Influence of Flight and Colour Morph on Susceptibility of Sitobion avenae to Infection by Erynia neoaphidis

The influence of clone and morph on the infection of Sitobion avenae with Erynia neoaphidis was examined in laboratory assays. The LC 50 -values were low; green alates: 0.8-1.8, green apterae: 2.9, brown alates: 1.5 and brown apterae: 2.6-3.6 conidia mm -2 . Within each of the two inoculation occasions, alates of both clones were significantly more susceptible than apterae, while there was no difference between the two clones. The LT 50 -values at 18°C varied between 6.4 and 7.5 days and there was no significant difference in LT 50 -values between either the two different coloured clones or between alates and apterae.     

Targeted Dispersal of the Aphid Pathogenic Fungus Erynia neoaphidis by the Aphid Predator Coccinella septempunctata

The potential of adult and larval C. septempunctata to vector the aphid-specific entomopathogenic fungus E. neoaphidis was assessed through a series of laboratory and field experiments. The ability of coccinellids to vector conidia from a colony of E. neoaphidis -infected pea aphids, Acyrthosiphon pisum, to a colony of uninfected A. pisum was demonstrated in a laboratory study. Adult coccinellids which had previously foraged on plants infested with different densities of sporulating cadavers (1, 5, 15, 30 cadavers per plant) initiated infection in a proportion of uninfected pea aphids (4, 0, 2 and 8%, respectively) when subsequently allowed to forage on A. pisum infested bean plants. Further laboratory studies demonstrated that fourth instar larvae and adult coccinellids artificially inoculated with conidia initiated infection in 11 and 13% of an A. pisum population in which they foraged, respectively. Furthermore, a proportion of A. pisum placed on bean plants which had previously been foraged on by inoculated larval and adult coccinellids also died from infection (3 and 10% of A. pisum, respectively). However, although coccinellid adults inoculated with conidia initiated infection in 19% of A. pisum, cereal aphids, S. avenae , exposed to the inoculated coccinellids did not become infected. A further laboratory study demonstrated that infection of A. pisum only occurred if inoculated coccinellids were transferred to A. pisum populations immediately post inoculation. However, a proportion of A. pisum placed on bean plants which had been foraged on by inoculated coccinellids transferred 0, 4 and 24 h post inoculation died from infection (9, 3 and 7%, respectively). A field study further demonstrated the potential of coccinellids to vector E. neoaphidis. Single spring sown field bean plants (Long Hoos Experimental Plots, IACRRothamsted Farm) were enclosed within nylon mesh bags and 25 adult A. pisum were added to each bag with one of the following treatments: no further addition (control), coccinellid adult (control), inoculated coccinellid adult, inoculated A. pisum or sporulating A. pisum cadavers. No aphids died of E. neoaphidis in the control treatments; 5, 16 and 33% of aphids were infected with E. neoaphidis on the other treatments, respectively.     

A novel computerised image analysis method for the measurement of production of conidia from the aphid pathogenic fungus Erynia neoaphidis

A semi-automated method has been developed for the quantification and measurement of conidia discharged by the aphid pathogen Erynia neoaphidis. This was used to compare conidiation by E. neoaphidis-mycosed pea aphid cadavers, mycelial plugs cut from agar plates, mycelial pellets from shake flasks and by mycelial pellets from different phases of liquid batch fermenter culture. Aphid cadavers discharged significantly more and significantly smaller conidia than plugs or pellets. The volume of conidia discharged was stable over the period of discharge (80 h), but more detailed analysis of the size frequency distribution showed that more very small and very large conidia were discharged after 5 h incubation than after 75 h incubation. Biomass harvested at the end of the exponential growth phase in batch fermenter culture produced significantly more conidia than biomass from any other growth phase. The implications of these findings for the development of production and formulation processes for E. neoaphidis as a biological control agent are discussed.     

Effect of oleic acid on vegetative growth of the aphid-pathogenic fungus Erynia neoaphidis

Abstract The aphid-pathogenic fungus Erynia neophidis grew on a semi-defined medium containing 16 g·1⁻¹ glucose, 3 g·1⁻¹ yeast extract and 5 g·1⁻ mycological peptone only when the medium was supplemented with low concentrations of certain fatty acids. Of these, oleic acid fulfilled the growth requirement at a concentration of 0.02% (v/v), but higher concentrations were toxic, causing complete loss of viability of cultures at a concentration of 0.2% (v/v) in liquid medium and at 4% (v/v) on solid medium. The reduced viability of the fungus in liquid culture compared to that on equivalent solid medium, and at low inoculum density compared to high inoculum density in liquid medium, is explicable in terms of this toxicity.     

The effect of introducing the aphid-pathogenic fungus Erynia neoaphidis into populations of cereal aphids

The aphid-pathogenic fungus Erynia neoaphidis, as dried fungus-infected aphids, was applied to caged plots of winter wheat infested with cereal aphids at two sites, one in Hertfordshire and the other in Hampshire, in 1983. In each trial, the fungus became established in the aphid populations in the treated plots even though conditions were drier than average and therefore sub-optimal for fungus spread. Treatment applied in the third week of June increased the proportion of infected aphids more than that applied two weeks later at one site, and the early application was the only treatment to have an obvious effect at the other. In spite of the observed effect of treatments on the proportion of infected aphids, the fungus failed to reduce the numbers of aphids relative to those in untreated plots, chiefly because in these plots many aphids were killed by fungi of the same species as that introduced and other related species from natural sources. Artificial introduction of E. neoaphidis acts too slowly and unpredictably to be likely to form a practical alternative to conventional insecticides for cereal aphid control.     

Effects of 20 fungicides on the infectivity of conidia of the aphid entomopathogenic fungus Erynia neoaphidis

The effect of 20 fungicides on theinfectivity of conidia of the entomopathogenicfungus, Erynia neoaphidis, were assessedin the laboratory. After projection on broadbean leaves, conidia were treated withfungicides applied at their recommended fieldrate. Afterwards, the infectivity of theseinocula was assessed using an aphid bioassay.Four fungicides, carbendazim, kresoxym-methyl, nuarimol and thiophanate-methyl reduced the infectivity of the conidia by less than 25% and can be considered harmless for this aphid pathogen. Propiconazole was a little more toxic, with 37% reduction. Other products reducedinfectivity by between 50% and 100%. These are, from the least to the most toxic:flutriafol, prochloraz, epoxyconazole,iprodione, hexaconazole, triadimenol,azoxystrobine, cyproconazole, cyprodynil,flusilazole and tridemorph. Chlorothalonil,fenpropimorph, spiroxamine and tebuconazoletotally inhibited infectivity of the fungi. Analysis of the results according to chemicalclass showed that the benzimidazoles were theleast toxic for E. neoaphidis and themorpholines the most toxic. Effects oftriazoles and strobilurines were variable, withreduction ranging from 37% to 100% fortriazoles and from 17% to 68% forstrobilurines.     

Formulation of Bacteria for Biological Weed Control Using the Stabileze Method

Two plant pathogenic Pseudomonas spp. were formulated using the 'Stabileze' method which involves the incorporation of bacteria in a water-absorbent starch matrix with oil and sucrose, then granulating the matrix with hydrated silica. In one experiment, P. syringae pv. tabaci formulated with the standard Stabileze formula was evaluated for storage viability at -15, 2 and 22°C. Bacteria stored for 1 year at -15 and 2°C lost only 0.2 and 0.5 log 10 colony forming units (CFU) g -1 respectively compared to a loss of log 10 3.5 CFU at 22°C. In a second experiment, the same pathogen was evaluated using variations of the formula with and without oil, and with and without sucrose. P.s. pv. tabaci formulated with sucrose and oil in combination, and sucrose and oil alone survived better than the formulation without oil or sucrose. A third experiment tested the effect of four levels of oil and four levels of sucrose (4 ×4 factorial) on survival of P.s. pv. tagetis over a 28 month period. Sucrose alone enhanced survival more than oil alone, and the beneficial effect of the sucrose was reduced when it was combined with oil. These experiments suggest that the Stabileze protocol is effective for stabilizing bacteria, but there are differences in response to different formulation components between species of bacteria.     

An Alginate Prill Formulation of Fusarium oxysporum Schlechtend: Fr. f. sp. erythroxyli for Biocontrol of Erythroxylum coca var. coca

A rice alginate prill formulation of isolate EN - 4 of Fusarium oxysporum f . sp . erythroxyli, pathogenic to Erythroxylum coca var . coca (coca) , was evaluated in greenhouse and field studies for its ability to enhance pathogen populations in the soil and cause disease in coca . The formulation was applied to four different soil types in the greenhouse at 33 . 6 kg ha 1 . It enhanced the population of EN - 4 in each soil and most ( > 90%) of the fungal population remained in the upper 5 cm of soil during the 49 - day experiment . When applied in field experiments , the formulation enhanced the population of EN - 4 in the soil . Isolate EN - 4 was present in the upper 7 . 6 cm of soil 28 days after application at populations similar to those in the greenhouse studies (1 103 to 1 104 colony - forming units (CFUs) / g of soil) . Elevated populations of the pathogen (1 102 CFUs / g of soil) were still present in treated soils 229 days after application of the formulation . The areas used for field studies were already infested with the pathogen and typically developed high levels of fusarium wilt within 2 years of planting with coca . The formulated F. oxysporum began having a significant effect on plant death 100 - 200 days after application based on repeated measures analysis . These data suggest that a formulation of F. oxysporum f . sp . erythroxyli which enhances the incidence of fusarium wilt in coca fields can be produced using established techniques .     

Evaluation of a Powder Formulation of Pseudomonas fluorescens Pf1 for Control of Rice Sheath Blight

Pseudomonas fluorescens strain Pf1, inhibitory to the growth of the rice sheath blight pathogen Rhizoctonia solani in vitro , was developed as a talc-based formulation. W hen the formulation was applied as a seed treatment, the bacteria established well in the rice rhizosphere. Root treatment or soil application of the formulation was less effective in establishing the bacteria into the rhizosphere. Effective control of rice sheath blight was obtained when the powder formulation was applied to seed, roots, soil and foliage, and these methods of application increased crop grain yield in four field trials. The possible commercial exploitation of the powder formulation of the bacteria is discussed.     

Oxalic Acid Production and Mycelial Biomass Yield of Sclerotinia minor for the Formulation Enhancement of a Granular Turf Bioherbicide

The fungus Sclerotinia minor is presently under development in this laboratory as a granular bioherbicide for broadleaf weed species. With a view to enhancing the virulence of the fungus, the effect of increasing endogenous oxalic acid concentration through modification of the growth conditions was investigated. S. minor was grown in 125 ml of eight different liquid culture media in shake flasks incubated at 20°C for 7 days. The final pH, mycelial dry weight, and oxalic acid content of the spent growth media were determined and the virulence of S. minor grown on each solid culture medium was screened on detached dandelion leaves. A 330% increase in oxalic acid was obtained plus 56 mM of sodium succinate to Modified Richard's solution (MRS) as compared to MRS alone. A concomitant increase in virulence of 218% was expressed as increased lesion diameter.     

Suppression of the Root-knot Nematode Meloidogyne javanica by Alginate Pellets Containing the Nematophagous Fungi Hirsutella rhossiliensis , Monacrosporium cionopagum and M. ellipsosporum

The endoparasitic fungus Hirsutella rhossiliensis and the nematode-trapping fungi Monacrosporium cionopagum and M. ellipsosporum were formulated as hyphae in alginate pellets. In a soil microcosm experiment, dried pellets of all three fungi decreased the invasion of cabbage seedlings by the root-knot nematode Meloidogyne javanica when juvenile nematodes were placed 2 cm from roots; M. cionopagum was more effective than the other two fungi, reducing nematode invasion by 40-95% with 0.24-0.94 pellets cm - 3 of soil. In a field microplot experiment, in which neither H. rhossiliensis nor M. ellipsosporum suppressed nematodes, 0.5 pellets of M. cionopagum cm - 3 of soil suppressed M. javanica invasion of tomato seedlings by 73%. In a second microplot experiment with only M. cionopagum , again at 0.5 pellets cm - 3 of soil, the fungus suppressed the invasion of tomato seedlings whether the pellets were added 0, 5 or 14 days before planting; the population density of M. cionopagum increased to nearly 3000 propagules g - 1 of soil by day 8 and then declined to less than 300 by day 22. Enchytraeid worms were observed in and around damaged and apparently destroyed pellets in both microplot experiments. Whether enchytraeids consumed the fungi or otherwise affected biological control requires additional research.     

Ultrastructure of mitosis in the aphid-pathogenic fungusErynia neoaphidis

Summary An account of mitosis in the aphid-pathogenic, entomophthoraceous fungusErynia neoaphidis is presented. The mitotic apparatus is characterized by a closed, intranuclear, polarized spindle. Chromosomes are permanently attached by kinetochore microtubules (kcMTs) to the poles during mitosis. The spindle develops as the spindle pole bodies migrate and separate. At metaphase the eccentric spindle contains only kcMTs and is located in a relatively chromatinfree zone. Paired sister kinetochores are arranged in a broad metaphase plate. During anaphase kcMTs shorten, astral and nonchromosomal microtubules develop and elongate and the interpolar distance increases.     

Influence of commercially derived lipids and a surfactant on the mode of germination and process of germ-tube formation in primary conidia of two species of Erynia subgenus Neopandora (Zygomycotina: Entomophthorales

Primary conidia of the entomopathogens Erynia (subgenus Neopandora) delphacis (1 isolate) and Erynia ( Neopandora) neoaphidis (3 isolates) were stimulated to form germ-tubes with Tween 20 and with free, long-chain fatty acids, each incorporated into Entomophthora complete medium (ECM). When combined with other basal media (three tested), these compounds did not stimulate germ-tube formation. Triacylglycerols and vegetable oils, added to the same media, allowed almost complete resporulation in the fungi. In both species, Tween 20 (0.1%) encouraged greater germ-tube production (41–69%) than the fatty acids (0.1%) (≤36%). For E. delphacis, Tween 20 and the fatty acids differed significantly, but for E. neoaphidis the differences were almost always insignificant. Myristic and oleic acids stimulated germ-tube formation in both species. Palmitic acid allowed almost complete resporulation of the fungi, except for one isolate of E. neoaphidis that formed germ-tubes. Linoleic acid, tested only for E. delphacis, was fungistatic to most conidia. Higher concentrations of the fatty acids (≤1%) did not increase germ-tube formation, except 1% oleic acid which affected E. delphacis alone (>80% germination and germ-tubes). Linoleic acid, and sometimes also myristic and oleic, were fungistatic and/or toxic, depending on their concentration and on medium composition. Addition of fatty acids to ECM usually extended the lag period, and altered the morphology of the conidia and germ-tubes. These phenomena were not observed with Tween 20. Colonies were formed by E. delphacis alone, stimulated by ECM supplemented with Tween 20 or fatty acids. The results are discussed with respect to biological and physiological aspects of germination, and with respect to the mode of action of the fatty acids and the surfactant. This revised version was published online in August 2006 with corrections to the Cover Date.     

Effectiveness of Entomopathogenic Nematodes in an Alginate Gel Formulation Against Lepidopterous Pests

An improved calcium alginate gel formulation was developed and tested as a carrier for entomopathogenic nematodes against Spodoptera littoralis and Helicoverpa armigera larvae. Mortality of 100% was caused in 4th instar larvae of the two insects by feeding them on 1000 infective juveniles (IJ) g -1 of Steinernema carpocapsae (ALL strain) in the gel for 24 h. Exposing 2nd to 5th instars of H. armigera and 3rd to 6th of S. littoralis to 500 IJ g -1 of S. carpocapsae (ALL strain) resulted in 70-100% larval mortality. Mature larvae were less susceptible to the nematodes. Mortality of larvae exposed to 500 IJg -1 of S. carpocapsae (ALL strain) ranged from about 45-55% at 4 h to 90-95% at 48 h. Fourth instar larvae fed for 24 h with 250 IJ g -1 of nematode strains in gel showed in S. littoralis ranges of susceptibility in the following descending order: S. feltiae (IS -7 strain) = S. carpocapsae (DT strain) = S. feltiae (IS-6 strain) > S. carpocapsae (Mexican strain) = S. carpocapsae (ALL strain) = Heterorhabditis bacteriophora (HP-88 strain) = H sp. (IS-5 strain) > S. riobravae (Texas strain); in H. armigera the rating was: S. feltiae (IS-7 strain) = H. bacteriophora (HP88 strain) > S. carpocapsae (ALL strain) = S. feltiae (IS-6 strain ) = Heterorhabditis sp. (IS5 strain) > S. carpocapsae (Mexican strain) > S. riobravae (Texas strain) . The number of nematodes per larval cadaver increased with mortality rates. In greenhouse tests at 28 &#45 2°C and 90% relative humidity, gel discs containing 500 IJ g -1 of nematodes were pinned to leaves of potted plants of cotton ( Gossypium hirsutum ) (Acala SJ2) and the plants were offered to S. littoralis larvae. Larval mortality of 89 &#45 12.7% was caused by S. feltiae (IS-7 strain) and most of the plant leaves were protected against the larvae by the nematodes. In the control, larval mortality was 3.3 &#45 0.05% and the plants were almost completely defoliated. Possibilities of using the gel-nematode formulation to protect sheltered crops against insect pests are discussed     

Laboratory and Field Trials of Bait Formulations of the Fungal Pathogen, Metarhizium flavoviride, Against a Tropical Grasshopper and Locust

Extrusion technology was used to produce maize-starch 'contact bait' carriers, and vegetable-oil suspensions of Metarhizium flavoviride were incorporated into the baits after extrusion. Laboratory bioassays using the locust, Schistocerca gregaria, demonstrated mortality and reduction of 8 1 feeding after exposure to the baits, and gave an LC 50 of 1.5 10 spores l slope 0.63 for fifth instar nymphs. In laboratory trials, maize-starch contact baits showed improved efficacy relative to wheat-bran edible bait formulations of M. flavoviride. Field cage experiments, undertaken in Mali, using fourth and fifth instar nymphs of the grasshopper, Hieroglyphus dagenensis, demonstrated significant mortality with both fresh and weathered maize-starch contact bait formulations of M. flavoviride, relative to untreated controls.     

The Potential of Stagonospora sp. as a Mycoherbicide for Field Bindweed

Field bindweed ( Convolvulus arvensis L.) is one of the 12 most important weeds worldwide. Stagonospora sp. Isolate LA39 was isolated from diseased field bindweed plants collected in Europe. No crop tested was susceptible to the fungus, but disease symptoms were observed on other Convolvulaceae species. On field bindweed, the fungus induces disease symptoms (i.e. lesions) mainly on leaves and less severely on stems. The application of spores in an oil emulsion (10% oil in water) enhanced the disease on field bindweed plants compared with spores suspended in a 0.1% aqueous solution of the surfactant agent, Tween 80. The necrotic leaf area of inoculated plants increased as the length of exposure to 100% relative humidity (RH) and the spore density applied increased. Severe disease developed on plants inoculated with 1 107 spores/ml in oil emulsion, even in the absence of exposure to 100% RH. A delay of exposure to 100% RH (up to 8 h) did not have a significant eVect on disease severity. Field bindweed was susceptible to the fungus at all growth stages tested, but older plants were more susceptible than younger ones. It was concluded that isolate LA39 has potential as a biocontrol agent of field bindweed, especially when applied in an oil emulsion. The oil emulsion maintains the aggressiveness of the pathogen during a dew-free period and provides a favourable microenvironment during the infection process.     

Effect of spatial heterogeneity on the role of Coccinella septempunctata as an intra-guild predator of the aphid pathogen Pandora neoaphidis

The foraging behavior of starved and non-starved adult and larval Coccinella septempunctata on groups of plants in the presence of Pandora neoaphidis-infected Acyrthosiphon pisum, uninfected aphids or a mixture of these two prey types was compared. In general results of these studies confirmed the results of previous work comparing foraging behavior on a smaller spatial scale in Petri dishes. However, behaviors were modified in response to spatial complexity, prey quality, and the host plant. Starved C. septempunctata adults and larvae fed for longer and consumed more aphids than non-starved coccinellids. Both larvae and adults fed on infected aphids and in some cases entirely consumed them. This was thought to be due to the ease of capture of infected (dead) aphids and the feeding stimuli provided by the presence of the host plant and, where there was a choice of prey, uninfected aphids in the environment. Both larvae and adults spent the majority of the time foraging in the upper regions of plants and visited more plants when they were not starved or when they were in the presence of less suitable, infected aphid prey.     

甲壳素促进茂源链霉菌发酵产酶

为了提高茂源链霉菌发酵生产谷氨酰胺转胺酶的产量,研究了甲壳素对茂源链霉菌发酵产酶的影响。结果表明,添加0.5%的甲壳素对茂源链霉菌发酵产酶的促进效果极显著,但甲壳素的添加量达到2%时反而会抑制菌株产酶。从菌株生长代谢过程中p H变化、产酶情况、发酵液中蛋白含量及总氮含量等方面,对甲壳素促进茂源链霉菌发酵产酶的作用机理进行了初步探讨。研究显示,甲壳素在茂源链霉菌发酵过程中对菌体生长产生一定的胁迫,刺激菌体大量分泌次级代谢产物,从而提高茂源链霉菌的产酶。对菌株发酵过程的显微观察则表明,甲壳素也可能通过分散菌体生长,提高菌体向胞外分泌谷氨酰胺转胺酶的量来促进产酶。