CN1245080C - 一种昆虫拒食剂及其用途 - Google Patents

一种昆虫拒食剂及其用途 Download PDF

Info

Publication number
CN1245080C
CN1245080C CN 01130078 CN01130078A CN1245080C CN 1245080 C CN1245080 C CN 1245080C CN 01130078 CN01130078 CN 01130078 CN 01130078 A CN01130078 A CN 01130078A CN 1245080 C CN1245080 C CN 1245080C
Authority
CN
China
Prior art keywords
antifeedant
insect
nematode
bacterium
xenorhabdus
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Expired - Fee Related
Application number
CN 01130078
Other languages
English (en)
Other versions
CN1372798A (zh
Inventor
韩日畴
刘秀玲
曹莉
李秋剑
陈镜华
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Guangdong Entomological Institute
Original Assignee
Guangdong Entomological Institute
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Guangdong Entomological Institute filed Critical Guangdong Entomological Institute
Priority to CN 01130078 priority Critical patent/CN1245080C/zh
Publication of CN1372798A publication Critical patent/CN1372798A/zh
Application granted granted Critical
Publication of CN1245080C publication Critical patent/CN1245080C/zh
Anticipated expiration legal-status Critical
Expired - Fee Related legal-status Critical Current

Links

Landscapes

  • Agricultural Chemicals And Associated Chemicals (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

本发明涉及一种昆虫拒食剂及其用途，它选自人工培养的Xenorhabdus或Photorhabdus属细菌的培养液、培养液的上清液或培养液中滤渣的水稀释液；将其应用于植物体特别是十字花科的蔬菜的表面或地下部分、生长环境中时，可阻止世界性的蔬菜主要害虫小菜蛾(Plutella xylostella)取食植物体。

Description

一种昆虫拒食剂及其用途
技术领域
本发明涉及一种昆虫拒食剂，进一步涉及这种昆虫拒食剂的用途。
技术背景
昆虫病原斯氏科Steinernematidae与异小杆科Heterorhabditidae线虫是新型生物杀虫剂。这类线虫具有广泛的寄主范围；对寄主具主动搜寻能力，特别是对土栖性及钻蛀性害虫；对人畜、环境安全。近年来，已广泛应用于防治农、林、牧草及卫生等害虫，受到国内外学者及商业部门的高度重视，并走向商品化。
这类线虫是以感染期虫态(infective juveniles，IJ)随寄主食物或从昆虫的自然开口(如肛门、气孔)、节间膜进入昆虫体内，随后释放肠腔中携带的Xenorhabdus属(目前与斯氏科线虫共生)或Photorhabdus属(目前与异小杆科线虫共生)的共生细菌。线虫以及共生细菌分泌的毒素(毒性因子)导致昆虫死亡。
原先，科学家们认为，Xenorhabdus属或Photorhabdus属的共生细菌只有被线虫携带或注射进入昆虫体内时才发挥杀虫作用[Akhurst(1980)Journal of General Microbiology121，303-309；Bedding(1984)Annals of Applied Biology 104，117-120；Forst & Nealson(1996)Microbiological Review 60，21-43]。最近发现，共生细菌可通过口服单独致死昆虫[李素春等(1995)第六届全国杀虫微生物学术讨论会论文摘要集p.43；Jarrett et al.(1998)PCT PatentApplication No.98/08388；Ragni et al.(1998)PCT Patent Application No.98/05212]，从共生菌中可分离出蛋白质毒素[Ensign et al.(1997)PCT Patent Application No.97/17432；Bowen et al.(1998)Science 280，2129-2132；Ensign et al.(2000)US Patent，Application No.072264]，因此，昆虫病原线虫共生细菌在虫害防治方面的应用也受到重视。杀虫蛋白毒素的发现为进一步研制新型微生物农药以及创建防虫转基因植物提供了新的途径。所有已发表或公开的文献指出，昆虫病原线虫的共生细菌或共生细菌分泌的杀虫蛋白对昆虫有毒杀作用(toxicactivity)，但未发现这类细菌对昆虫的拒食作用。
技术内容
本发明的目的是提供一种昆虫拒食剂，进一步目的是提供这种昆虫拒食剂的用途。
本发明的一种昆虫拒食剂，选自人工培养的Xenorhabdus或Photorhabdus属细菌的培养液、培养液的上清液或细菌培养液中滤渣的水稀释液。
本发明所公开Xenorhabdus和Photorhabdus属细菌分离自斯氏科Steinernematidae斯氏属Steinernema与异小杆科Heterorhabditidae异小杆属Heterorhabditis昆虫病原感染期线虫，目前在分类上归属于Enterobacteriaceae科。Xenorhabdus属细菌目前经描述的种有Xenorhabdus nematophilus、X.poinarii、X.bovienii、X.beddingii或X.kushidaii。Photorhabdus属细菌目前经描述的种有P.luminescens。这些细菌的分类地位以后的可能改变并不会影响本发明的限制范围。
Xenorhabdus属细菌选自Xenorhabdus nematophilus、X.poinarii、X.bovienii、X.beddingii或X.kushidaii种细菌。Photorhabdus属细菌有P.luminescens种细菌。
滤渣中包含有灭活菌体、灭活菌体的片段以及其他固体物质。
本发明的一种昆虫拒食剂的制备方法即培养方法，是将所属细菌经分离、纯化后，以人工培养基(如蛋白胨水培养基或其它能够培养这类细菌的培养基)体外培养，包括以发酵罐或反应器系统培养，所获得的菌液可直接或经分离、提取和提纯，得到的有效组分用做昆虫拒食剂。
本发明的这种昆虫拒食剂的用途是将其应用于植物表面或地下部分、生长环境中，可阻止世界性的蔬菜主要害虫小菜蛾(Plutella xylostella)取食植物体。特别是将其应用于十字花科的蔬菜的表面或地下部分、生长环境中，特别能阻止世界性的蔬菜主要害虫小菜蛾(Plutellaxylostella)取食十字花科的蔬菜。
施用量和施用次数根据小菜蛾的发生期、发生数量以及气候条件等而定。这些拒食剂可通过不同的剂型如水剂、可湿性粉剂、颗粒剂和浓缩剂等配置，首选水剂，既经济又方便；也可加入辅助剂如吸附剂、湿润剂、防腐剂、粘着剂、引诱剂、保护剂和增效剂等使用。
具体实施方式
下列实验及操作实例是进一步对本发明的说明，不应该当作对本发明的限制。实例中仅描述X.nematophilus和P.luminescens，但适用于所有Xenorhabdus或Photorhabdus属细菌。
实例一
按下列方法分别分离Xenorhabdus和Photorhabdus初生型共生细菌：昆虫病原线虫感染的大蜡螟末龄(七龄)幼虫Galleria mellonella，置于25℃下，昆虫死亡后(一般2至4天)，以70％酒精体表消毒死虫，然后用无菌剪刀剪开虫体，取血淋巴划营养琼脂和麦康凯平板。根据Akhurst(1980)(Journal of General Microbiology 121，303-309)描述的方法鉴定线虫共生细菌。从S.carpocapsae A24线虫中分离出X.nematophilus共生菌；从H.bacteriophora线虫中分离出P.luminescens共生菌。
取二个500ml三角瓶中，分别加入100ml液体培养基(1％蛋白胨，0.5％NaCl)，121℃下高压消毒30分钟。然后分别接入从S.carpocapsae A24和H.Bacteriophora H06线虫中分离出的X.nematophilus和P.luminescens初生型共生菌(5×108菌体)，置于25℃、150rpm下摇床分别培养48小时，所获得的培养菌液作为昆虫拒食剂，采用叶蝶法分别测定二种菌液对小菜蛾幼虫的拒食作用：
用打孔器(直径＝2cm)将椰菜叶作成叶蝶，将其浸泡在不同的菌液中(50ml菌液中加入3滴高压消毒的甘油)5秒钟，取出并凉干叶蝶，将其放入垫有2层无菌中速滤纸(新华)的培养皿中(直径＝7.5cm)。实验分两种方法处理：
一、选择性测定；每培养皿加两片叶蝶，一片浸泡菌液的叶蝶和另一片对照叶蝶(不浸泡菌液，仅浸泡加入高压消毒的甘油的无菌水)，然后往每个培养皿中接入3龄小菜蛾幼虫10头。每处理设3个平板的重复。置于25℃下。试验后24，48和72小时，检查叶蝶被取食的面积。
二、非选择性测定：每培养皿加两片浸泡菌液的叶蝶，另设对照叶蝶(不浸泡菌液，仅浸泡加入高压消毒的甘油的无菌水)放于另一培养皿(对照培养皿)中，然后往每个培养皿中接入3龄小菜蛾幼虫10头。每处理设3个平板的重复。置于25℃下。试验后24，48和72小时，检查叶蝶被取食的面积。
实验证明，用X.nematophilus和P.luminescens培养的菌液无论是在选择性测定或非选择性测定中，对小菜蛾幼虫均具有拒食作用。72小时后，在选择性测定平板中，未经菌液处理的菜叶已被小菜蛾幼虫吃光，但经菌液处理的菜叶面积仍完好；在非选择性测定中，经菌液处理的菜叶面积完好，未被小菜蛾幼虫破坏，即使小菜蛾幼虫由于饥饿而死亡也不取食菜叶。由此可见，X.nematophilus和P.luminescens培养的菌液对小菜蛾幼虫均具有强烈拒食作用。
实例二
按实例一同样的方法从S.carpocapsae A24线虫中分离出X.nematophilus共生菌；从H.bacteriophora线虫中分离出P.luminescens共生菌，并分别制备X.nematophilus和P.luminescens的培养菌液，所获得菌液经14000rpm，10分钟离心分离，上清液以0.2μm孔径的细菌过滤器过滤，滤渣(菌体)则以等量无菌水替代上清液稀释，分别以上清液和滤渣(菌体)水稀释液作为昆虫拒食剂，然后也按实施例一的叶蝶法测定其对小菜蛾幼虫的拒食作用。
实验证明，X.nematophilus和P.luminescens培养的菌液，无论是上清液还是菌体水稀释液，无论是在选择性测定或非选择性测定中，对小菜蛾幼虫均具有强烈拒食作用。

Claims (1)

1.Xenorhabdus nematophilus或Photorhabdus luminescens细菌的培养液在制备昆虫拒食剂中的用途。
CN 01130078 2001-12-14 2001-12-14 一种昆虫拒食剂及其用途 Expired - Fee Related CN1245080C (zh)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN 01130078 CN1245080C (zh) 2001-12-14 2001-12-14 一种昆虫拒食剂及其用途

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN 01130078 CN1245080C (zh) 2001-12-14 2001-12-14 一种昆虫拒食剂及其用途

Publications (2)

Publication Number Publication Date
CN1372798A CN1372798A (zh) 2002-10-09
CN1245080C true CN1245080C (zh) 2006-03-15

Family

ID=4669708

Family Applications (1)

Application Number Title Priority Date Filing Date
CN 01130078 Expired - Fee Related CN1245080C (zh) 2001-12-14 2001-12-14 一种昆虫拒食剂及其用途

Country Status (1)

Country Link
CN (1) CN1245080C (zh)

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101638434B (zh) * 2008-07-30 2011-09-07 中国农业科学院植物保护研究所 嗜线虫致病杆菌拒食蛋白质及其基因序列和该拒食蛋白及其基因的用途

Also Published As

Publication number Publication date
CN1372798A (zh) 2002-10-09

Similar Documents

Publication Publication Date Title
Llácer et al. Evaluation of the efficacy of Steinernema carpocapsae in a chitosan formulation against the red palm weevil, Rhynchophorus ferrugineus, in Phoenix canariensis
Gerson et al. Mites (Acari) for pest control
Jones et al. The uses of pollen and its implication for entomology
Sharma et al. Entomopathogenic nematodes, a potential microbial biopesticide: mass production and commercialisation status–a mini review
Gumus et al. A novel approach to biocontrol: Release of live insect hosts pre-infected with entomopathogenic nematodes
Pérez et al. Use of entomopathogenic nematodes to suppress Meloidogyne incognita on greenhouse tomatoes
Pérez et al. Suppression of Meloidogyne incognita and Meloidogyne hapla with entomopathogenic nematodes on greenhouse peanuts and tomatoes
Prasad et al. Evaluating prospects of fungal biopesticide Beauveria bassiana (Balsamo) against Helicoverpa armigera (Hubner): An ecosafe strategy for pesticidal pollution
Kaaya et al. Pathogenicity of Beauveria bassiana, Metarhizium anisopliae and Serratia marcescens to the banana weevil Cosmopolites sordidus
Ishibashi et al. Dynamics of the entomogenous nematode Steinernema feltiae applied to soil with and without nematicide treatment
Wilson et al. Mass cultivation and storage of the rhabditid nematode Phasmarhabditis hermaphrodita, a biocontrol agent for slugs
Morton et al. Ecological characterization of entomopathogenic nematodes isolated in stone fruit orchard soils of Mediterranean areas
Patlar et al. Aphid (Hemiptera: Aphididae) species in Burdur urban parks with three records for the fauna of Turkey, their host plants and predators
Perry et al. EVect of the Entomopathogenic Nematodes, Steinernema feltiae and S. carpocapsae, on the Potato Cyst Nematode, Globodera rostochiensis, in Pot Trials
Atwa Entomopathogenic nematodes as biopesticides
Peters Application and commercialization of nematodes
CN1115408C (zh) 一种昆虫病原线虫的液体培养方法
Mahar et al. Microbial control of diamondback moth, Plutella xylostella L.(Lepidoptera: Yponomeutidae) using bacteria (Xenorhabdus nematophila) and its metabolites from the entomopathogenic nematode Steinernema carpocapsae
CN101892170A (zh) 昆虫病原线虫共生菌及其应用
CN1245080C (zh) 一种昆虫拒食剂及其用途
Saleh et al. Commercialization of biopesticides based on entomopathogenic nematodes
SUSURLUK Effectiveness of the entomopathogenic nematodes Heterorhabditis bacteriophora and Steinernema feltiae against Tenebrio molitor (yellow mealworm) larvae in different soil types at different temperatures
Sivinski The attraction of lepidoptera to flowering plants also attractive to parasitoids (Diptera, Hymenoptera)
Mahar et al. Control of insects with entomopathogenic bacterium Xenorhabdus nematophila and its toxic secretions.
Drew et al. The biology and behavior of flies in the tribe Dacini (Dacinae)

Legal Events

Date Code Title Description
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
C14 Grant of patent or utility model
GR01 Patent grant
C17 Cessation of patent right
CF01 Termination of patent right due to non-payment of annual fee

Granted publication date: 20060315

Termination date: 20101214