (Oryza sativa) with the bioherbicidal fungus Colletotrichum gloeosporioides f. sp. aeschynomene formulated in an invert emulsion

Abstract In greenhouse and field experiments, an invert emulsion (MSG 8.25) was tested with dried, formulated spores of the bioherbicidal fungus Colletotrichum gloeosporioides f. sp. aeschynomene, a highly virulent pathogen of the leguminous weed Aeschynomene virginica (northern jointvetch), but considered ‘immune’ against another more serious leguminous weed, Sesbania exaltata (hemp sesbania). A 1:1 (v/v) fungus/invert emulsion mixture resulted in 100% infection and mortality of inoculated hemp sesbania seedlings over a 21-day period under greenhouse conditions. In replicated field tests of the fungus/invert formulation conducted in Stuttgart, AR, and Stoneville, MS, hemp sesbania was controlled 85 and 90%, respectively. These results suggest that this invert emulsion expands the host range of C. gloeosporioides f. sp. aeschynomene, with a concomitant improvement of the bioherbicidal potential of this pathogen.

Evaluation of selected pharmaceuticals as potential herbicides: bridging the gap between agrochemicals and pharmaceuticals.

Much has been written in the field of agrochemicals about the application of phenoxy compounds as herbicidal agents. The illustrious history of these agents includes their accelerated development by the Allies for use during World War II. Among the most famous of these agents are 2,4-dichlorophenoxyacetic acid (2,4-D) and its congener, 2,4,5-trichlorophenoxyacetic acid (2,4,5-T). Because of their high specific activities, these compounds became templates for the production of a number of plant growth regulators such as Fruitone and Tomato Bloom. Interestingly, the pharmaceutical industry was developing a group of phenoxy derivatives which proved to be useful in the treatment of type III hyperlipoprotenemia, a condition where triglycerides and total cholesterol levels are elevated. The most recognized agent in this pharmaceutical class is clofibrate which inhibits sn-glyceryl-3 phosphate acyltransferase, the enzyme responsible for the conversion of acetate to mevalonate. Furthermore, its mechanism of action also includes inhibition of 3-hydroxy-3-methylglutaryl-CoA (HMGCoA) reductase. In addition, the aftermath of World War II served as a catalyst in the development of the benzodiazepines, a well recognized pharmaceutical class possessing sedative/hypnotic and tranquilizing properties. It is of interest to note that these agents are actually allelopathic natural products whose chemical structures were ignored until the development of Valium as a pharmaceutical agent. Had these natural product chemical templates not been relegated to library book shelves, they may have served a useful purpose in their development as medicinals. Recently, some benzodiazepines have been shown to possess agrochemical properties including inhibition against Phytophthora infestans, late blight of potato. The separate, but distinct, agrochemical and pharmaceutical development of various classes of agents will be discussed.

Bioherbicidal potential of Xanthomonas campestris for controlling Conyza canadensis

The effects of environmental parameters on bioherbicidal activity of the bacterium Xanthomonas campestris, against glyphosate-resistant and – susceptible Conyza canadensis (horseweed), were studied under greenhouse conditions. Rosette leaf-stage plants were more susceptible than older plants, and increasing inoculum from 105 to 109 cells/mL caused significantly greater plant mortality and biomass reduction of plants in both the rosette and bolting growth stages. A dew period at 25°C was required to cause an 80% and 60% mortality of plants in the rosette and bolting growth stages, respectively. Results indicate that X. campestris can infect and kill horseweed, demonstrating its bioherbicidal potential.