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松材线虫是进境木质包装中的重要检疫对象之一。本研究以进境木质包装中截获的12种伞滑刃属线虫,包括4种不同来源的松材线虫、3种不同来源的拟松材线虫、豆伞滑刃线虫、大尖尾伞滑刃线虫、阿苏里伞滑刃线虫、伯氏伞滑刃线虫及拟小刺伞滑刃线虫为材料,进行实时荧光PCR检测。结果显示,仅松材线虫可观察到明显的荧光强度变化,而其他线虫的荧光强度没有变化。通过研磨样品的方法,提取单条松材线虫的DNA,进行实时荧光PCR试验,检测成功率为100%,且不论是雄虫、雌虫或幼虫均能检测出来。该研究对于我国口岸进境木质包装中松材线虫的检测具有一定的实际意义。 相似文献
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松材线虫种群遗传多样性AFLP标记的建立及其应用 总被引：2，自引：0，他引：2
松材线虫是极具危险性的外来入侵生物,其引起的松材线虫病,目前正在我国部分地区迅速扩展和蔓延,对我国林业生产造成了严重的经济损失。开展松材线虫的种群遗传学研究,是了解其成功入侵和爆发成灾内在机理的重要途径。但迄今为止,尚未找到很有效的分子标记方法来检测松材线虫入侵种群的遗传变异。本文采用AFLP分子标记技术,通过对各反应体系和反应条件的优化及对多态性引物组合的筛选,成功地建立了松材线虫的AFLP分子标记实验体系,并筛选出52对高效多态性的引物组合。应用4对引物对27个松材线虫种群样品进行遗传多样性检测,结果表明,AFLP是进行松材线虫种群遗传学研究的一种很灵敏和可靠的分子标记。此外,本文还对AFLP技术在松材线虫研究中的应用前景进行了讨论。 相似文献
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应用TaqMan-MGB探针进行松材线虫的实时荧光定量检测技术研究 总被引：7，自引：0，他引：7
松材线虫是国际公认的最重要的检疫性有害生物之一,也是我国2类检疫危险性有害生物,我国口岸多次从货物的木质包装中截获该线虫。由于松材线虫与拟松材线虫在形态上极其相似,难以区分,幼虫更无法用于鉴定。传统的形态学鉴定、生化以及其他分子技术等方法存在费时、准确度不高、灵敏度低等缺点,不易形成标准。我们设计筛选一对引物以及一条MGB探针,对松材线虫进行实时荧光PCR检测。建立了一条从1 pg到104pg标准曲线,相关系数r=0.965。该检测方法省时、准确、快速、无污染。 相似文献
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松材线虫(Bursaphelenchus xylophilus)是一种对我国林业危害极大的外来入侵种,具有繁殖力强,扩散范围广的特点。为探究松材线虫的发育调控机制,本研究通过转录组分析和实时荧光定量PCR技术,比较了不同虫态松材线虫发育差异表达基因及功能富集分析。利用LC/MS技术检测了松材线虫代谢物中的蛔甙成分及含量。结果显示:繁殖型和扩散型虫态的差异表达基因主要与滞育和发育调控相关。繁殖型虫态的daf-9基因表达水平显著高于扩散型松材线虫,扩散型LIV期线虫的daf-12、daf-2及daf-16基因表达量显著高于繁殖型松材线虫。Ln和LIII的蛔甙成分比较分析结果显示:繁殖型松材线虫短链蛔甙asc-C3、asc-ΔC7含量显著高于扩散型松材线虫,长链蛔甙asc-C9含量低于扩散型松材线虫。推测松材线虫DAFs基因与蛔甙可能参与松材线虫扩散型虫态的形成。 相似文献
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两种分子技术检测松木中松材线虫的效果评价 总被引：1，自引：0，他引：1
为更好地解决当前松材线虫Bursaphelenchus xylophilus检测工作中频现的假阴性问题,通过不同分子技术检测效果的比较评价,摸索出了一套可提高松材线虫检测准确性的技术体系。针对早期感病松木的低线虫量,建立了线虫的最大量分离方法和基于rDNA ITS序列的分子检测体系,该体系可高效检测出单条松材线虫,准确率达93.75%。以来自浙江省不同疫区的96份松木样品为检测材料,比较了SCAR标记和ITS序列2种分子检测技术的阳性率。结果显示,通过首次PCR,ITS_Ⅰ和ITS_Ⅱ序列的检测阳性率分别为52.08%和55.21%,SCAR标记的检测阳性率为30.21%;通过第二次或巢式PCR,ITS_Ⅰ和ITS_Ⅱ序列的检测阳性率提高到了97.92%和100.00%,SCAR标记的检测阳性率提高到了59.38%,表明基于ITS序列的检测阳性率明显高于SCAR标记,且通过第二次或巢式PCR方法可进一步提高检测灵敏度,降低假阴性率。因此,通过线虫的最大量分离并基于rDNA ITS序列的分子检测可明显提高检测准确性,更适用于松材线虫的常规检测。 相似文献
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ABSTRACT The pinewood nematode, Bursaphelenchus xylophilus, has caused significant damage to pine plantations both in East Asia and North America and is an important quarantine organism. A real-time polymerase chain reaction (PCR) assay was developed to detect B. xylophilus. A set of primers and probe specific for B. xylophilus was designed to target the ribosomal DNA internal transcribed spacer region. Optimal primer concentration, Mg(2+) concentration, and extension temperature were 400 nM, 3.0 mM, and 60 degrees C, respectively. The assay was highly specific and sensitive, detecting as little as 0.01 ng of B. xylophilus DNA. The real-time PCR assay also successfully detected B. xylophilus in field samples, and it should be very useful for quarantine purposes. 相似文献
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The pine wood nematode Bursaphelenchus xylophilus is the causal agent of pine wilt disease. It is a quarantine pest for most countries in the world. Surveys for the occurrence of the nematode may be necessary in the framework of internationally agreed phytosanitary standards, in order to delimit infested areas or to demonstrate the absence of B . xylophilus and thus to identify pest free areas. Import inspections of wood and wood products also need sound monitoring systems. It is obvious that the quality of survey/monitoring, as well as the results of diagnosis, are strongly dependent on the sampling procedure. Although, in the literature, numerous results of surveys and monitoring are documented, sampling procedures vary and there is no scientific or statistically based sampling system published for B. xylophilus . The current paper relates background information concerning the biology of the nematode, its vector beetles and the development of pine wilt disease and the influence of these factors on sampling procedures, and attempts to define a more systematic methodology. 相似文献
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《EPPO Bulletin》1998,28(1-2):27-38
The main technical theme of the 2nd EPPO Worksh op for Heads of Plant Quarantine Services of central and eastern European countries was the risks presented to the forests of the EPPO region by quarantine pests. Measures currently recommended by EPPO concentrate on A1 quarantine pests, not present in the region. Nevertheless, measures applied by many European countries, and in particular by the EU, target non-European forest pests. This presents a particular problem for Russia, since its territory lies partly in Europe and partly in northern Asia and the export of forest products is very important for its economy. Against the background of the new global developments in plant protection (revision of the IPPC, SPS agreement), the Workshop opened the question whether there are forest pests in northern Asia which present a risk to Europe (and vice versa). Recognizing that one of the main non-European pests targeted by the regulations of European countries is Bursaphelenchus xylophilus , the Workshop took note of the fact that this pest does not occur in Russia. Certain known or potential vectors of B. xylophilus in the genus Monochamus do occur in Russia, but also occur widely throughout the palaearctic region. The forest fauna of northern Russia remains fairly homogeneous from Europe to the River Yenisei, and then becomes progressively more different. Nevertheless, specific pests from this eastern region presenting a risk to Europe remain to be identified and subjected to pest risk analysis. 相似文献
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马尾松松材线虫病与松材蓝变相关性研究 总被引：1，自引：0，他引：1
研究表明:松材线虫引起的马尾松(Pinus massoniana)病树死亡后期,必然有蓝变(Blue-stain)现象。松材蓝变程度与松树树干部位相关性不大。但是,对于同一棵树,若蓝变程度高,则每一部位的蓝变程度都比较高;若蓝变程度低,则每部位的蓝变程度可能都比较低。蓝变程度与松材线虫(Bursaphelenchus xylophilus)的含量也没有正相关关系。在松材线虫病木的检疫过程中,松木蓝变仅可以作为松材线虫病的参考指标。 相似文献
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J. C. Ostojá‐Starzewski 《EPPO Bulletin》2014,44(1):34-36
The finding of two pests which are on the EPPO Lists of pests recommended for regulation and are EU quarantine listed pests: Monochamus alternatus and Bursaphelenchus xylophilus, within the structural components of imported furniture are discussed. This highlights the plant health risks associated with this neglected pathway. 相似文献
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由松材线虫(Bursaphelenchus xylophilus)侵染引起的松材线虫病是松属植物的重要病害之一,其繁殖和传播速度快,防治困难。RNAi技术是目前研究植物寄生线虫基因功能的重要手段。本文利用农杆菌介导灰葡萄孢菌表达松材线虫ncr基因的dsRNA(PDH-RH-NCR),并连续多代喂饲松材线虫使ncr基因表达稳定下调后分析其表型。结果表明,在转有PDH-RH-NCR的灰葡萄孢菌上连续培养10代的松材线虫,ncr基因表达量显著下调13倍。ncr基因表达下调后,线虫发育缓慢,繁殖力也显著降低;处理4 d后发育成成虫的百分率为61.75%,显著低于PDH-RH-s GFP(89.75%)和空载体(88.75%)处理;而9 d的繁殖量则较PDH-RH-s GFP和空载体处理分别少2 314和2 347条。松材线虫ncr基因表达下调后,4龄幼虫平均寿命(20.9 d)和体内脂肪量(1.340 nmol)则显著高于PDH-RH-s GFP(15.5 d,0.336 nmol)和空载体(15.2 d,0.323 nmol)处理。通过喂饲法RNAi发现松材线虫ncr基因可能与松材线虫的发育速度、繁殖量、寿命、脂肪积累等表型相关。 相似文献
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松材线虫rDNA的测序和PCR-SSCP分析 总被引：37，自引：3，他引：37
本文为克服形态鉴定的不足,用分子生物学的方法鉴别松材线虫。线虫核糖体DNA(r DNA)的内部转录间隔区(ITS1)区(约308bp)的测序结果显示:松材线虫种内区别很小,不超过1bp;拟松材线虫种内区别较大,最大达7bp;这2种线虫的种间区别为32~39bp。根据以上测序结果,本文结合单条线虫DNA的提取技术,对14个松材线虫和拟松材线虫样本进行了单链构象多态性(PCR-SSCP)分析,结果表明PCR-SSCP分析技术可明确区分这2种线虫,该技术可为单条松材线虫的鉴定提供一套灵敏而可靠的方法。 相似文献