Ecological factors in the inundative use of fungal entomopathogens

Fungal entomopathogens have been developed in numerous countries as biocontrol agents with more than 100 mycoinsecticide products commercially available in 2006. The chief, perhaps sole, use of these mycoinsecticides has been as inundative agents, within a chemical paradigm. Large numbers of propagules are applied in an attempt to overwhelm by brute force many of the factors that keep a pathogen in nonepizootic equilibrium with its host. This review attempts to summarize what we know about the abiotic and biotic factors that affect the efficacy of these mycoinsecticides in both foliar and soil applications. Sunlight, humidity, temperature, and phylloplane-associated factors can affect both immediate efficacy and persistence on plants. Likewise, soil texture-moisture interactions, temperature, and a host of biotic factors can affect mycoinsecticides in the soil. Despite much research, our understanding of these ecological aspects is imperfect, especially in a holistic, dynamic sense.     

The impact of biotechnology on hyphomycetous fungal insect biocontrol agents

The potential for the control of insect pests by entomopathogenic fungi has been touted for decades, if not centuries. Only recently have advances in biotechnology provided the tools for indepth analysis of the mechanisms involved in pathogenesis and host death at the molecular level. This review outlines the current state of knowledge regarding the mode of infection and targets several key components that are amenable to improvement via biotechnology. Realization of the considerable economic potential of fungal bioinsecticides can occur only through a combined and coordinated effort involving fundamental science, formulation technology and field applications.     

Genetic engineering of fungal biocontrol agents to achieve greater efficacy against insect pests

Molecular biology methods have elucidated pathogenic processes in several fungal biocontrol agents including two of the most commonly applied entomopathogenic fungi, Metarhizium anisopliae and Beauveria bassiana. In this review, we describe how a combination of molecular techniques has: (1) identified and characterized genes involved in infection; (2) manipulated the genes of the pathogen to improve biocontrol performance; and (3) allowed expression of a neurotoxin from the scorpion Androctonus australis. The complete sequencing of four exemplar species of entomopathogenic fungi including B. bassiana and M. anisopliae will be completed in 2010. Coverage of these genomes will help determine the identity, origin, and evolution of traits needed for diverse lifestyles and host switching. Such knowledge combined with the precision and malleability of molecular techniques will allow design of multiple pathogens with different strategies to be used for different ecosystems and avoid the possibility of the host developing resistance.     

The impact of nutrition on spore yields for various fungal entomopathogens in liquid culture

Spore yields were measured for various fungal entomopathogens grown in six nutritionally different liquid media with low and high carbon concentrations (8 and 36 g l^–1, respectively) at carbon-to-nitrogen (C:N) ratios of 10:1, 30:1 and 50:1. Six fungi were tested: two Beauveria bassiana strains, three Paecilomyces fumosoroseus strains and one Metarhizium anisopliae strain. Spore yields were examined after 2, 4 or 7 days growth. In general, highest spore yields were obtained in media containing 36 g/l and a C:N ratio of 10:1. After 4 days growth, highest spore yields were measured in the three Paecilomyces isolates (6.9–9.7 × 10⁸ spores ml^–1). Spore production by the B. bassiana isolates was variable with one isolate producing high spore yields (12.2 × 10⁸ spores ml^–1) after 7 days growth. The M. anisopliae isolate produced low spore concentrations under all conditions tested. Using a commercial production protocol, a comparison of spore yields for the coffee berry borer P. fumosoroseus and a commercial B. bassiana isolate showed that highest spore concentrations (7.2 × 10⁸ spores ml^–1) were obtained with the P. fumosoroseus isolate 2-days post-inoculation. The ability of the P. fumosoroseus strain isolated from the coffee berry borer to rapidly produce high concentrations of spores prompted further testing to determine the desiccation tolerance of these spores. Desiccation studies showed that ca. 80% of the liquid culture produced P. fumosoroseus spores survived the air-drying process. The virulence of freshly produced, air-dried and freeze-dried coffee berry borer P. fumosoroseus blastospores preparations were tested against silverleaf whiteflies (Bemisia argentifolii). While all preparations infected and killed B. argentifolii, fresh and air-dried preparations were significantly more effective. These results suggest that screening potential fungal biopesticides for amenability to liquid culture spore production can aid in the identification of commercially viable isolates. In this study, P. fumosoroseus was shown to possess the production and stabilization attributes required for commercial development.     

Advances in fundamental and applied studies in China of fungal biocontrol agents for use against arthropod pests

Entomopathogenic fungi, such as Beauveria bassiana and Metarhizium anisopliae, are environmentally friendly biocontrol agents (BCAs) against various arthropod pests. We provide an overview to the past-decade advances in fungal BCA research and application in China. Since 1960s, fungal BCAs have been mass-produced for application and at present, thousands of tons of their formulations are annually applied to control forest, agricultural, greenhouse and grassland insect pests throughout the country. Apart from technical advances in mass production, formulation and application of fungal BCAs, basic studies on the genomics, molecular biology, genetic engineering and population genetics of fungal entomopathogens have rapidly progressed in the past few years in China. The completed genomic studies of M. anisopliae, Metarhizium acridum, B. bassiana and Cordyceps militaris provide profound insights into crucial gene functions, fungal pathogenesis, host–pathogen interactions and mechanisms involved in fungal sexuality. New knowledge gained from the basic studies has been applied to improve fungal virulence and stress tolerance for developing more efficacious and field-persistent mycoinsecticides by means of microbial biotechnology, such as genetic engineering. To alleviate environmental safety concerns, more efforts are needed to generate new data not only on the effects of engineered BCAs on target and non-target arthropods but also on their potential effects on gene flow and genetic recombination before field release.     

Targeting the click beetle Agriotes obscurus with entomopathogens as a concept for wireworm biocontrol

Applications of Metarhizium brunneum Petch (Ascomycota: Hypocreales: Clavicipitaceae) isolate LRC112 conidia caused high mortality to Agriotes obscurus L. (Coleoptera: Elateridae) click beetles in field trials. Banded conidiated rice (4.4 × 10¹⁴ conidia ha^?1) and conidia dust (5.0 × 10¹³ conidia ha^?1) resulted in 93.3 % ± 7.3 and 91.3 % ± 3.0 mortality after 18 days, while aqueous conidia suspension spray (5.0 × 10¹³ conidia ha^?1) with and without 80 g ha^?1 spinosad resulted in 68.2 % ± 17.7 and 52.6 % ± 17.4 mortality. Differences in results between 2012 and 2013 were attributed to rainfall, with pronounced effects in 2012 (rain beginning 35 h post treatment) and minimal effects in 2013 (rain beginning at 4 h). In another field experiment, beetles dosed with 1.49 × 10⁷ ± 5.08 × 10⁶ conidia per beetle retained 4.6 % of conidia after seven days while conidia viability on beetle bodies remained unchanged. The results inferred opportunities for controlling click beetles using fungal entomopathogens, and for horizontal transmission of the inoculum.     

Principles from community and metapopulation ecology: application to fungal entomopathogens

Fungal entomopathogens are often studied within the context of their use for biological control, yet these natural enemies are also excellent subjects for studies of ecological interactions. Here, we present selected principles from community ecology and discuss these in relation to fungal entomopathogens. We discuss the relevance of apparent competition, food web construction, intraguild predation and density-mediated and trait-mediated indirect effects. Although current knowledge of community interactions involving fungal entomopathogens are limited, fungal entomopathogens can be important, interactive members of communities and the activities of fungal entomopathogens should be evaluated in the context of ecological principles. We also discuss aspects of metapopulation ecology and the application of these principles to fungal entomopathogens. Knowledge of ecological interactions is crucial if we are to understand and predict the effects of fungal entomopathogens on host populations and understand the interactions among fungal entomopathogens and other organisms in the communities in which they occur.     

Challenges in modelling complexity of fungal entomopathogens in semi-natural populations of insects

The use of fungal entomopathogens as microbial control agents has driven studies into their ecology in crop ecosystems. Yet, there is still a lack of understanding of the ecology of these insect pathogens in semi-natural habitats and communities. We review the literature on prevalence of fungal entomopathogens in insect populations and highlight the difficulties in making such measurements. We then describe the theoretical host-pathogen models available to examine the role that fungal entomopathogens could play in regulating insect populations in semi-natural habitats, much of the inspiration for which has been drawn from managed systems, particularly forests. We further emphasise the need to consider the complexity, and particularly the heterogeneity, of semi-natural habitats within the context of theoretical models and as a framework for empirical studies. We acknowledge that fundamental gaps in understanding fungal entomopathogens from an ecological perspective coupled with a lack of empirical data to test theoretical predictions is impeding progress. There is an increasing need, especially under current rapid environmental change, to improve our understanding of the role of fungi in insect population dynamics beyond the context of forestry and agriculture.     

Endophytic fungal entomopathogens with activity against plant pathogens: ecology and evolution

Dual biological control, of both insect pests and plant pathogens, has been reported for the fungal entomopathogens, Beauveria bassiana (Bals.-Criv.) Vuill. (Ascomycota: Hypocreales) and Lecanicillium spp. (Ascomycota: Hypocreales). However, the primary mechanisms of plant disease suppression are different for these fungi. Beauveria spp. produce an array of bioactive metabolites, and have been reported to limit growth of fungal plant pathogens in vitro. In plant assays, B. bassiana has been reported to reduce diseases caused by soilborne plant pathogens, such as Pythium, Rhizoctonia, and Fusarium. Evidence has accumulated that B. bassiana can endophytically colonize a wide array of plant species, both monocots and dicots. B. bassiana also induced systemic resistance when endophytically colonized cotton seedlings were challenged with a bacterial plant pathogen on foliage. Species of Lecanicillium are known to reduce disease caused by powdery mildew as well as various rust fungi. Endophytic colonization has been reported for Lecanicillium spp., and it has been suggested that induced systemic resistance may be active against powdery mildew. However, mycoparasitism is the primary mechanism employed by Lecanicillium spp. against plant pathogens. Comparisons of Beauveria and Lecanicillium are made with Trichoderma, a fungus used for biological control of plant pathogens and insects. For T. harzianum Rifai (Ascomycota: Hypocreales), it has been shown that some fungal traits that are important for insect pathogenicity are also involved in biocontrol of phytopathogens.     

Inhibition of fungal spore germination by gramicidin S and its potential use as a biocontrol against fungal plant pathogens

Gramicidin S, as well as being sporicidal to Bacillus spores, also inhibits germination and emergence of fungal-like spores of Dictyostelium discoideum . The fungal plant pathogen Fusarium nivale is also inhibited and gramicidin S, therefore, is a sporicidal and antifungal antibiotic. Considering these findings the potential use of this antibiotic and its producer organism Bacillus brevis as a biocontrol is discussed.     

比较两种昆虫病原真菌中结构相似的ATP结合匣转运蛋白的抗氧化力与多药抗性

[目的]比较分析球孢白僵菌(Beauveria bassiana)BbT1和玫烟棒束孢(Isaria fumosorosea)IfT1两种结构相似的ATP结合匣转运蛋白的生物学功能.[方法]基于Bb2860野生株构建BbT1的敲除株和回补株,并将IFT1在BbT1敲除株中异源重组表达,比较各菌株的表型变化.[结果]与野生株、回补株及异源重组株相比,敲除株对20 - 40 mmol/L过氧化氢和2-8 mmoL/L甲萘醌氧化胁迫的抵抗力下降27％ -2.1倍,对多菌灵、伊曲康唑、菌核净、放线菌酮、乙嘧酚和4-硝基喹啉N-氧化物等不同类型化学药物的抗药性下降28％ -4.7倍,对斜纹夜蛾Spodoptera litura二龄幼虫的毒力下降20％左右,而野生株、回补株及异源重组株之间无任何表型的显著差异.[结论]BbT1和IfT1是结构相似且功能一致的转运蛋白,分别是两种生防真菌多药抗性的决定因子之一,因参与抗氧化反应而对毒力有所贡献.     

Occurrence and diversity of fungal entomopathogens in soils of low and high Arctic Greenland

Knowledge of the occurrence, distribution and diversity of pathogens of insects and arachnids (entomopathogens) in the Arctic is very limited. Climate change is expected to affect Arctic terrestrial arthropod communities and therefore also host–pathogen interactions, given that entomopathogens are present. We conducted a survey of fungal entomopathogens in soil samples collected at four localities in Greenland; two at low Arctic sites (Ritenbenk and Disko Island) and two at sites in the high Arctic (Zackenberg and Danmarkshavn). Fungi were isolated from soil samples using larvae of the insect species Galleria mellonella (Lepidoptera) and Tenebrio molitor (Coleoptera) as baits providing evidence that the fungal isolates were indeed entomopathogenic. Five fungal species (Ascomycota; Hypocreales) were found: Isaria fumosorosea Wize, Isaria farinosa (Holmsk.) Fr., Beauveria bassiana (Bals.) Vuill., Beauveria pseudobassiana Rehner and Humber and Tolypocladium inflatum W. Gams (syn.?=?T. niveum). I. farinosa was found at all four localities, while I. fumosorosea was detected in single samples at each of three localities including both high Arctic sites. Only the locality on Disko Island revealed B. bassiana, whereas B. pseudobassiana was isolated at the three other sites. T. inflatum was only found on Disko Island and only isolated with T. molitor as a bait insect. The results document that fungal entomopathogens are widely distributed in the soil environment in Greenland. Entomopathogens should therefore be included in future studies of arthropod ecology in the Arctic.     

Susceptibility to entomopathogens and modulation of basal immunity in two insect models at different temperatures

In this work, we analysed the efficacy of different commercial bio-insecticides (Steinernema feltiae, Steinernema carpocapsae, Heterorhabditis bacteriophora and Bacillus thuringiensis) by valuating the mortality induced on two insect models, Galleria mellonella (Lepidoptera) and Sarcophaga africa (Diptera) after exposure to different temperatures (10, 20 and 30 °C). Moreover, we investigated the effects of temperature on the basal humoral immunity of the two target insects; particularly, phenoloxidase (PO) and lysozyme activity. Our results show that G. mellonella is susceptible to all bio-insecticides at all the examined temperatures, except when infected at 10 °C with S. carpocapsae and at 30 °C with S. feltiae and B. thuringiensis. S. africa is more susceptible at 30 °C to all bioinsecticides; whereas, when infected at 10 and 20 °C, H. bacteriophora is the most efficient. Temperature modulates PO activity of both G. mellonella and S. africa, otherwise variations in lysozyme activity is observed only in G. mellonella. Except for a possible correlation between the increased lysozyme activity and the delayed Bt efficacy recorded on G. mellonella at 30 °C, a different resistance to bio-insecticides at different temperatures does not seem to be associated to variations of the host basal immunity, probably due to immunoevasive and immunodepressive strategies of these entomopathogens.     

Epicoccum nigrum for biocontrol agents in vitro of plant fungal pathogens

Epicoccum nigrum strains were evaluated in vitro as potential biological agents for control of the growth of Fusarium avenaceum, F. graminearum, F. oxysporum and Botrytis cinerea. Individual biotic effects of five strains of E. nigrum on the various fungi were determined using the biotic series method elaborated for fungi, on potato dextrose agar (PDA - Difco) medium. Our results show that E. nigrum strains limited the growth of all isolates of Fusarium spp., but not of those of Botrytis cinerea.     

Efficacy of commercially available predators,nematodes and fungal entomopathogens for augmentative control of Drosophila suzukii

The recent arrival of Drosophila suzukii, an invasive pest of soft‐skinned fruit with a wide host range, has resulted in increased production costs for growers and the need for additional insecticide applications each growing season. There are few effective organic insecticides for D. suzukii, and insecticide use in conventional farms may be disruptive to natural enemies, suggesting a need for effective biological control to combat D. suzukii. Commercially available natural enemies were evaluated for their potential use in augmentative releases, including: the predators Orius insidiosus and Dalotia coriaria; the entomopathogenic fungi Metarhizium anisopliae, Beauveria bassiana and Paecilomyces fumosoroseus; and the entomopathogenic nematodes Heterorhabditis bacteriophora, Steinernema feltiae and S. carpocapsae. This suite of natural enemies was chosen to target D. suzukii adults as well as larvae in hanging or dropped fruit. Of the cultured fungal strains tested, only M. anisopliae significantly decreased D. suzukii survival, but it had low residual activity and no effect on D. suzukii fecundity. O. insidiosus decreased D. suzukii survival in simple laboratory arenas but not on potted blueberries or bagged blueberry branches outdoors. D. coriaria did not decrease D. suzukii survival in infested blueberries in simple laboratory arenas. The nematodes tested showed low infection rates and were not able to affect D. suzukii survival. Although this suite of natural enemies showed limited ability to suppress D. suzukii under the tested conditions, these and related natural enemies are present as part of the endemic natural enemy community in agricultural fields, where they may contribute to D. suzukii suppression.     

Rhizobia: a potential biocontrol agent for soilborne fungal pathogens

Rhizobia are a group of organisms that are well known for their ability to colonize root surfaces and form symbiotic associations with legume plants. They not only play a major role in biological nitrogen fixation but also improve plant growth and reduce disease incidence in various crops. Rhizobia are known to control the growth of many soilborne plant pathogenic fungi belonging to different genera like Fusarium, Rhizoctonia, Sclerotium, and Macrophomina. Antagonistic activity of rhizobia is mainly attributed to production of antibiotics, hydrocyanic acid (HCN), mycolytic enzymes, and siderophore under iron limiting conditions. Rhizobia are also reported to induce systemic resistance and enhance expression of plant defense-related genes, which effectively immunize the plants against pathogens. Seed bacterization with appropriate rhizobial strain leads to elicitation and accumulation of phenolic compounds, isoflavonoid phytoalexins, and activation of enzymes like L-phenylalanine ammonia lyase (PAL), chalcone synthase (CHS), peroxidase (POX), polyphenol oxidase (PPO), and others involved in phenylpropanoid and isoflavonoid pathways. Development of Rhizobium inoculants with dual attributes of nitrogen fixation and antagonism against phytopathogens can contribute to increased plant growth and productivity. This compilation aims to bring together the available information on the biocontrol facet of rhizobia and identify research gaps and effective strategies for future research in this area.     

Deep space and hidden depths: understanding the evolution and ecology of fungal entomopathogens

Entomopathogens are important natural enemies of many insect and mite species and as such have been recognised as providing an important ecosystem service. Indeed, fungal entomopathogens have been widely investigated as biological control agents of pest insects in attempts to improve the sustainability of crop protection. However, even though our understanding of the ecology of fungal entomopathogens has vastly increased since the early 1800s, we still require in-depth ecological research that can expand our scientific horizons in a manner that facilitates widespread adoption of these organisms as efficient biological control agents. Fungal entomopathogens have evolved some intricate interactions with arthropods, plants and other microorganisms. The full importance and complexity of these relationships is only just becoming apparent. It is important to shift our thinking from conventional biological control, to an understanding of an as yet unknown “deep space”. The use of molecular techniques and phylogenetic analyses have helped us move in this direction, and have provided important insights on fungal relationships. Nevertheless, new techniques such as the PhyloChip and pyrosequencing might help us see beyond the familiar fields, into areas that could help us forge a new understanding of the ecology of fungal entomopathogens.     

Toxicity of non-ionic surfactants and interactions with fungal entomopathogens toward Bemisia tabaci biotype B

Several non-ionic surfactants can be used to enhance insecticidal performance of various bio-based products and agrochemicals. In this study, we describe the toxicity of four commercial non-ionic surfactants against immature Bemisia tabaci (Gennadius) biotype B whiteflies along with their spreading ability. Results revealed that trisiloxane-based surfactants (Break-thru^® and Silwet^® L-77) exhibited the highest toxicity to 1st–2nd (early) and 3rd–4th (late) instar nymphs as well as the greatest wetting performance. We also investigated whether sub-lethal concentrations of Break-thru^® and Silwet^® L-77 improved the insecticidal efficacy of the fungal entomopathogens Beauveria bassiana (Balsamo) Vuillemin (strain CG1229) and Isaria fumosorosea Wise (strain CG1228) conidial suspensions against whiteflies. Germination of hydrophobic conidia of both isolates were unaffected by these surfactants over the range 100–1000 ppm. The combinations of both fungi with trisiloxane carriers significantly increased nymphal mortality with mostly additive and synergistic effects on early and late nymphs, respectively. In screenhouse trials, both fungi (10⁷ conidia ml^?1) mixed with 200 ppm of Silwet^® L-77 significantly improved effectiveness against early nymphs (72–74 % mortality) compared with controls treated with water and Silwet^® L-77 alone. In addition, reduced volume rates of I. fumosorosea in Silwet^® L-77 (equivalent to 100 l ha^?1) were at least as effective against late nymphs as higher volume rates (200 l ha^?1) at equivalent conidial doses. Our findings underline the compatibility and enhanced activity of silicon-based surfactants with B. bassiana and I. fumosorosea for use in integrated whitefly management programs.     

Screening for candidate bacterial biocontrol agents against soilborne fungal plant pathogens

Previous studies relating root systems and drought tolerance in oil palm focused mainly on biomass. Yet, total root length (TRL), total root surface area (TRS), and root distribution in the soil better determine water uptake. These morphological traits were studied on 3 oil palm genotypes displaying a contrasting drought tolerance. A new concept of potential root water extraction ratio (PRER) was developed using measured half-distances between roots and some assumptions about the distance of water migration from soil to root. PRER was determined in conjunction with soil moisture extraction efficiency (SMEE). The presumed tolerant genotype (T) had higher TRL, TRS and PRER than the susceptible genotype (S), whilst the performance of the control genotype (I) was intermediate. Surprisingly, during a period of moderate water deficit, T had a lower SMEE than S, which was interpreted successfully with PRER, as the result of a better access to a large volume of soil and of a slower drying out of the soil around the roots. PRER appears as a helpful indicator for comparing or ranking genotypes, and for addressing better the complexity of the genetic variability of drought tolerance.