鹰嘴豆象与四纹豆象的分子检测技术

结合聚合酶链反应(polymerase chain reaction,PCR)和酶切方法,对鹰嘴豆象Callosobruchus analis(Fabricius)和四纹豆象Callosobruchus maculatus(Fabricius)的分子检测技术进行了研究。根据两种豆象m tDNACOⅠ基因序列的比对结果设计了2对特异性PCR引物,并筛选到2个特异性内切酶SnaⅠ和MboⅠ,通过PCR反应(最适退火温度为55℃)和PCR产物的酶切反应检测2个近缘种。可靠性检测表明,2对引物针对不同地理种群和个体以及不同DNA浓度的鹰嘴豆象和四纹豆象均能扩增出相应的目的条带。限制性内切酶验证结果表明,PCR技术具有较高的准确性和灵敏度,弥补了形态鉴定的不足。     

高温与磷化铝对四纹豆象的处理技术

采用不同温度和不同剂量的磷化铝,在不同的处理时间一对四纹豆象的高温灭虫及磷化铝熏蒸试验,结果表明：四纹豆象肾的而高温能力最强,卵次之,在５５℃温度下幼虫和成虫经５０ｍｉｎ校正死亡率为１００％,肾和卵要７５ｍｉｎ,在６０℃温度下卵、幼虫、成虫经１５ｍｉｎ全部死亡,而肾需要４５ｍｉｎ才达到１００％的死亡率。用经铝熏蒸,幼虫和成虫较第敏感,用１．５ｇ／ｍ＾３处理３天,人武部死亡,但卵、蛹的校正死亡率仅为     

豇豆对四纹豆象的抗性

四纹豆象[Callosobruchus maculatus(F.)]是危害豇豆[Vigha unguiculata(L.)Walp]种子的一种害虫,常可造成贮藏种子的严重损失。四纹豆象首先在田间侵入豆荚、然后随种子进入仓库(Pr-evett,1961),此后便开始大量繁殖,成虫产卵于种子表面,卵在5—7天内孵化。幼虫孵化后钻入种子,在种子内完成其生长发育过程。成虫羽化后便离开种子,在种子     

6种植物源农药对四纹豆象的触杀活性

采用药膜滤纸法测定除虫菊素、烟碱、鱼藤酮、蛇床子素、印楝素、苦参碱6种植物源农药四纹豆象的触杀活性。结果表明,6种供试药剂对四纹豆象的LC50分别为7.0771、7.5804、66.3049、16.1044、25.4777和85.5233μg/cm2;触杀活性测定结果表明,除虫菊素对四纹豆象的触杀活性最强,烟碱次之,其它4种供试药剂对四纹豆象的触杀活性较弱。     

谷斑皮蠹分子检测方法

根据Simon(1994)发表的COI基因的通用引物,扩增了谷斑皮蠹、黑斑皮蠹、条斑皮蠹和花斑皮蠹不同地理种群的COI基因片段.经序列比对后,依据多态位点,设计了检测谷斑皮蠹的特异引物.PCR结果表明:引物可成功扩增出谷斑皮蠹样品的基因片段.而其他斑皮蠹属种类基因片段不能被扩增.     

5种仓储豆象的分子鉴定和检测

本研究以在检疫中经常被截获的5种豆象为研究对象,通过对GenBank中5种豆象mtDNA COⅠ序列的查询,并进行序列比对,共设计了30对引物,通过筛选获得5对能分别鉴定5种豆象的特异引物,并用50、25、10、5、1、0.5、0.1、0.05ng/μL 8个不同浓度系列的豆象DNA模板来检测灵敏度,结果表明,豌豆象和四纹豆象特异引物对的灵敏度为0.1ng/μL,蚕豆象、菜豆象和绿豆象特异引物对的灵敏度为0.5ng/μL。该方法可用于快速准确地鉴定5种豆象,对于豆象类害虫的检测监测具有重要意义。     

红火蚁实时荧光分子检测方法

根据GenBank上发表的火蚁属(Solenopsis)COI基因序列,手工设计了检测红火蚁(Solenopsis invicta)的引物与探针.实时荧光PCR结果表明:红火蚁样品可被成功地检测到,而近似种热带火蚁没有被检测到,同种不同虫态的个体得到一致结果.     

四纹豆象生物、生态学特性及防治方法研究进展

四纹豆象(Callosobruchus maculatus Fabricius)是我国进境植物检疫性有害生物之一,严重危害豇豆、豌豆、绿豆、鹰嘴豆、大豆、扁豆、赤豆、胡豆等豆类。本文对四纹豆象的生物学和生态学特性以及防治等方面的研究概况进行了综述,以期引起人们对四纹豆象的充分重视。     

基于DNA条形码技术的河南省夏季重要实蝇害虫分子鉴定

以夏季采自河南省洛阳市、郑州市的实蝇幼虫、成虫为研究材料,采用DNA条形码技术,通过测定、比对线粒体DNA细胞色素氧化酶I(COI)基因片段序列,对其进行种类鉴定。结果表明,检测的实蝇样品种类为橘小实蝇(Bactrocera dorsalis)、具条实蝇(Zeugodacus scutellatus)和三点棍腹实蝇(Dacus trimacula),与相关种已知序列的相似度均在99.39%以上,其系统发育树明显与我国其他常见实蝇近缘种区分开。     

线粒体COI基因作为根结线虫条形码的适用性分析

为评估线粒体COI基因作为根结线虫(Meloidogyne spp.)DNA条形码的适用性,对收集的25个根结线虫群体的线粒体COI基因进行扩增、测序,检测其扩增和测序成功率。将测序获得的COI基因序列与Q-bank中下载的根结线虫COI基因序列共同进行比对分析,利用MEGA 6.0计算种内、种间遗传距离进行"Barcoding Gap"检验以及构建NJ系统进化树作聚类分析,检验COI基因对根结线虫的物种识别能力。结果表明:线粒体COI基因扩增和测序成功率极高,扩增成功率达96%,测序成功率为100%。种间平均遗传距离为0.126,远远超过种内平均遗传距离0.002的10倍,种内遗传距离与种间遗传距离存在明显的"Barcoding Gap"间隔区。系统发育分析显示,COI基因可有效分辨出本研究15种根结线虫中的10种,物种识别率约为67%。线粒体COI基因具有极高的扩增和测序成功率、较高的物种识别能力,且扩增片段长度适宜,符合物种鉴定DNA条形码基因的标准,可作为根结线虫候选条形码基因。     

Effects of Hyptis spicigera Lam. (Labiatae) on the behaviour and development of Callosobruchus maculatus F. (Coleoptera: Bruchidae), a pest of stored cowpeas

Abstract

The use of beni seed, Hyptis spicigera Lam. (Labiatae), to protect stored cowpeas from bruchid attacks is a widespread practice in West Africa. More knowledge about its actual effects on storage pests is needed to enhance its effectiveness. In this laboratory study, repellent and lethal effects of powder and essential oil from H. spicigera on Callosobruchus maculatus, the main pest of stored cowpeas, were studied. Repellency tests undertaken with a Y-tube olfactometer showed that powder was repulsive for C. maculatus adults. On the other hand, this product had no insecticidal activity. Essential oil had a dose-dependent insecticidal effect while sublethal doses repelled the adults. Both products reduced oviposition by 15 – 19 and 7 – 78% according to the doses of powder and essential oil, respectively, tested. Similarly, egg viability decreased with increasing doses of powder and essential oil used in a range of 40 – 75 and 24 – 86%, respectively. Only essential oil was lethal to C. maculatus larvae developing within cowpea seeds; however, oil activity was age-dependent, younger instars being more susceptible. These results are discussed with a view of controlling C. maculatus populations in stored cowpeas.     

Evaluation of powdered fruits of four peppers (Capsicum spp.) for the control of Callosobruchus maculatus (F) on stored cowpea seed

Abstract

The efficacy of two application rates of powdered fruits of four types of peppers (Capsicum spp.) was evaluated for the control of Callosobruchus maculatus (F). The various pepper powders at 2.5 g‐5.0 g per 250 g of seed were effective in reducing oviposition and damage to cowpea seeds. The pepper type Capsicum frutescens var. yarmunchi was the most effective in discouraging oviposition and minimizing damage to the seed, as indicated by the significantly lower number of emergence holes. The seed quality and viability were not affected by the powdered pepper fruit treatment. The result is indicative of the potential for the use of powdered fruits of some peppertypes in protecting stored cowpea seeds against C. maculatus.     

基于DNA条形码技术的泰国番石榴中实蝇幼虫分子鉴定研究

实蝇是世界重要的检疫性害虫之一,对果蔬生产及其国际贸易具有很大的影响。而以线粒体DNA细胞色素C氧化酶亚单位I(mtDNACOI)基因的部分序列作为实蝇的DNA条形码能减少对实蝇成虫形态特征的依赖,可检测其任何虫态的样品,有助于实蝇样品的快速鉴定。本研究采用DNA条形码技术,针对采自泰国四色菊市番石榴烂果中的5头实蝇幼虫进行COI扩增测序,与生命条形码数据库(BOLD)中的序列进行比对并利用PAUP4.0软件构建了其系统进化树。根据序列分析和系统进化关系分析的结果,将5头实蝇幼虫样品鉴定为番石榴实蝇(Bactrocera correctaBezzi),并将本研究获得的2条COI序列在GenBank中注册,GenBank的登录号为HM590450和HM590451。     

基于DNA条形码的红翅大小蠹和黄杉大小蠹分子鉴定

外来有害生物的快速准确鉴定有助于提高口岸检疫效率。红翅大小蠹(Dendroctonus rufipennis)和黄杉大小蠹(D. pseudotsugae)作为大小蠹属的非中国种是我国口岸原木检疫中频繁截获的检疫性有害昆虫。本研究以mtDNA COI基因5′端和3′端序列为标记对红翅大小蠹与黄杉大小蠹的11个样本开展分子鉴定有效性评价。结果显示,当以COI基因5′端和3′端为靶标时,两种大小蠹间的平均遗传距离分别为0.181和0.144,是种内遗传距离的60.3和48倍,且种内、种间遗传距离间无重叠现象,在系统发育树上都能聚为独立的分支,表明mtDNA COI基因5′端和3′端序列均可有效区分红翅大小蠹与黄杉大小蠹。     

以Ypt1基因序列为靶标的辣椒疫病菌快速分子检测

PCR primers were designed based on the sequence of Ras-related protein gene (Ypt1) of P. capsici. According to the multiple sequence alignment, Ypt1 has the sufficiently polymorphic intron region for the development of P. capsici-specific primers (PcYpt1F/PcYpt1R). One primer pair was developed which can amplify one P. capsici-specific fragment of 156 bp. Using the primer pair, the P. capsici infected plants and soils were detected. Additionally, Ypt1 has an appropriate region for the development of Phytophthora genus-specific primers (Ypt1F/Ypt1R), which can amplify a fragment of about 540 bp from 14 different Phytophthora specices and a fragment of about 350 bp in Pythium species, with no amplification from fungal species. By PCR optimization using P. capsici genomic DNA, the detection sensitivities of 10 pg and 10 fg DNA were achieved in standard PCR (PcYpt1F/PcYpt1R) and nested PCR (Ypt1F/Ypt1R and PcYpt1F/PcYpt1R), respectively. The developed primers were proved to be efficient in detection of Phytophthora pathogens from diseased plant tissues and residues in soils.     

Efficacy of Artocarpus altilis (Parkinson) Fosberg extracts on contact mortality,repellency, oviposition deterrency and fumigant toxicity of Callosobruchus maculatus (F.) (Coleoptera:Bruchidae)

Callosobruchus maculatus (F.), a major pest of grain legumes, causes considerable losses worldwide. C. maculatus management relies on synthetic insecticides with their adverse environmental and health concerns. The current study investigated the use of Artocarpus altilis (breadfruit) extracts on contact and fumigant toxicity, repellency and oviposition deterrency of C. maculatus as a management strategy. Fumigant mortality (LC₅₀) of the dry inflorescence extract of A. altilis was 126.65 µL L^?1 air at 53.4 h. Fruit extract gave 73.3% repellency at 1000 ppm after 24 h. Percent repellency was dose and time dependent. Green inflorescence extract of A. altilis reduced egg laying with 50% oviposition occurring after 37.78 h at 1000 ppm. We conclude that extracts of leaf, fruit, green and dried inflorescence of A. altilis displayed contact mortality, repellency, deterred oviposition and had fumigant activity toward C. maculatus.     

苹果异形小卷蛾的分子鉴定研究

苹果异形小卷蛾（Thaumatotibia leucotreta）是世界范围内重要的检疫性有害生物,对水果和农作物等生产具有重要影响,并影响国际贸易发展。本研究选用线粒体 COI 基因作为物种鉴定的条形码,结合 GenBank 数据库、BOLD 数据库比对和系统进化树构建,对2014年1月广州白云国际机场口岸入境加纳牛油果中截获的卷蛾科昆虫进行分子鉴定。根据 GenBank 数据库比对分析、NJ方法构建的系统进化树分析以及 BOLD 数据库查询分析,最终鉴定该样品为苹果异形小卷蛾。认为基于 DNA 条形码的分子鉴定能快速、准确地鉴定苹果异形小卷蛾。