绿僵菌生物农药的研制与应用

绿僵菌生物农药的生产目前普遍采用液固双相发酵工艺,其固体载体多为大米及稻壳等材料,国内外的研究多集中于培养参数的优化与选择,如培养基成分、温度、光照、pH值、含水量及接种量等。绿僵菌制剂的剂型主要有粉剂、可湿性粉剂、干菌丝和油剂,其助剂的成分及配比是当前的研究热点。在应用方面,全球已登记的绿僵菌商品有17个,应用效果较好,防治对象集中于直翅目的蝗虫类、蜚蠊目、同翅目的蚜虫、粉虱、叶蝉类以及鞘翅目的蛴螬等害虫。本文从工艺研究、剂型研发、商品化及应用等角度对绿僵菌生物农药现状进行综述,重点介绍近年来的研究进展和新兴技术。     

绿僵菌治蝗研究进展

介绍了目前对防治多种蝗虫效果较好的虫生真菌--绿僵菌的种类、株系,以及绿僵菌对蝗虫的致病机理、 对环境的安全性等,并讨论了国内外应用绿僵菌防治蝗虫的田间试验效果、剂型研制、产业化和推广应用前景。     

白僵菌和绿僵菌作为防控红火蚁生物农药的潜力及前景

红火蚁SolenopsisinvictaBuren已从南美洲入侵到北美洲、亚洲多个国家和一些太平洋岛国,在我国扩散至12个省区500多个市县,严重为害农林业生产,威胁人居安全和生态平衡。为了实现对红火蚁的可持续治理,需要引入和加强生物防治的作用。本文概述了昆虫病原真菌白僵菌、绿僵菌作为生物杀虫剂的总体现状,以及针对红火蚁研发的详细进展,包括优良菌株选育、规模化生产、产品剂型、登记注册及防控应用等,分析了白僵菌、绿僵菌制剂防控红火蚁的优势和尚需解决的问题,讨论和展望了真菌杀虫剂防控红火蚁的前景。     

巴西批准5种生物农药用于有机作物种植

巴西于今年5月批准了5种生物农药用于防治有机作物种植中不同的靶标害虫。其中,生物杀虫剂绿僵菌（metarhizium anisopliae）IBCB425可用于防治沫蝉及牧草沫蝉。该生物农药有可湿性粉剂和颗粒剂2种剂型,并对甘蔗沫蝉及牧草沫蝉有一定的防效。     

金龟子绿僵菌CQMa128新制剂对花生蛴螬的田间防控效果

为明确金龟子绿僵菌CQMa128微粒剂和乳粉剂对花生蛴螬的田间防治效果,设计随机区组小区试验,采用绿僵菌微粒剂在花生播种期防治上一年的3龄蛴螬老熟幼虫;采用绿僵菌乳粉剂在花生幼果期防治当年的1～2龄蛴螬.同期调查绿僵菌新制剂的施用对蛴螬主要天敌臀钩土蜂的影响.金龟子绿僵菌CQMa128微粒剂(最适剂量375 kg/hm2)施药后30～40 d,平均校正防效为75.58％～80.03％;金龟子绿僵菌CQMa128乳粉剂(最适剂量7.5 kg/hm2)施药后15～20 d,平均校正防效为77.59％～86.46％.花生果荚收获期平均保果率达85.76％～87.13％,花生果荚鲜重平均增产量达832.5～916.4kg/hm2,控害保产效果显著.且绿僵菌新制剂的施用对臀钩土蜂的存活未见有明显不良影响.表明金龟子绿僵菌CQMa128新制剂能够时田间花生蛴螬进行持续有效的防控.     

金龟子绿僵菌CQMa421农药及应用情况

金龟子绿僵菌Metarhizium anisopliae是一种重要的生防真菌,具有安全、易在害虫种群中流行、环境友好、不易产生抗药性等优点,已在中国、美国、欧盟等多个国家或地区登记和应用。本文对我国广谱金龟子绿僵菌CQMa421农药的菌株选育、产品登记、应用技术及产品应用情况等方面进行了概述。     

绿僵菌CQMa128乳粉剂对蛴螬时间-剂量-死亡率模型分析

用拌土法测定了金龟子绿僵菌Metarhizium anisopliae菌株CQMa128乳粉剂对蛴螬3龄幼虫的毒力。结果表明：土壤含有1×106~1×108孢子/g绿僵菌乳粉剂时,蛴螬的校正死亡率随着绿僵菌孢子浓度的增加和作用时间的延长而增大,土壤含有5×107孢子/g绿僵菌乳粉剂时蛴螬的校正死亡率达85.2%。运用时间-剂量-死亡率模型对生测数据进行拟合,通过Hos-mer-Lemeshow拟合异质性检验,并估计了该制剂对蛴螬的致死剂量与致死时间。随着接种天数的增加,相应的LD50和LD90随之降低,剂量效应逐渐增强。土壤中含有3×107、5×107和1×108孢子/g绿僵菌乳粉剂处理的LT50值分别为10.18、8.42、6.79d。当土壤含有1×108孢子/g绿僵菌乳粉剂时,蛴螬死亡率可达90%以上,LT90为10.50d。该绿僵菌制剂在蛴螬的生物防治中具较强的应用潜力。     

不同温度下绿僵菌对东亚飞蝗3龄蝗蝻的致病力影响

采用体表接菌法测定了商品生物农药金龟子绿僵菌油悬浮剂PD20080671在15、20、25、30、35和40℃等6个不同温度条件下对东亚飞蝗3龄蝗蝻的致病力的影响。结果表明：东亚飞蝗3龄蝗蝻的累积死亡率在接菌后19 d内均随天数的增加而升高。35℃的累积死亡率最高（82.50%）,显著高于其他处理;40℃的累积死亡率最低,为55.00%。死亡个体的湿度培养结果显示绿僵菌在高温条件下萌发的时间较短,40℃为4.26 d,15℃为19.36 d。结果表明环境温度低于15℃或高于35℃都不利于绿僵菌杀死蝗虫。因此,在田间施用金龟子绿僵菌防治东亚飞蝗时应尽量避免高温天气,建议在上午低温时进行防治。     

绿僵菌对云南松切梢小蠹的致病性

利用3个绿僵菌菌株2060-d、X-m、2014分生孢子分别制成液剂、油剂和粉剂通过室内和林间试验,3个绿僵菌菌株对云南松切梢小蠹成虫都具有较高的致病力。室内校正死亡率77.1%~97.9%,死虫僵虫率达100%;用2060-d菌株分生孢子制成不同的剂型在林间防治切梢小蠹成虫,试验结果,成虫死亡率依次为油剂(73.3%)>液剂(70.0%)>粉剂(20.0%),死虫僵虫率分别为67.0%、65.0%和20.0%。上述结果表明,绿僵菌对云南松切梢小蠹有一定的致病力,可以将绿僵菌分生孢子制成油剂进行低量喷雾防治。     

不同浓度增效剂对金龟子绿僵菌、黄绿绿僵菌和球孢白僵菌产孢的影响

生防菌作为一种重要的生物杀虫剂,被开发成多种制剂应用于农林害虫的防治;微生物制剂的生产成本高、耗时长,极大影响其生产效率。本试验通过液固两相发酵法筛选出提高金龟子绿僵菌Metarhizium anisopliae、黄绿绿僵菌Metarhizium flavoviride和球孢白僵菌Beauveria bassiana产孢量的最优增效剂浓度。结果表明0.1%浓度增效剂对金龟子绿僵菌的促产孢作用达最大化,浓度达4.48×10⁹孢子/g;黄绿绿僵菌的最适增效剂浓度为0.05%,浓度达2.75×10⁹孢子/g;当增效剂浓度小于0.1%时利于黄绿绿僵菌的产孢,增效剂高于0.1%时抑制产孢;而球孢白僵菌的产孢量与增效剂浓度正相关,最大浓度达7.41×10⁹孢子/g。在真菌液体发酵过程中添加产孢增效剂,有效提高了真菌产孢量,缩短了真菌培养周期,为提高真菌制剂生产效率、节约成本提供理论支持。     

Genetically engineering better fungal biopesticides

Microbial insect pathogens offer an alternative means of pest control with the potential to wean us off our heavy reliance on chemical pesticides. Insect pathogenic fungi play an important natural role in controlling disease vectors and agricultural pests. Most commercial products employ Ascomycetes in the genera Metarhizium and Beauveria. However, their utilization has been limited by inconsistent field results as a consequence of sensitivity to abiotic stresses and naturally low virulence. Other naturally occurring biocontrol agents also face these hurdles to successful application, but the availability of complete genomes and recombinant DNA technologies have facilitated design of multiple fungal pathogens with enhanced virulence and stress resistance. Many natural and synthetic genes have been inserted into entomopathogen genomes. Some of the biggest gains in virulence have been obtained using genes encoding neurotoxic peptides, peptides that manipulate host physiology and proteases and chitinases that degrade the insect cuticle. Prokaryotes, particularly extremophiles, are useful sources of genes for improving entomopathogen resistance to ultraviolet (UV) radiation. These biological insecticides are environmentally friendly and cost‐effective insect pest control options. © 2017 Society of Chemical Industry     

Biological control in Hungary: present situation and possibilities1

Biological control has already achieved certain results in Hungary, especially against soil-borne fungi and glasshouse pests. Nevertheless, experience has shown that locally found biological agents, however effective, could not be developed into registered, ready-for-sale plant protection products because of the lack of companies willing to invest. Biological control in Hungary is therefore limited to the conservation of natural antagonistic flora and fauna and to the application of imported biopesticides and bioagents, which are subject in Hungary to a registration procedure similar to that for chemical plant protection products. Current work includes the development of an anti-nematode product from a local strain of Arthrobotrys oligospora, application techniques for a local mirid Dicyphus hyalinipennis against insect pests, studies with native and imported strains of the nematodes Steinernema and Heterorhabditis against soil-borne insect pests, and introduction techniques for the chrysomelid Zygogramma sutularis against the weed Ambrosia elatior.     

Susceptibility of larvae and adults of Monochamus galloprovincialis to entomopathogenic fungi under controlled conditions

Six entomopathogenic fungal isolates were tested under controlled conditions, as biological control agents against Monochamus galloprovincialis (Coleoptera: Cerambycidae) (Olivier), the vector of the pine wood nematode Bursaphelenchus xylophilus (Nematoda: Parasitaphelenchidae) (Steiner and Buhrer). The fungi Beauveria bassiana (Bals.-Criv.) Vuill, Metarhizium anisopliae (Metchnikoff) Sorokin, Metarhizium sp. and Fusarium sp. were tested against larvae and adults of the insect, by comparing two exposure techniques: continuous contact and spraying. Regarding the larvae, only Fusarium sp. and Metarhizium sp. killed more than 40% of the individuals, while B. bassiana and M. anisopliae were found to kill 50% of the adults (LT₅₀) within five days. Exposure to B. bassiana reduced adult longevity by 19 days, when compared to untreated beetles, and had a direct efficacy of 46%. Continuous contact with the fungi proved to be more effective than spraying. A larval disinfection protocol was developed to eliminate other fungal isolates from the wood-collected tested individuals. The implications of the results and possible applications of selected strains as bio-control agents against M. galloprovincialis are discussed.     

Regulation and use of biological control agents in Hungary

A wide range of biological control agents (BCAs) have been authorized and used in Hungary for the control of pests (including pathogens). BCAs are key elements of both integrated pest management and organic farming for different crops. Authorization of BCAs depends on the type of ‘active substance’. Micro‐organisms can be authorized under the EU Regulation (EC) No. 1107/2009 as plant protection products (PPP), while macro‐organisms are not within its scope. Plant protection tools (traps, mating disruption) are registered at a national level in Hungary, although mass traps and mating disruption pheromones are considered to be PPP. Plant extracts can be authorized as PPP, but some of them can be approved as a ‘basic substance’ and in this case they do not need further authorization. In Hungary, the vast majority of macro‐organisms used as BCAs have been mainly used in greenhouses and polytunnels for the control of thrips, whiteflies, aphids, leaf mining flies, phytophagous mites and lepidopteran pests in vegetable and ornamental crops. Recently a non‐indigenous chalcid wasp, Torymus sinensis, has been successfully introduced for the control of the oriental chestnut gall wasp, Dryocosmus kuriphilus, on chestnut (Castanea sativa) on different sites.     

In vitro and in vivo antifungal activities of decursin and decursinol angelate isolated from Angelica gigas against Magnaporthe oryzae, the causal agent of rice blast

Blast is considered the most important fungal disease of rice due to its wide distribution and destructiveness under favorable conditions. Development of new effective and environmentally benign agents against the causal pathogen, Magnaporthe oryzae, is of great interest. In the course of a search for natural antifungal compounds in medicinal plants, we found that the methanol extract of Angelica gigas roots showed a potent control efficacy against rice blast caused by M. oryzae. We isolated antifungal coumarins from the extract, and they were identified as decursin and decursinol angelate. Antifungal activities of these compounds, along with kasugamycin, were tested on M. oryzae in vivo and in vitro. In an in vivo assay, the three compounds effectively suppressed the development of rice blast at concentrations more than 100 μg/mL. Coumarins showed relatively weak inhibitory effect on fungal mycelial growth when compared to kasugamycin. However, they strongly inhibited M. oryzae spore germination, which was not observed in kasugamycin treatments. This is the first report demonstrating that decursinol angelate can provide control against rice blast and that the two coumarins inhibit M. oryzae spore germination. In addition, the wettable powder formulation of the crude extract of A. gigas prohibited the development of blast symptoms on rice plants more effectively than liquid concentrate formulation of kasugamin, a commercial fungicide. Based on our study, we propose that coumarin compounds as well as the A. gigas root crude extract can be used as natural, benign fungicides for controlling rice blast.     

Thermal behaviour and biological activity of pyrethroids in smoke-generating formulations

Smoke-generating insecticide formulations are nowadays very important for the control of Chagas' disease vectors. In this work, the thermal decomposition of different pyrethroid insecticides in smoke-generating formulations has been studied. Their recovery from the smoke has been determined, and the effect on this of the addition of foaming agents such as cyanoguanidine (CNG) or azodicarbonamide (ADC), or antioxidant agents such as butylated hydroxytoluene (BHT) or butylated hydroxyanisole (BHA) has been assessed. In each case the best smoke-generating formulation was established. Calorimetric studies were done to justify the behaviour of the smoke-generating mixtures. The isomerization process for different pyrethroids was also studied, to establish the influence of the foaming agents in inhibiting isomerization to less active pyrethroids. Smoke-generating mixtures containing β-cypermethrin and cis-permethrin as insecticides and CNG or ADC as foaming agents were evaluated for their insecticidal activity against nymphs I of Triatoma infestans, vector of Chagas' disease. The best effect was found with β-cypermethrin formulated with ADC, with LT₅₀ values lower than 5 min. © 1999 Society of Chemical Industry     

Evaluation of the entomopathogenic fungiMetarhizium anisopliae andBeauveria bassiana against the red palm weevilRhynchophorus ferrugineus

The red palm weevil (RPW,Rhynchophorus ferrugineus (Olivier) (Coleoptera: Curculionidae) is one of the most severe pests of various palm species, including date palms. While examining the susceptibility of RPW to two entomopathogenic fungi,Metarhizium anisopliae andBeauveria bassiana, strains of the former were found to be more virulent than those of the latter, achieving 100% larval mortality within 6–7 days. The most virulent strains ofM. anisopliae were then tested on RPW eggs and adults. Incubation in a substrate treated withM. anisopliae spores increased egg mortality and reduced their hatchability. The total percentage mortality of eggs and hatched larvae was 80–82%, compared with 34% in the controls. RPW adults were challenged with two types of fungal formulation: dry powder and aqueous suspension. Cumulative adult mortality of 100% was achieved in 2–3 weeks for the dry rice-based formulation and in 4–5 weeks for the spore suspension. As a result of decreased longevity, treated females had a shorter oviposition period and three times lower fertility than the controls. Possible strategies for fungus application are discussed in the light of the high susceptibility of eggs and larvae to fungal infection. http://www.phytoparasitica.org posting July 19, 2006.     

Integrated crop management, the basis for environment friendly crop protection of potatoes

Relatively large amounts of pesticides are needed to control diseases and pests in modern, intensive potato production. Integrated crop management offers a way of reducing the need for pesticides. It aims to reduce costs and improve the quality of the product and of the production methods, while maintaining soil fertility and the quality of the environment. The components of integrated crop management are described. Prevention of diseases and pests has high priority. If diseases or pests are present, non-chemical control methods are preferred and chemical control is based on economic criteria and the monitoring of the soil and crops. The contribution of integrated crop management to the control of two important fungal diseases, late blight (Phytophthora infestans), and stem canker (Rhizoctonia solani) and of one pest, potato cyst nematode (Globodera spp..) is described. The prospects for further development of integrated crop protection are discussed.     

Bt Transgenic Crops: Risks and Benefits

Bt crops, predominantly maize and cotton hybrids, transgenically expressing cry genes derived from Bacillus thuringiensis, were planted on approximately 14 million hectares (worldwide) in 1999. Preliminary reports suggest that in most cases pesticide use was reduced, and in some situations there were significant increases in yields and profits. However, assemblages of secondary pests – such as aphids, plant bugs and thrips – also exist in Bt crops, and although the overall need for scouting and chemical control is reduced in Bt crops, there may be a requirement for additional, conventionally applied chemicals to control such non-target pests.Naturally-occurring Bt toxins with activity against a wide variety of pest species have been discovered and are thus potentially available for engineering into Bt crops to control a broader spectrum of pests than are currently targeted. New Bt crops and second-generation products incorporating an expanding range of Cry toxins and other arthropod targeted genes are in development and could become available for introduction to the market within the next few years.Insecticide resistance management (IRM) strategies for Bt crops are reviewed in the context of studies on selection pressures and the potential for resistance development in target populations. The so-called, high dose strategy, combined with the use of refuges, is widely agreed to be the best technical approach for managing resistance, and evidence is accumulating that separate refuges are more effective at conserving pest susceptibility than mixed refuges. A widespread consensus on the necessity for such measures, and an appreciation of the importance of multi-tactical approaches, has developed. Monitoring programmes, protocols and studies relevant to detecting the early development of resistance to Bt Cry toxins are described.Field monitoring of non-target entomofauna has not revealed significant differences in the abundance or biodiversity of beneficial insects associated with Bt maize. Indeed, laboratory studies of effects on parasitoids suggest that Bt plants may even have an environmental advantage over broad spectrum pesticides. However, more complex, multi-trophic, long-term experiments are needed to thoroughly assess the compatibility of Bt crops with non-target invertebrates and to define the complex relationship between IRM, target species and their natural enemy assemblages. Studies on the effects of transgenically-expressed Cry toxins on non-target insects, and their persistence in soil and on leaves, is reviewed. It is suggested that there is currently no generally agreed framework, or methodology, within which ad hoc experimental results can be accommodated, and each crop-transgene combination has to be assessed on a case-by-case basis. Studies proposing a conceptual approach to evaluating risks associated with Bt crops are highlighted and potential benefits and hazards are reviewed.     

Potential of biopesticides in agriculture

All living organisms are subject to predation, parasitism or competition from other organisms. The study of these interactions has led to the identification of many potential opportunities for the use of living organisms as biopesticides to protect agricultural crops against insect pests, fungal, bacterial and viral diseases, weeds, nematodes and mollusc pests. A range of biopesticide products (including as active agents bacteria, fungi, nematodes, protozoa, viruses and beneficial insects) are now available commercially for control of insect pests, fungal and bacterial diseases and weeds. However, world biopesticide sales in 1990 were estimated to be $120 million, representing less than 0–5% of the world agrochemical market. Over 90% of biopesticide sales are represented by a single product type, containing Bacillus thuringiensis Berl., for control of insect pests. Nevertheless, biopesticide sales are estimated to be increasing at 10–25% per annum whilst the world agrochemical market is static or even shrinking. There has been a significant renewal of commercial interest in biopesticides as evidenced by the substantial number of alliances forged between major agrochemical companies and biotechnology companies which allow these major companies access to marketing rights to novel biopesticides. This paper reviews the current commercial status of biopesticides and discusses the technical and commercial constraints which have impeded development of biopesticides in the past. Novel developments in R&D, which may enable some of these constraints to be overcome, are examined by reference to a number of specific examples (some of which arise from the author's own experience in a biotechnology company). The future prospects for biopesticides are discussed in the light of technical advances and commercial and regulatory requirements.