Seed treatment technology: An attractive delivery system for controlling root parasitic weed Striga with mycoherbicide

Coating sorghum seeds with Fusarium oxysporum (Foxy 2) for control of the root parasitic weed Striga, appears to be an attractive option for minimizing the inoculum amount, establishing the biocontrol agent in the potential infection zone of the host plants, and offering a simple, easy and economical delivery system. Our investigations resulted in the selection of appropriate seed coating materials and a suitable type and form of fungal inoculum. The coating materials tested were arabic gum (AG10%, 20%, 40%), carboxymethylcellulose (CMC1%, 2%) and pectin (LS 440, LM-5 CS) 1%, while the fungal inoculum included microconidia and fresh and dried chlamydospores produced using different substrates. Foxy 2 survived the seed treatment processing and showed excellent viability on seeds for at least 8 months of storage after coating. In general, the performance of 40% arabic gum in combination with dried chlamydospores was the best among the other types of inoculum and coating material tested. Regardless of the type and form of inoculum and coating materials tested, Foxy 2 was able to colonize all roots, even root tips and hairs of the host (sorghum), thereby meeting important criteria of a promising candidate for controlling Striga when applied as a seed treatment. The efficacy of treated sorghum seed with Foxy 2 using different coating materials in reducing S. hermonthica infestation was evaluated in pot and root chamber trials. Foxy 2 markedly reduced Striga emergence and dry weight and increased the percentage of the diseased emerged Striga shoots. However, the efficacy of seed coating varied according to the type and form of fungal inoculum as well as coating material. Coating sorghum seed with dried chlamydospore inoculum homogenized into 20% arabic gum (as adhesive) showed the highest efficacy of 81 and 77% (i.e., percent reduction in healthy emerged Striga shoots compared to the control treatment) against Striga using either sterilized or non-sterilized soil, respectively. In root chamber bioassays, the application of Foxy 2 in combination with AG40% significantly caused disease in 77% of the germinated Striga seeds and in all tubercles after 25 days of sowing. These findings provide an optimized coating protocol as an attractive delivery system for bioherbicides for root parasitic weeds.     

Development and assessment of fusarium oxysporum-based mycoherbicide for control of Striga Hermonthica in sorghum

Abstract

Experiments were carried out from 2002 to 2003 to determine the most suitable form of fungal delivery for possible use by farmers in biological control of Striga hermonthica. Six mycoherbicides were developed, based on Fusarium oxysporum isolated from wilted S. hermonthica. In mycoherbicide formulation, rock phosphate powder, sorghum bran and gum arabic powder were used as carriers. Besides its role as a carrier, gum arabic powder was used as a sticker. There were three carriers with two formulations each, making six treatments altogether. Living propagule studies were based on colony, mycelium and conidium number of F. oxysporum. In greenhouse evaluation of mycoherbicides, each kg sorghum seed was coated with 10 g mycoherbicide before sowing. Carrier rock phosphate powder with gum arabic powder as a sticking agent was the most suitable form of its delivery for use by peasant farmers.     

Induction, Morphogenesis, and Germination of the Chlamydospore of Candida albicans

Early log phase yeast cells of Candida albicans transformed into suspensor cells and chlamydospores when streaked on washed agar without added nutrients. The transformation was apparently a result of endogenous metabolism since starved yeast cells did not form chlamydospores. Addition of glucose (5 mg/ml) to washed agar completely suppressed chlamydospore formation. Size of inoculum and age of inoculum markedly affected chlamydospore yield. Electron microscopy of thin sections revealed the chlamydospore wall to be double layered, the outer thin layer being continuous with the wall of the suspensor cell. A technique was devised to study germination of chlamydospores. Chlamydospores germinated by budding, and the fluorescent antibody technique was used to study the budding process.     

Isolation of Fusarium oxysporum with Potential for Biocontrol of the Witchweed (Striga hermonthica) in the Nigerian Savanna

A fungus isolated from wilted Striga hermonthica plants was identified as Fusarium oxysporum. A comparison of this isolate (PSM-197) with four other Fusarium spp. for control of S. hermonthica using conidial foliar sprays, showed that it was the most pathogenic and virulent. The isolate, grown on sorghum grain and incorporated into soil, completely inhibited the emergence of S. hermonthica compared with other substrates.     

Influence of Agricultural By-products in Liquid Culture on Chlamydospore Production by the Potential Mycoherbicide Fusarium oxysporum Foxy 2

Economically feasible inoculum mass production methods are required for successful application of Fusarium oxysporum Foxy 2 as a potential mycoherbicide. Therefore, different substrates (agricultural by-products) and the factors that influence the production of spores, especially chlamydospores, of Foxy 2 were investigated in liquid cultures. The substrates tested were cotton seed cake, maize stover, wheat and triticale stillage. The presence of plant fibers in the medium of unfiltered cotton seed cake (2.5%, w/v) significantly enhanced chlamydospore, micro- and macroconidia production by 150, 185, and 300%, respectively, compared to the filtered (fiber-free) medium. Regardless of the type of substrate tested, Foxy 2 was able to produce abundant chlamydospores (0.14-2.7×10⁷ mL^-1) in all growth media. Generally, increasing the concentrations of cotton seed cake and maize stover in the medium significantly increased chlamydospore formation; however, this was not the case for wheat-based stillage. To optimize conditions required for chlamydospore production of Foxy 2, the effect of near ultra-violet (NUV) light, substrate combinations (synergism), level of agitation, incubation time and their interaction were studied. A liquid culture of 2.5% (w/v) filtered cotton seed cake, exposed to continuous NUV for 15 days, doubled the yield of chlamydospores (4.7×10⁶ mL^-1) and macroconidia (5×10⁵ mL^-1), and increased microconidial production by one-third (1×10⁸ mL^-1) compared to natural light. An apparent synergistic effect of substrate combination was observed, since the addition of 20% (v/v) of either wheat or triticale-based stillage to maize stover medium (1%, w/v) increased the number of chlamydospores produced by 16 or 12 times, respectively, compared to maize stover alone (1.4×10⁶ mL^-1), and 2 times more than either of the stillages. A significantly positive effect between a high level of agitation tested during incubation and chlamydospore production of Foxy 2 was recorded. All in all, substantial chlamydospore production of Foxy 2 (4.3×10⁷ mL^-1) was successfully achieved within 12 days of incubation in a single-step liquid fermentation through the combination of 0.5% (w/v) maize stover plus 20% (v/v) wheat-based stillage and a high level of agitation (200 rpm).     

Identification of Intestinal Bacteria Responsible for Fermentation of Gum Arabic in Pig Model

Acacia spp. produce gum exudates, traditionally called gum arabic or gum acacia, which are widely used in the food industry such as emulsifiers, adhesives, and stabilizers. The traditional gum arabic is highly variable with average molecular weights varying from 300,000–800,000. For this reason a standardized sample was used for the present experiments, based on a specific species of gum arabic (Acacia(sen)SUPER GUM^TM EM2). The literature indicates that gum arabic can be fermented by the intestinal bacteria to short chain fatty acid, particularly propionate. However, the bacteria responsible for the fermentation have not been determined. In this study, we used enrichment culture of pig cecal bacteria from the selected high molecular weight specific gum arabic of (M_W 1.77 × 10⁶). We found Prevotella ruminicola-like bacterium as a predominant bacterium that is most likely to be responsible for fermentation of the gum arabic used to propionate.     

Agricultural By-products as Substrates for Growth, Conidiation and Chlamydospore Formation by a Potential Mycoherbicide, Fusarium oxysporum Strain EN4

Characteristics, such as mycelial growth, conidiation and chlamydospore formation, of three distinct variants (EN4-FT, EN4-FF and EN4-S), of the potential mycoherbicide, Fusarium oxysporum, strain EN4 were studied in liquid cultures. Agricultural by-products, such as cornmeal, corn cob, soya bean hull fiber (Dietfiber-Soyfiber), cotton seed embryo flour (Pharmamedia), cane molasses yeast extract and potato dextrose medium were used as substrates. The mycelial growth and conidia and chlamydospore formation were different for the three strains and varied with the substrate used. The quantity of conidia and chlamydospores produced depended on the concentration of the substrates. Irrespective of carbon:nitrogen ratios, chlamydospores were formed readily in liquid media and in greater amount on substrates with low utilizable carbon content (aqueous extracts of soya bean hull fiber and corn cob) than on those with higher utilizable carbon content (potato dextrose broth, aqueous extracts of cotton seeds and molasses yeast extract broth). In liquid cultures, increases in the concentration of substrates high in utilizable carbon resulted in reduced chlamydospore formation; however, this was far less in those with a lower carbon content. The effect on fungal growth characteristics due to the presence or absence of plant fibers in various concentrations of unfiltered and filtered extracts of soya bean hull fiber depended on the fungal variant tested. However, it had a greater effect on viable and microconidial counts than on chlamydospore and macroconidial counts. The viable and microconidial counts were significantly higher in unfiltered than in filtered extracts. The final pH of the liquid cultures (4.1-8.5), after 10 days of growth, was affected by the fungal variant, the substrate used and by the substrate concentration. In liquid cultures with 1% soya bean hull fiber, chlamydospore formation corresponded to an increase in the pH of the medium from 6.0 to 7.7. The interaction between fungal variants, the substrate used and the substrate concentration was highly significant for all the variables tested, indicating that optimization of the production of fungal propagules is complex. Overall, for all three variants, the best substrates for optimum conidiation and chlamydospore formation were aqueous extracts of soya bean hull fiber and corn cob at concentrations of 1-2.5%.     

Sorghum straw for xylanase hyper-production by Thermomyces lanuginosus (D2W3) under solid-state fermentation

This paper reports the production of very high levels of cellulase free xylanase and associated hemicellulases by an indigenous thermophilic isolate of Thermomyces lanuginosus (D(2)W(3)) using solid-state fermentation. Sorghum straw, an inexpensive and abundant source of carbon supported maximal xylanase activity (11,855 units/g dry substrate). Culturing T. lanuginosus D(2)W(3) on sorghum straw and optimizing other culture conditions (media types, particle size of carbon source, inoculum level, inoculum age and additives), yielded increased levels of xylanase (39,726 units/g dry substrate). Further optimization of enzyme production was carried out using Box-Behnken design of experiments with three independent variables (inoculum level, glycerol and ammonium sulphate concentrations) which resulted in very high levels of xylanase, 48,000+/-1774 units/g dry substrate, and 2.6+/-0.2, 13.4+/-0.56, 68+/-1.7, 1.4+/-0.08, 1.2+/-0.05 (units/g dry substrate) of beta-xylosidase, alpha-galactosidase, pectinase, beta-mannosidase and alpha-L-arabinofuranosidase, respectively.     

Interactions of Phytophthora cinnamomi and Trichoderma spp. in relation to propagule production in soil cultures at 26 degrees C1

Effects of Trichoderma harzianum and T. polysporum on chlamydospore production by two isolates of Phytophthora cinamomi were determined over a 21-day period in nonsterile, sterilized, and sterilized amended soil. Trichoderma was either coinoculated with P. cinnamomi or added to the cultures on day 3 of the incubation period. In nonsterile soil, conversion of mycelial fragments in the P. cinnamomi inoculum to chlamydospores resulted in an initial slight increase in chlamydospore numbers. In cultures where either of the Trichoderma isolates was added on day 3, a significiant reduction in chlamydospore numbers was observed on day 4; however, neither of the Trichoderma isolates at either inoculation time significantly affected clamydospore numbers by day 21. Results from studies with sterilized amended soil indicated that a reduction in P. cinnamomi chlamydospore numbers by either of the Trichoderma isolates was dependent upon availability of simple carbohydrates. In sterilized nonamended soil, the number of chlamydospores was increased in cultures containing either Trichoderma isolate; in sterilized amended soil chlamydospore numbers were initially reduced, followed by a general increase as the nutrients were depleted. Neither of the Trichoderma isolates exhibited significant antagonistic qualities toward P. cinnamomi.     

Exudate gums: occurrence,production, and applications

This paper presents a review of the industrially most relevant exudate gums: gum arabic, gum karya, and gum tragacanth. Exudate gums are obtained as the natural exudates of different tree species and exhibit unique properties in a wide variety of applications. This review covers the chemical structure, occurrence and production of the different gums. It also deals with the size and relative importance of the various players on the world market. Furthermore, it gives an overview of the main application fields of the different gums, both food and non-food.     

Induction and morphogenesis of chlamydospores in an agerminative variant of Candida albicans

A strain of Candida albicans that did not form germ tubes was endowed with a pronounced ability for massive production of chlamydospores under appropriate environmental conditions. Development of chlamydospores was induced by N-acetyl hexosamines, especially N-acetyl-D-glucosamine, and this induction did not depend on non-specific utilization of N-acetyl hexosamines as metabolic sources nor did it correlate with induction of germ-tube formation. Formation of chlamydospores in N-acetyl hexosamine-agar medium occurs through a multiplication stage (10-12 h) consisting of a few cycles of budding leading to short, "pseudo-hypha-like" structures, followed by progressive differentiation of most cells into young chlamydospores (16-18 h) which go to complete maturation in 36-48 h. There were marked differences in chlamydospore formation among different strains of C. albicans but, when induced, the morphology and kinetics of sporulation were identical in all strains. This study shows that chlamydospore formation is not necessarily associated with the mycelial phase and suggests that N-acetyl hexosamines may induce sporulation by controlling endogenous metabolism rather than through products of their own metabolism.     

Liquid fermentation to produce biomass of mycoherbicidal strains of Fusarium oxysporum

Conditions for optimizing spore production, especially chlamydospores, by host-specific mycoherbicidal strains of Fusarium oxysporum causing vascular wilts in coca (Erythroxylum coca) and poppy (Papaver somniferum) were studied in 2.5-1 fermentors. The fermentor dissolved oxygen and pH had significant effects on the growth characteristics of F. oxysporum strains. The effect of the fungal strain, however was not significant for most of the variables studied except for chlamydospore formation. After 14 days of fermentation, the spore types produced were microconidia and chlamydospores, with very little production of macroconidia. While the total viable counts were significantly higher under high than under low dissolved O₂, the chlamydospore counts were significantly higher under low than under high dissolved O₂. The percentage of chlamydospores obtained, as a proportion of total viable was significantly higher when the fermentor pH was increased, than when it was not. Scaling-up the liquid fermentation to 20 l, yielded log₁₀ c = 6.8 (where c = chlamydospores ml⁻¹) after 14 days' fermentation, with biomass viable counts of log₁₀ v∼8.0 (where v = viable counts g⁻¹ air-dried biomass). A single-step liquid fermentation reported in this study increased chlamydospore yields and reduced the time required for their production with techniques currently available from 5 weeks to less than 2 weeks. Received: 24 April 1997 / Received revision: 6 August 1997 / Accepted: 29 August 1997     

Production of Chlamydospores and Conidia in Submerged Culture by Rhynchosporium alismatis, a Mycoherbicide of Alismataceae in Rice Crops

The objective of this work was to evaluate the production of chlamydospores and conidia of Rhynchosporium alismatis in a liquid Czapex-Dox based medium supplemented with increasing concentrations of sodium nitrate and malt extract. In addition, the germination of chlamydospores was evaluated. A high concentration of malt extract (4.4 g L^-1) as the sole carbon source and a high level of sodium nitrate as the sole nitrogen source (3.3 g L^-1) were shown to increase chlamydospore production while agitation (150 rpm) enhanced conidial yields. Maximum chlamydospore production (2.03×10⁵±0.7 total chlamydospores mg DW^-1) was achieved in cultures grown in a medium supplemented with 8.8 g L^-1 malt extract and 5.74 g L^-1 sodium nitrate. Two days growth was required for maximum chlamydospore and conidial production, while 6 days was necessary to obtain maximum dry weight accumulation (350 mg per flask). Germination of chlamydospores (90%) was significantly higher than germination of conidia (47%) after 2 days growth.     

Effects of cover crops, weeds and plant debris on development of Mycocentrospora acerina in caraway

This paper reports on the search for inoculum sources of Mycocentrospora acerina on caraway (Carum carvi L.). Obvious suspects are cover crops of biennial caraway and preceding crops of annual caraway. Other suspects are weeds in or alongside the field. Finally, survival structures of the fungus, chlamydospore chains, packed in plant debris or naked, are suspected. M. acerina is able to infect many plant species, including cover crops of caraway such as spinach for seed production and peas. However, the agronomical suitability of a crop to serve as a cover crop of biennial caraway proved to be a more important factor in determining caraway yield than the susceptibility of the cover crop to M. acerina. This finding was corroborated by the fact that spinach and peas as preceding crops had no significant effects on M. acerina development in spring caraway sown the next year. Dill, barley and four weed species were found as new hosts of M. acerina. The role of weed hosts, susceptible crops and plant debris in the survival of the fungus in years without caraway is discussed. Caraway sown on soil containing infested caraway straw, infested debris of other plant species or chlamydospores grown in pure culture, became infected by M. acerina. Only high inoculum densities of chlamydospores in the soil caused severe damping-off of caraway seedlings. The opportunity for disease management by agronomical means is quite limited.     

Roles of low pH, carbon and inorganic nitrogen source use in chlamydospore formation by Fusarium solani.

Citrate and malate were poorer sources of exogenous carbon than several hexose, pentose, or disaccharide sugars for supporting macroconidial germination by Fusarium solani at high conidial density (1 X 10(5) condia/ml). Only citrate, however, failed to block chlamydospore morphogenesis to a degree comparable to glucose or other readily used sugars. Mostly immature chlamydospores were formed in the presence of citrate. At low conidial density (5 X 10(3) conidia/ml), exogenous carbon-independent macroconidial germination and subsequent rapid chalmydospore formation on germ tubes was not inhibited by ammonium or nitrate nitrogen. The citrate-phosphate buffered, low pH (4.0) medium of Cochrane induced more immature chlamydospore formation by F. solani than a pH 6.0 medium, but few mature chlamydospores were formed in either medium. Condensation of hyphal cytoplasm into developing chlamydospores, a character typical of chlamydospore formation, did not occur extensively and macroconidia, hyphae, and immature chlamydospores stained deeply with Sudan III, suggesting lipid biosynthesis. This inhibition of chlamydospore maturation may be due partly to nitrogen deficiency imposed by the high C:N ratio of the medium and to the presence of citrate. Only vesiculate hyphal cells were formed by F. solani f. sp. phaseoli in both media. Field soils to which the clone of F. solani used is indigenous had mean pH values ranging from 5.2 to 6.0.     

Composition of the gum from Combretum paniculatum and four other gums which are not permitted food additives.

Only three gum exudates are permitted for pharmaceutical and food use by international regulatory authorities, viz. gum tragacanth (Asiatic Astragalus spp.), gum karaya (Sterculia spp.) and gum arabic [Acacia senegal (L.) Willd.], but a wide range of other tree exudates is used for a variety of uses in their countries of origin. This paper presents analytical data for the gum exudates from Atalaya hemiglauca, Cassine aethiopica, Combretum paniculatum, Sclerocarya birrea, and Pseudocedrela kotschyi. These gums may have local technological applications, but are not recommended for addition to foodstuffs.     

In vivo-Untersuchungen zur Entstehung und Funktion der Chlamydosporen von Botrytis cinerea Pers. am Wirt-Parasit-System Fuchsia hybrida – B. cinerea

In vivo-investigations on the formation and function of chlamydospores of Botrytis cinerea Pers. in the host-parasite-system Fuchsia hybrida–B. cinerea On naturally with Botrytis cinerea Pers. infected and artificially inoculated outdoor- and greenhouse-plants of Fuchsia hybrida the extra- and intramatrical formation of the B. cinerea- chlamydospores was investigated. The chlamydospores served 1. as structures of survival, which were tested with regard to their tolerance of drought, nutrient- and oxygen-deficiency, attack by bacteria and pH-requirements. 2. The chlamydospores represented dispersal units, which were capable of germination. 3. The chlamydospores could function as structures of infection, because after chlamydospore germination the outgrowing mycelium – either directly or after production of macroconidia – could serve as secondary inoculum and start new infections.     

家蝇幼虫对两种秸秆的消化及利用研究

本文旨在探明家蝇Musca domestic幼虫对高粱秸秆和小麦秸秆营养成分利用和木质纤维素的降解情况,以及不同配比秸秆饲料对家蝇生物学指标的影响,为今后产业化利用家蝇来生物降解秸秆类有机废弃物奠定理论基础.本研究采用70％高粱秸秆粉、70％麦秸秆粉和纯麦麸作为饲料,每种饲料各设置对照组、发酵组和家蝇幼虫取食组3种处理...