Prospects for the development of a genetically engineered baculovirus insecticide

Baculoviruses are insect-specific pathogens which have been employed for the natural control of insect pests for many years. They present an alternative to the use of chemical agents for crop protection. Their restricted host range enables particular pest species to be targeted, and this prevents any loss of beneficial insects from a treated area. However, their large-scale development as insect control agents has been limited because they take longer to kill the host following infection than rapidly-acting chemical insecticides. The aim of genetic modification of baculoviruses is to increase their speed of action, while preserving their beneficial characteristics, so creating a fast-acting, environmentally friendly insecticide. The prospects for achieving this are discussed in this paper, with consideration of scientific progress and the influence of economic and legislative factors.     

杆状病毒在昆虫中的持续感染

杆状病毒(Baculovirus)是一类特异性感染节肢动物的环状双链DNA病毒,是野外控制害虫种群的重要生物因子,并已被开发为一种生物杀虫剂加以应用。杆状病毒感染昆虫宿主并不一定导致昆虫死亡,其持续感染(per-sistent infection)在昆虫种群中普遍存在,且在某些刺激条件下,持续感染可被激活为增殖性感染并引发病毒流行病爆发。因此,杆状病毒持续感染对昆虫种群动力学以及病毒流行病学的研究具有重要意义。     

Natural products in agriculture—a view from the industry

The paper discusses the use of natural products and biological control agents in crop protection from an industrial viewpoint. The criteria which must be satisfied are noted. Examples are given from the genetic engineering of baculoviruses and proteins. The final section considers the utility of natural products as a source of leads for conventional agrochemicals, and the screens needed. © 1998 SCI.     

改造病毒基因组增强病毒杀虫剂杀虫效率研究进展

由于基因工程技术的发展和安全性研究的深入,以重组杆状病毒为主的重组昆虫病毒杀虫剂的应用正面临着突破。文章综述了通过修饰或删除昆虫病毒某些基因、插入控制宿主发育、代谢激素或酶的基因、删除、修饰昆虫病毒基因组中特定的基因以扩大宿主域、应用RNA干扰技术提高昆虫病毒杀虫效率等构建重组昆虫病毒杀虫剂的技术路线,展望重组病毒杀虫剂的发展趋势。     

Genetically engineered viral insecticides–A progress report 1986 1989

Among the viruses that are pathogenic for insect species, baculoviruses have been shown to be useful as insecticides for pest control. In some cases they have been used as cost-effective and environmentally acceptable alternatives to chemical insecticides. However, because viruses need to be ingested and replicate extensively in their host before they kill it, baculovirus insecticides are much slower than chemicals or other reagents that kill insects either on contact or shortly after ingestion. The objective of the programme of genetic engineering of baculovirus insecticides is to improve their speed of action while maintaining their host specificity and other attributes that make them desirable alternatives to chemical pesticides. Since 1986 four field releases have been undertaken involving genetically engineered baculovirus insecticides. The first release used a genetically marked Autographa californica nuclear polyhedrosis virus (AcNPV). The study began in 1986 and was terminated in 1987. The results demonstrated that an innocuous piece of DNA, appropriately positioned in the AcNPV genome, was an effective means to tag the virus without affecting its phenotype, allowing it to be identified in bioassays of plant and soil samples. The second release, in 1987, involved a genetically marked virus from which the gene coding for the protective polyhedrin protein of the virus had been removed. The field data obtained with this virus showed that it did not persist in the environment, neither in soil, nor on vegetation, nor in the corpses of caterpillars. The third and fourth releases were undertaken in 1988. For one of these studies the marked, polyhedrin-negative virus was again used. In the other study a polyhedrin-negative virus that contained a junk' (/?-galactosidase) gene was employed.     

Scorpion neurotoxins: structure/function relationships and application in agriculture

Continued use of non‐specific chemical insecticides poses potential risks to the environment and to human health resulting from non‐target toxicity and increased insect resistance to these agents. Scorpions produce anti‐insect selective polypeptide toxins that bind to and modulate voltage‐sensitive ion channels in excitable tissues, thus offering alternative, environmentally safer means for insect pest control. Despite this potential, little is known about their structural elements dictating anti‐insect preference, which may be useful for the design of selective insecticides. We used a bacterial system for expression and genetic dissection of two pharmacologically distinct scorpion toxins: alpha and excitatory. By exploiting a multi‐disciplinary approach consisting of mutagenesis, protein chemistry, electrophysiology, binding and toxicity assays, and structural studies, we elucidated the bioactive surface of two anti‐insect toxins, LqhαIT and Bj‐xtrIT. In both polypeptides the bioactive surface is composed of residues surrounding the C‐terminal region. In addition, a direct, immediate approach in using the toxin genes was demonstrated by engineering baculoviruses with cDNAs encoding LqhIT2 (depressant toxin), and LqhIT1 (excitatory toxin) resulting in viral vectors with significantly improved insecticidal efficacy. © 2000 Society of Chemical Industry     

Genetically engineered viral insecticides: New insecticides with improved phenotypes

Baculoviruses are natural pathogens of insects which have been used as biopesticides. In contrast to many chemical agents, baculoviruses affect only a limited number of insects and so can be used to target particular insect species. Unfortunately, unless the host receives a very high virus dose, the insect continues to feed and causes damage to crops, because the virus takes several days to kill it. This lag is unacceptable in the protection of many crops, especially where cosmetic damage seriously reduces the value of a crop (e.g. fruit). Strategies have been devised recently to circumvent this problem. The baculovirus genome has been modified, using genetic engineering techniques, to incorporate foreign genes encoding insect-specific toxins, or hormones or enzymes. Expression of some of these genes in the virus-infected host insect has been shown to reduce both the feeding damage to crop plants and the time taken to kill the insect pest. The current status of this developing field is described, together with an assessment of the possible risks involved in using such genetically modified agents in the environment.     

Development of the Multi-nucleocapsid Nucleopolyhedroviruses (MNPVs) Infectious to Loopers (Lepidoptera: Noctuidae: Plusiinae) as Microbial Control Agents

A number of the members of the Noctuidae (subfamily Plusiinae) are polyphagous insects that infest a variety of economically important crops world-wide. These hosts include but are not limited to cotton, vegetable and field crops and also such specialty crops as mint. Many of these crops require multiple applications of chemical pesticides to protect them from various loopers and other economic pests. One proposed alternative to the use of chemical pesticides to protect these crops is the development of insect-specific viruses of the family Baculoviridae. Hundreds of these viruses of the genus Nucleopolyhedrovirus have been isolated, many from economically important Lepidoptera. The advent of the discovery that some of these viruses may have a broad host range increased the interest in the development of them as microbial agents throughout the world. The broad host range (over 30 species in some cases) provided the possibility to control not only one species, but a complex of lepidopterous pests infesting specific or multiple crops. From the 1960s to the present, extensive basic and applied research has been conducted on the multi-nucleocapsid (MNPV) forms within which a broad host range appears to be characteristic. Of these, the virus isolated from the alfalfa looper, Autographa californica (Speyer) (AcMNPV) has been the most studied, particularly as related to microbial control. Two of the viruses isolated from loopers have been registered in the USA. Although efficacy has been demonstrated for loopers, there is a paucity of data on the other important species that may simultaneously infest looper hosts. Because of their relatively slow action, the viruses will probably be used in integrated programs with chemical pesticides applied during critical periods of high population pressure. Further research needs to be conducted to establish efficacy for other hosts, improved production methods either in vivo or in vitro, increased field persistence as related to timing and frequency of applications, and finally the utility of genetically engineered MNPVs as microbial pesticides. The potential of these organisms has been demonstrated and new developments are being made to increase the overall efficiency and economics of their use. These improvements can only increase interest in use of the baculoviruses in pest management systems for loopers and other pests.     

Baculoviruses as vectors for foreign gene expression in insect cells

Recombinant DNA technology has enabled the expression of a large number of eukaryotic genes in vitro. There has been growing interest in recent years in the development of baculovirus expression vectors as an easily manipulated gene expression system. Both virus and insect cell culture are relatively easy to handle, and biologically active proteins have been produced abundantly from a variety of eukaryotic, prokaryotic and viral genes. This review describes the molecular biology of baculoviruses and some of the current applications of the baculovirus expression system in insect cells. Specific emphasis is placed on those features relevant to the use of this system in pesticide research.     

Persistence of naturally occurring and genetically modified Choristoneura fumiferana nucleopolyhedroviruses in outdoor aquatic microcosms

BACKGROUND: To assess the persistence of genetically modified and naturally occurring baculoviruses in an aquatic environment, replicate (three) outdoor, aquatic microcosms were spiked with spruce budworm viruses [Ireland strain of Choristoneura fumiferana multiple nucleopolyhedrovirus (CfMNPV) and the recombinant CfMNPVegt(-)/lacZ(+)] at a rate of 1.86 x 10(10) occlusion bodies (OBs) m(-2) of surface area. The presence of virus in water samples collected at various times after inoculation was determined by PCR amplification of baculoviral DNA extracted from OBs.RESULTS: Although UV radiation rapidly degrades baculoviruses under natural conditions, both viruses persisted above the level of detection (>100 OBs 450 microL(-1) of natural pond water) for at least 1 year post-inoculation, with little difference between the viruses in their patterns of persistence.CONCLUSION: The present microcosm study suggests that occlusion bodies of baculoviruses can persist in the flocculent layer of natural ponds. On disturbance, OBs could re-enter the main water column and thus be available for transport to new locations. Implications for environmental risk assessment are discussed. (c) 2008 Her Majesty the Queen in Right of Canada. Canadian Forestry Service. Published by John Wiley & Ltd.     

Can a chitin‐synthesis‐inhibiting turfgrass fungicide enhance black cutworm susceptibility to a baculovirus?

BACKGROUND: Developmental resistance, i.e. reduced virulence and speed of kill of late instars, is a limiting factor in the use of baculoviruses for caterpillar control. Agrotis ipsilon multicapsid nucleopolyhedrovirus (AgipMNPV) is highly infective to young black cutworms, Agrotis ipsilon, but too slow‐acting against late instars for effective curative control on golf courses or sports fields. Chitin‐synthesis‐inhibiting fungicides containing the active ingredient polyoxin‐d are used to control fungal diseases in turfgrass, and similar compounds have been shown in the laboratory to synergize baculoviruses by disrupting peritrophic membrane function. This study tested whether applying the virus together with such a fungicide can synergize AgipMNPV activity against A. ipsilon in turfgrass. RESULTS: The addition of a chitin synthesis inhibitor failed to increase AgipMNPV infectivity to A. ipsilon in the field. Rather, delayed and slightly reduced mortality from viral infection was seen when larvae fed on fungicide/virus‐treated grasses as opposed to virus‐only treatments. Choice tests revealed the fungicide residues to be a mild feeding deterrent. CONCLUSION: Because polyoxin‐d does not deactivate AgipMNPV, the two substances are compatible. However, combination applications of polyoxin‐d and Agip MNPV on turfgrass might interfere with larval ingestion of a lethal virus dose, resulting in prolonged larval feeding in the field. Copyright © 2011 Society of Chemical Industry     

马铃薯生长模型研究进展及其应用

马铃薯是世界第四大粮食作物,然而马铃薯的生产却易受到环境因素和遗传因素的影响。通过利用生长模型将马铃薯生理生态过程和相互关系进行量化表达,分析其内部结构与外界环境的关系,从而提出合理的管理方式,达到增产高效的目的。目前,国际上开发了一些较成熟的马铃薯生长模型,如SUBSTOR-potato,NPOTATO,LINTUL-POTATO模型等,应用在国内马铃薯生产实践中取得了一定成果,但总体来说中国马铃薯生长模型的应用还处于探索发展阶段。本文通过对国内外马铃薯生长模型研究的分析总结,得出国内外马铃薯生长模型的主要特点是偏重C、N素研究,缺少P、K素研究;模型所需要的参数较其他作物模型更为复杂。同时,综合分析现有各个生长模型的原理与技术路线的异同点与优缺点,指出马铃薯生长模型需在遗传参数和气候参数方面加以深化研究,并展望了利用作物生长模型指导国内马铃薯生产相关的发展趋势。     

United States Department of Agriculture-Agricultural Research Service: advances in the molecular genetic analysis of insects and their application to pest management

USDA-ARS scientists have made important contributions to the molecular genetic analysis of agriculturally important insects, and have been in the forefront of using this information for the development of new pest management strategies. Advances have been made in the identification and analysis of genetic systems involved in insect development, reproduction and behavior which enable the identification of new targets for control, as well as the development of highly specific insecticidal products. Other studies have been on the leading edge of developing gene transfer technology to better elucidate these biological processes though functional genomics and to develop new transgenic strains for biological control. Important contributions have also been made to the development and use of molecular markers and methodologies to identify and track insect populations. The use of molecular genetic technology and strategies will become increasingly important to pest management as genomic sequencing information becomes available from important pest insects, their targets and other associated organisms.     

玉米抗旱性的QTL分析研究进展和发展趋势

由于分子标记技术的广泛应用,使玉米抗旱性的复杂遗传机制得到逐步了解,其中的一个重要研究进展是通过对玉米抗旱性的数量性状位点(QTL)进行作图和定位,大致了解了一些对抗旱性有显著贡献的基因的染色体位置及其效应,这些结果为分子标记辅助选择和进一步的深入研究打下了基础。在总结分析国际上近十年来在玉米抗旱性QTL分析方面的研究资料的基础上,对研究中涉及到的主要性状及其相互关系、QTL分析中需要注意的主要问题和在该领域的主要研究进展进行了分析评述。认为构建通用的连锁图谱、与功能基因组学研究相结合、应用新的QTL分析技术和抗旱QTL的克隆及其利用将是今后的发展趋势。     

Genetic resistance for the sustainable control of plant virus diseases: breeding,mechanisms and durability

Plant viruses are important agricultural pathogens, and are responsible for a significant number of commercially relevant plant diseases. There are very few efficient control measures for viral diseases, but the use of genetic resistance appears to be the most promising strategy, often conferring effective protection without additional costs or labour during the growing season, and without damaging the environment. Sources of virus resistance have been identified for most crop species, and many resistant cultivars are already commercially available and of widespread cultivation; however, much remains to be learned about genetic resistance. This review article considers three main aspects that require intense investigation. First, we review the identification of sources of resistance and how plant breeders and pathologists have focused on aspects of the breeding process particularly relevant to viruses, such as germplasm screening and the dissection of resistance phenotypes. Second, we review how molecular mechanisms controlling resistance have been unravelled, looking at case studies where resistance mechanisms are now understood in detail for each stage of the infection cycle. Third, we turn to the durability of resistance in a global context, examining factors that influence durability and how this can be predicted. We conclude with a short discussion of the technological and scientific opportunities provided by recent advances in the field.     

新疆野生啤酒花群体遗传多样性的RAPD分析

用RAPD技术分析新疆7个啤酒花(Humulus lupulus L.)的天然居群遗传多样性及群体间的遗传分化,12个随机引物共检测到121个可重复的位点,其中多态位点119个,占总位点的98.35%.由Shannon表型多样性指数和Nei基因多样性指数估计,居群内遗传分化百分比分别为63.55%和65.65%,表明啤酒花种内的遗传变异主要存在于群体内.分析认为,新疆境内啤酒花自然居群的遗传多样性很丰富,居群间产生了一定的分化.啤酒花居群的基因流Nm=1.0037,相对有限的基因流可能在居群遗传分化的维持中起着作用.     

海洋真菌次级代谢产物在植物保护中的研究与应用

病虫害严重影响我国农业生产,每年造成巨额经济损失,传统化学农药虽然在一定程度上能够减轻这些危害,但其不合理的使用也造成严重的负面影响,亟需开发新型生物农药。海洋真菌生存于独特的海洋环境中,形成了有别于陆生真菌的生存繁殖方式和遗传代谢机制,能够产生许多结构新颖、生物活性显著的次级代谢产物,其中许多化合物具有抗植物病原菌、杀虫和抗病毒等农用生物活性,具有开发成为新型生物农药的巨大潜力。本文综述了2010年以来前人在海洋真菌次级代谢产物农用生物活性方面的研究成果,涵盖了90个化合物,结构类型主要包括聚酮类、萜类、生物碱类和甾体类化合物等,以期为生物农药的开发提供依据。     

EPPO Council Colloquium on strategies for reducing use of plant protection products Lyngby (DK), 1996–09–19

The 1996 EPPO Council Colloquium provided an opportunity for European plant protection services to present their alternative strategies for reducing the use of plant protection products. Several countries have set fixed targets for use reduction by certain dates, and have attained many of them. This approach has thus achieved visible success. It has, however, been criticized because use can be measured by different indices, because the real aim should be to reduce risks (to user, consumer and environment), and because it is mainly relevant for countries which have a high level of use in the first place. Other countries try to target their strategies more precisely, particularly by supporting IPM and related approaches. These strategies can be better justified, but it is difficult to assess to what extent they are reducing risks in practice. Yet other countries see the need to use plant protection products more effectively, which does not necessarily imply reducing their use. This is the case particularly for countries which have till recently had moderate levels of use and, in the case of most of central and eastern Europe, have seen this fall sharply in the last few years with the change in economic system. EPPO promotes an approach known as 'good plant protection practice', which seeks to use products optimally, within the limits set by the label recommendations on the one hand, and the possibilities for successful IPM on the other.     

Fitness costs associated with insecticide resistance

Insects are exposed to a variety of stress factors in their environment, and, in many cases for insect pests to agriculture, those factors include toxic chemical insecticides. Coping with the toxicity of insecticides can be costly and requires energy and resource allocation for adaptation and survival. Several behavioural, physiological and genetic mechanisms are used by insects to handle toxic insecticides, sometimes leading to resistance by constitutive overexpression of detoxification enzymes or inducing mutations in the target sites. Such actions are costly and may affect reproduction, impair dispersal ability and have several other effects on the insect's fitness. Fitness costs resulting from resistance to insecticides has been reported in many insects from different orders, and several examples are given in this mini‐review. Copyright © 2012 Society of Chemical Industry     

Characterization of Two Major Genetic Factors Controlling Quantitative Resistance to Melampsora larici-populina Leaf Rust in Hybrid Poplars: Strain Specificity, Field Expression, Combined Effects, and Relationship with a Defeated Qualitative Resistance Gene

ABSTRACT Two genetic factors explain a significant proportion of the variability for quantitative resistance to Melampsora larici-populina leaf rust in a Populus deltoides x P. trichocarpa F(1) progeny. One is inherited from P. deltoides and is associated with a defeated qualitative resistance gene R(1), and the other, R(US), is inherited from P. trichocarpa. To assess the potential contribution of these two factors for durable resistance breeding, 284 genotypes from this F(1) progeny were studied in laboratory experiments with three M. larici-populina strains and in a field experiment under natural inoculum pressure. Results confirmed that both factors can have strong beneficial effects in the laboratory. These effects were strain specific, thus impairing their chances for durability. However, association of both factors led to synergistic effects in most situations. In accordance with good field-laboratory relationships, especially those involving uredinia-size laboratory measurements, field effects of both resistance factors were significant. R(US) led to a significant reduction of rust colonization on the most infected leaf in the field, and its effect was significant both in the presence and the absence of R(1). In contrast, the presence of R(1) was useful in the field only when R(US) was absent. The nature of the genetic relationship between both factors remains unknown, but benefits from their association should be quantified over a longer period to evaluate potential adaptation of the pathogen.