Identification of Candidatus Liberibacter asiaticus from Wampee in Guangdong
英国植物新品种保护测试_下_
分类号:B J A农民日报/2001年/07月/26日/第006版/种业论坛英国植物新品种保护测试(下)徐一力D U S测试的进行(2)对照品种库的建立和对照品种的筛选。
英国的植物新品种保护中重要的工作之一是建立了各类作物的对照品种库,因为这样才能使申请保护的品种有对照和对比,才能准确地确定品种的特异性、一致性和稳定性。
对照品种来自现有的已知品种,包括各国受保护的、进人各国国家目录的、已提出申请植物育种者权利或进入国家目录的、在生产中已使用或被销售的以及出版物上刊登的品种等。
有些品种则向育种者或植物园购买。
英国D U S测试机构的对照品种库中的对照品种种类繁多,保证了对照品种的完整性。
对照品种入库按某一性状进行分级、分组保存,这些信息全部由计算机来管理。
入库前进行登记、称重、测定纯度、发芽率、水分含量。
有时还采用电子扫描与电脑结合的现代化技术来对照品种进行分级分组。
而对花卉等观赏植物的对照品种则放在活体对照库中进行保存,对马铃薯等采用脱毒薯块进行保存。
对照品种库的管理也是十分复杂的。
英国测试机构对照品种库在入库品种的水分及库内的湿度、温度都摸索出了十分丰富的经验,在投入最节省的条件下(节电、节能、节省人工),要使对照品种保存25年以上。
目前,英国正在探索用现代化的生物技术手段来分类,以减少库内种子的数量。
在进行D U S测试前,英国的测试人员对申请者提供的申请品种的各种性状进行研究,同时,按申请者要求的对照品种的类型,从计算机管理的对照品种库中选取对照品种,这样对照品种的筛选就更加科学和准确。
(3)技术问卷的填写和品种材料的提供。
进行D U S测试前,英国的育种者首先填写申请调查表,详细地填写申请品种的各种性状、选育过程、亲本来源或材料来源。
同时,要填写品种的发芽率、纯度、净度以及品种的质量等。
有些品种,如观赏的菊花,还需要交照片。
育种者必须提供充足的种子(材料)用于测试和保藏,一般大田作物需3k g种子,蔬菜种子需120g左右。
ERIC_PCR鉴别苏云金芽胞杆菌与蜡状芽胞杆菌的研究
ERIC PCR 鉴别苏云金芽胞杆菌与蜡状芽胞杆菌的研究*李 强1金莉莉1王 芳2宋树森1王秋雨1**(辽宁大学生命科学学院 沈阳 110036)1 (沈阳出入境检验检疫局 沈阳 110016)2摘要:利用ERIC PCR 技术对苏云金芽孢杆菌(B t)、蜡状芽孢杆菌(Bc)和对照菌基因组DNA 进行扩增,回收、标记Bt PCR 扩增片段,分别与各菌株的基因组DNA 进行斑点杂交和Southern 杂交,筛选Bt 标识序列。
结果显示:Bt 各菌株均可扩增得到250bp 的特异片段;Bt 和Bc 均可得到600bp 的共有扩增片段;以筛选得到的569bp 片段为探针,可特异性地与B t 基因组DNA 杂交;ERIC PCR 技术可以在DNA 指纹图谱水平区分鉴别Bt 与Bc 菌,正确反映出两者的亲缘关系。
结果表明ERIC PCR 技术在Bt 的检测及在Bt 与Bc 的鉴定中具有较强的实用性。
关键词:苏云金芽孢杆菌,蜡状芽孢杆菌,ERIC PCR,指纹图谱,Sou thern 杂交中图分类号:Q93 3 文献标识码:A 文章编号:0253 2654(2007)06 1184 04Identification of Bacillus thuringiensis and Bacillus cereus by ERIC PCR *LI Qiang 1 JIN Li Li 1 WANG Fang 2 S ONG Shu Sen 1 WANG Qiu Yu 1**(Li f e Sc ienc e De pa rtme nt o f Liaon in g U ni versit y ,Sh en ya ng 110036)1(Liaon in g En try Exit Inspection an d Qu aran tine Bu re au ,Shen yan g 110016)2Abs tract :To screen the id entifier of B .thu rin gien sis ,gen omic D NA of B .th uring iensis ,B .ce re us and con trol strains were amplificated b y ERIC PCR.Fragments amp lified from B .th u rin gien sis were retrieved ,cl on ed ,marked b y 32P,d ot bl ot and southe rn h ybri dized with genomic DNA of these strains.It sho ws th at every strai ns of B .thu ring ie nsis can be a mp li fied to produce a 250b p fragmen t,and b oth B .th uring iensis an d B .ce reu s can be amplified to produce a 600bp fragment.Hyb ridi zation resul t bet w een 569bp probe and genomic DNA of B .thu rin gien sis sh ows a hi gh specificity.Discri mi nation an d genetic relati on shi p be tween B .thu rin gien sis and B .ce re us can be re vealed b y ERIC fin gerprin t.This researc h i ndicates th at ERIC PCR techn iqu e is a p ractical method in the d etec ti on and c haracteri zati on of B .thu rin gien sis an d B .ce reu s .K ey words :B .thu ring iensis ,B .cereus ,ERIC PCR,D NA fin gerprin ts,S ou thern b lotti ng*辽宁出入境检验检疫局资助项目(No.LK 30 2002)**通讯作者 Tel:024 ********,E mail:qiuyuwang@ 收稿日期:2007 01 27,修回日期:2007 07 20苏云金芽孢杆菌(Bacillus thuringiensis ,简称Bt)是一种自然界中广泛分布的革兰氏阳性菌,属于芽孢杆菌属。
拟金钱菌属的4个中国新记录种
拟金钱菌属的4个中国新记录种
杨越婷;熊勇;兰勇;岑进;彭巧华;栾丰刚;易伶俐
【期刊名称】《生物灾害科学》
【年(卷),期】2024(47)1
【摘要】【目的】对采集于江西省官山国家级自然保护区的9份拟金钱菌属Collybiopsis标本进行物种鉴定。
【方法】依据宏观形态和显微结构比较,结合ITS 和LSU基因序列系统发育进行分析与鉴定。
【结果】9份拟金钱菌属标本分属于7个种,即韩国拟金钱菌(Collybiopsis. Koreana)、富尔瓦拟金钱菌(C. fulva)、梅尼胡内拟金钱菌(C.menehune)、东方缺裸拟金钱菌(C. orientisubnuda)、安杜拉塔拟金钱菌(C. undulata)、茂盛拟金钱菌(C. luxurians)和多纹拟金钱菌(C. polygramma)。
【结论】参考中国截至目前已报道的拟金钱菌属物种,确定韩国拟金钱菌、富尔瓦拟金钱菌、梅尼胡内拟金钱菌和东方缺裸拟金钱菌为我国新记录种,其余3种为江西省新记录种。
【总页数】10页(P57-66)
【作者】杨越婷;熊勇;兰勇;岑进;彭巧华;栾丰刚;易伶俐
【作者单位】江西农业大学林学院;江西官山国家级自然保护区管理局
【正文语种】中文
【中图分类】Q949.32
【相关文献】
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2.中国粉孢革菌科一新记录属(白缘皱孔菌属)和一新记录种(拟软白缘皱孔菌)
3.中国新记录属——乌菌蚊属及一新种记述(双翅目:扁角菌蚊科)(英文)
4.干脐菇属中国新记录种——金钱菌型干脐菇
5.网柄菌属的2个中国新记录种和2个中国大陆新记录种
因版权原因,仅展示原文概要,查看原文内容请购买。
意大利苍耳提取物抗植物病原真菌活性研究
意大利苍耳提取物抗植物病原真菌活性研究意大利苍耳(Cetraria islandica)是一种生长在寒冷地区的苔藓类植物,具有多种药用和生物活性成分。
本研究通过提取意大利苍耳的苍耳素(usnic acid)并对其抗植物病原真菌活性进行研究。
本研究采集了生长在寒冷地区的意大利苍耳植物,并将其经过适当的处理,并使用醇提取的方法,得到了纯度较高的苍耳素。
然后,使用适当的溶剂稀释苍耳素溶液,并进行了一系列不同浓度的体外抗真菌活性实验。
真菌菌株的来源包括了几种常见的植物病原真菌,如白色念珠菌(Candida albicans)、白色念珠丝菌(Candida tropicalis)和黑曲霉(Aspergillus niger)等。
结果显示,意大利苍耳提取物对上述植物病原真菌显示出一定的抑制活性。
通过浓度梯度的实验观察到,苍耳素对这些真菌的生长和代谢活动产生了明显的抑制效果。
而且,这种抑制效果呈现出明显的剂量依赖性,随着浓度的增加,抑制效果也越强。
对于白色念珠菌和白色念珠丝菌来说,苍耳素在低浓度下对其生长速率有显著的抑制效果;而对于黑曲霉来说,即使在高浓度下也表现出了一定的抑制效果。
进一步研究发现,苍耳素抑制真菌活性的机制可能与其对真菌细胞膜的作用有关。
研究发现,苍耳素可以改变真菌细胞膜的结构和功能,导致其失去正常的细胞膜完整性和功能。
这种改变进一步抑制了真菌的生长和代谢活动。
本研究通过提取意大利苍耳的苍耳素,并对其体外抗植物病原真菌活性进行了研究。
结果显示,苍耳素对常见的植物病原真菌具有一定的抑制活性,并且这种抑制活性可能与其对真菌细胞膜的作用有关。
这些发现对于进一步开发苍耳素作为植物病原真菌的控制剂具有重要的指导意义,也为理解意大利苍耳的药用和生物活性提供了科学依据。
意大利苍耳提取物抗植物病原真菌活性研究
意大利苍耳提取物抗植物病原真菌活性研究
意大利苍耳(Cetraria islandica)是一种自然生长于北纬20-60度寒冷地区的苔藓植物,具有广泛的药用价值。
研究发现,意大利苍耳中含有丰富的活性成分,具有抗菌、抗病毒、抗氧化等多种生物活性。
为了研究意大利苍耳提取物的抗植物病原真菌活性,本研究采用经典的生物活性评价方法,包括抑菌圈实验和最小抑菌浓度(MIC)测定。
我们收集了不同地区的意大利苍耳样本,并进行干燥和粉碎处理。
然后,利用不同溶剂(如乙醚、乙醇、水)对样品进行提取,并得到相应的苍耳提取物。
接下来,我们选取了一些重要的植物病原真菌,如白菌、麦秆镰刀菌、黑曲霉等,对苍耳提取物进行抗菌活性测试。
结果显示,意大利苍耳提取物在不同溶剂中均表现出良好的抗菌活性。
根据抑菌圈实验,可以看出苍耳提取物对不同真菌菌株都具有一定的抑制效果。
我们进一步测定了苍耳提取物的最小抑菌浓度,结果表明苍耳提取物对大多数测试菌株都具有较低的最小抑菌浓度,表明其抗菌活性较高。
进一步的研究表明,苍耳提取物对植物病原真菌的抗菌活性可能与其所含的活性成分有关。
前期研究发现,意大利苍耳中含有多种生物活性成分,如苍耳多糖、苍耳酚类化合物等。
这些成分具有良好的抗氧化和抗菌活性,可能是苍耳提取物抗植物病原真菌活性的重要原因。
意大利苍耳提取物具有良好的抗植物病原真菌活性。
这为开发新型的植物抗菌剂提供了一个有前景的方向。
未来的研究可以进一步筛选和分离意大利苍耳中的活性成分,并探究其抗菌机制,为植物病害的防治提供新的思路和方法。
还需要进一步研究苍耳提取物的安全性和稳定性,以保证其在实际应用中的效果和效能。
基础微生物必记的拉丁学名
Staphylococcus aureus
苏云金芽胞杆菌
Bacillus thuringiensis
满江红鱼腥蓝细菌
Anabaena azollae
古菌
甲烷球菌属
Methanococcus
盐杆菌属
Halobacter
真菌
裂殖酵母属
Schizosaccharomyces
假丝酵母属
Candida
基础微生物必记的拉丁文学名
(注意:细斜体标记的微生物名称既要会写拉丁文学名又要会认;粗体标记的只要求会认拉丁文学名,会写中文学名。)
细菌
假单胞菌属
Pseudomonas
固氮菌属
Azotobacter
根瘤菌属
Rhizobium
蛭弧菌属
Bdellovibrio
脱硫弧菌属
Desulfovibrio
衣原体属
Chlamydia
支原体属
Mycoplasma
鱼腥蓝细菌属
Anabaena
土壤杆菌属
Agrobacterium
链球菌属
Streptococcus
乳杆菌属
Lactobacillus
芽胞杆菌属
Bacillus
梭菌属
Clostridium
链霉菌属
Streptomyces
大肠杆菌
Escherichia coli
毛霉属
Mucor
根霉属
Rhizopus
曲霉属
Aspergillus
青霉属
Penicillium
酿酒酵母
Saccharomyces cerevisiae
葡枝根霉(黑根霉)
Rhizopus stolonifer
猪胸膜肺炎放线菌分离鉴定与耐药性检测
文章编号:1673-887X(2023)05-0144-03猪胸膜肺炎放线菌分离鉴定与耐药性检测吕凤禄,赵欢(1.北镇市动物疫病预防控制中心(动物卫生监督中心),辽宁北镇121300;2.北镇市农业综合行政执法队,辽宁北镇121300)摘要为了分析某地区规模化养殖场及散养户中猪胸膜肺炎放线菌流行情况与对临床中常用抗菌药物耐药性情况,文章对2021年—2023年采集某地区规模化养殖场及散养户中疑似患猪胸膜肺炎放线菌病不同日龄猪的咽拭子、鼻拭子及死亡猪的肺脏组织170份,采用细菌分离培养、形态学观察、生化鉴定及PCR 方法对猪胸膜肺炎放线菌进行鉴定,采用K-B 药敏纸片法检测分离菌株的耐药性。
结果显示:从170份病料样品中分离到65株猪胸膜肺炎放线菌;分离的65株猪胸膜肺炎放线菌对新霉素、磺胺间甲氧嘧啶、庆大霉素等10种药物耐药率在50.8%以上,对头孢噻肟、头孢噻呋、环丙沙星等8种药物耐药率在6.2%~33.8%,为该地区仔猪源沙门菌病防控和合理用药提供参考。
关键词仔猪;沙门菌;分离鉴定;耐药性分析中图分类号S858.28文献标志码Adoi:10.3969/j.issn.1673-887X.2023.05.054Isolation Identification and Drug Resistance Detection of Porcine Pleuropneumonia ActinomycetesLyu Fenglu,Zhao Huan(1.Beizhen Animal Disease Prevention and Control Center (Animal Health Supervision Center),Beizhen 121300,Liaoning,China;2.Beizhen Agricultural Comprehensive Administrative Enforcement Team,Beizhen 121300,Liaoning,China)Abstract :In order to analyze the prevalence of porcine pleuropneumonia actinomycetes in large-scale farms and free-range house ‐holds in a certain area and their resistance to commonly used antibiotics in clinical practice,the paper collected 170throat swabs,nose swabs and lung tissues of dead pigs of different ages suspected to be infected with porcine pleuropneumonia actinomycetes in large-scale farms and free-range households in a certain area from 2021to 2023.The actinomycetes porcine pleuropneumoniae were identified by isolation and culture,morphological observation,biochemical identification and PCR.The drug resistance of the iso ‐lates was detected by K-B drug sensitive disk method.The results showed that 65strains of actinomyces pleuropneumoniae were iso ‐lated from 170samples.The resistance rates of 65strains of actinomyces pleuropneumoniae isolated to 10drugs such as neomycin,sulfamethoxil and gentamicin were above 50.8%,and the resistance rates of 8drugs such as cefotaxime,ceftiofur and ciprofloxacin were 6.2%~33.8%,providing reference for the prevention and control of salmonellosis of piglets in this area and rational drug use.Key words :piglet,salmonella,isolation and identification,drug resistance analysis在猪养殖过程中,不同日龄猪的呼吸道疾病成为临床中的常见疾病,对不同阶段猪的感染率和死亡率均较高,尤其是对仔猪的危害较大。
法氏囊毒株毒力鉴定标准
法氏囊毒株毒力鉴定标准英文回答:The virulence determination of Francisella tularensis strains involves various factors and criteria. One of the commonly used methods is the mouse LD50 (lethal dose 50%) assay, which measures the lethal dose required to kill 50% of the infected mice. This assay helps in comparing the virulence of different strains by determining theirrelative potency.Another important criterion for assessing the virulence of F. tularensis strains is the ability to cause disease in humans. This is determined through epidemiological studies and clinical observations. Strains that are associated with severe disease manifestations and high mortality rates are considered more virulent.In addition to these criteria, genetic analysis plays a crucial role in identifying virulence factors in F.tularensis strains. The presence of specific genes or genetic variations can contribute to the pathogenicity of the strain. For example, the F. tularensis subsp.tularensis strain, also known as Type A, is considered highly virulent due to the presence of certain genes that enhance its ability to evade the host immune response.Furthermore, the ability of F. tularensis strains to survive and replicate within host cells is another important factor in determining their virulence. Strains that can efficiently invade and multiply within host cells are more likely to cause severe disease. This ability is often assessed through in vitro cell culture studies or in vivo infection models.Overall, the virulence determination of F. tularensis strains involves a combination of factors such as mouseLD50 assay, clinical observations, genetic analysis, and host cell invasion studies. These criteria help in assessing the pathogenic potential of different strains and understanding their virulence mechanisms.中文回答:法氏囊毒株的毒力鉴定涉及多种因素和标准。
用于诊断路易体痴呆的遗传标记[发明专利]
专利名称:用于诊断路易体痴呆的遗传标记
专利类型:发明专利
发明人:K·拜尔,M·多明戈萨巴特,A·阿利茨费尔南德斯申请号:CN201180011143.7
申请日:20110224
公开号:CN102869786A
公开日:
20130109
专利内容由知识产权出版社提供
摘要:已经发现了BChE基因中的特异性改变,这种特异性改变允许确定患者是否患有路易体痴呆(DLB),并且使得可以将其与阿尔茨海默病区别开来。
本发明提供了用于诊断DLB的体外方法,包括在来自受试者的生物学样本中确定丁酰胆碱酯酶(BChE)基因的下列改变的基因型:NCBI登录号
NG_009031位置68974处的多态性位点(即SEQ ID NO:28中的位置934),和多态性位点3687、4206、4443、以及位置4780至4786的多聚胸腺嘧啶区,所述位置参照对应于人BChE基因核苷酸序列的NCBI登录号NG_009031(即分别为SEQ ID NO:1中的位置3687、4206和4443)。
申请人:巴塞罗那自治大学,日耳曼人特里亚斯艾普霍尔健康科学研究院基金会
地址:西班牙加泰罗尼亚
国籍:ES
代理机构:北京派特恩知识产权代理事务所(普通合伙)
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细菌鉴定方法
细菌鉴定方法English:The identification of bacteria involves a variety of methods, including morphological, biochemical, and genetic techniques. Morphological identification entails examining the physical characteristics of the bacteria, such as their shape, size, and arrangement under a microscope. Biochemical tests involve assessing the metabolic capabilities of the bacteria, such as their ability to ferment specific sugars or produce certain enzymes. Genetic techniques, such as polymerase chain reaction (PCR) and DNA sequencing, can be used to analyze the genetic makeup of the bacteria and compare it to known sequences in databases. Additionally, mass spectrometry can be employed to analyze the protein profiles of bacteria by ionizing and then measuring the mass-to-charge ratios of their molecules. Ultimately, a combination of these methods is often used to accurately identify and characterize bacteria in clinical, research, and industrial settings.中文翻译:细菌的鉴定涉及各种方法,包括形态学、生化学和遗传学技术。
中科院理化所 抗菌实验认证 英语
中科院理化所抗菌实验认证英语Title: Certification of Antimicrobial Experiment at Institute of Physical and Chemical Research, Chinese Academy of SciencesIntroductionInstitute of Physical and Chemical Research (IPC) is a leading research institution in China that focuses on the study of physical and chemical properties of various substances. Recently, IPC conducted a series of experiments to test the antimicrobial properties of certain compounds. The results of these experiments have been certified by independent authorities, establishing IPC as a reliable source for research in antimicrobial agents.Experimental ProcedureThe experiments conducted at IPC followed rigorous protocols to ensure accuracy and reliability. First, various compounds were selected for their potential antimicrobial properties. These compounds were then tested against different strains of bacteria and fungi to determine their effectiveness. The experiments were conducted in controlled environments to eliminate any external factors that could influence the results.Certification ProcessAfter the completion of the experiments, the results were sent to independent authorities for certification. The authorities reviewed the data and procedures used in the experiments to ensure that they met the standards for antimicrobial testing. Once the authorities were satisfied with the results, they issued a certificate of approval, confirming the accuracy and reliability of the experiments conducted at IPC.Significance of CertificationThe certification of the antimicrobial experiments conducted at IPC holds significant importance in the field of research. It establishes IPC as a reputable institution for studying antimicrobial agents and provides credibility to the results obtained from these experiments. Researchers and scientists can now rely on the information provided by IPC when conducting their own studies on antimicrobial properties of compounds.ConclusionThe certification of antimicrobial experiments at IPC by independent authorities solidifies the institution's reputation as a reliable source for research in antimicrobial agents. The rigorous procedures followed during the experiments and the approval from independent authorities demonstrate the accuracy and reliability of the results obtained. Researchers andscientists can now confidently refer to IPC for information on antimicrobial properties of compounds, further advancing research in this field.。
欧洲食品安全局动物饲料添加剂和饲料产品委员会关于动物营养中屎肠球菌的安全性评估指南
欧洲食品安全局动物饲料添加剂和饲料产品委员会关于动物营养中屎肠球菌的安全性评估指南
王黎文(译)
【期刊名称】《中国饲料》
【年(卷),期】2014(000)015
【摘要】本指南旨在为临床分离获得的从属于细菌亚群的屎肠球菌菌株提供一种鉴定方法。
区分的关键在于对氨苄青霉素是否敏感以及是否含有临床分离菌株所含的3个遗传标记物。
欧洲食品安全局动物饲料添加剂和饲料产品委员会认为,当屎肠球菌菌株对氨苄青霉素的抗性大于2 mg/L或含有上述3个遗传标记物任意其一时,均不得作为饲料添加剂使用。
【总页数】4页(P37-40)
【作者】王黎文(译)
【作者单位】中国饲料工业协会
【正文语种】中文
【中图分类】S816.17
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© 2007 Plant Management Network.Accepted for publication 20 December 2007. Published 19 April 2007.Detection of Candidatus Liberibacter asiaticus from Wampee (Clausena lansium Skeels) by Nested PCRXiaoling Deng, Gen Zhou, and Huaping Li, Laboratory of Citrus Huanglongbing Research, Department of Plant Pathology, South China Agricultural University, Guangzhou, Guangdong 510642, People's Republic of China; and Jianchi Chen and Edwin L. Civerolo, San Joaquin Valley Agricultural Sciences Center, Crop Diseases, Pests, and Genetics Research Unit, USDA-ARS, Parlier, CA 93648Corresponding author: Jianchi Chen. jichen@Deng, X., Zhou, G., Li, H., Chen, J., and Civerolo, E. L. 2007. Detection of Candidatus Liberibacter asiaticus from wampee (Clausena lansium Skeels) by nested PCR. Online. Plant Health Progress doi:10.1094/PHP-2007-0419-01-BR.Wampee (Clausena lansium Skeels) is native to southern China and Southeast Asia. Wampee trees are attractive, with grape-like fruits and a muscat taste and are popular in home gardens. Like other members of Rutaceae, wampee has long been suspected to have "yellow shoot" disease or Huanglongbing (HLB) and Diaphorina citri, the disease vector, was capable of a long-term survival on Wampee (2). The importance of wampee HLB is its potential to serve as a source of inoculum of the HLB pathogen, Candidatus Liberibacter spp., for sweet orange, mandarin, and other economically important citrus crops. Yet, the presence of Ca. Liberibacter spp. in wampee has not been confirmed with the 16S rDNA signature sequence that define this unculturable bacterium (1,3).In August of 2006, we identified two wampee trees showing "yellow shoots" symptoms (Fig. 1A) adjacent to a mandarin orchard with HLB in Luoding City, Guangdong Province, People’s Republic of China. Symptomatic leaves ranged from mottling and to yellowing (Fig. 1B). Leaf samples were collected, and DNA was extracted using the CTAB (cetyltrimethylammoniumbromide) method (4). DNA samples were assayed by nested-PCR. The general bacterial 16S rDNA primer set fDl/rD1 (AGA GTT TGA TCC TGG CTC AG / AAG GAG GTG ATC CAG CC) (1) was used for the outer round amplification. The PCR reaction (25; 100 µl) mixture contained:10 mM Tris-HCl, pH 8.3; 50 mM KCl; 1.5 mM MgCl2µM each dNTPs; 400 mM each primer, 1 U of Taq DNA polymerase and 1 µL of template DNA. Amplification was conducted with an initial denature at 96°C for 10 min, followed by 10 cycles consisting of: denaturing at 96°C for 30 s, annealing at 55°C for 30 s, and extension at 72°C for 30 s. Two µl of the amplicon were then used for inner round amplification using the same procedure but 35 PCR cycles with primer set OI1/OI2c (5' GCG CGT ATG CAA TAC GAG CGG CA 3' / 5' GCC TCG CGA CTT CGC AAC CCA T 3') specific for Ca. L. asiaticus (1,3). The amplified DNAs were resolved through agarose gel electrophoresis.As shown in Fig. 2, a 1.1 kb amplicon was obtained from symptomatic but not asymptomatic leaf samples. Non-nested PCR using primer set OI1/OI2c-only did not yield any detectable DNA bands from either symptomatic or asymptomatic leaves. Xba I digestion yielded two fragments of 520 bp and 640 bp, characteristic to Ca. L. asiaticus. PCR amplicons were further sequenced to be 1,095 bp and shared a > 98% similarity to sequences of Ca. L. asiaticus in the GenBank database. No DNA was amplified with primer set GB1/GB2 (AAG TCG AGC GAG TAC GCA AGT ACT / CCA ACT TAA TGA TGG CAA ATA TAG) specific to Ca. L. americanus (5). We note that nested-PCR is necessary for Ca. L. asiaticus detection in wampee, suggesting low bacterial titer in this host. We recommend that eradication of wampee trees surrounding citrus orchards should be part of the overall management of citrus HLB.Fig. 1. Huanglongbing symptoms in Clausena lansium : (A ) "yellow shoots"; (B ) comparison of symptomatic and asymptomatic leaves.Literature Cited1. Jagoueix, S., Bove, J. M., and Garnier, M. 1994. The phloem-limited bacterium of greening disease of citrus is a member of the alpha subdivision of the Proteobacteria. Int. J. Syst. Bacteriol. 44:379-386.2. Koizumi, M., Prommintara, M., and Ohtsu, Y. 1996. Wood apple, Limoniaacidissima: A new host for the huanglongbing (greening) vector, Diaphorina citri . Pages 271-275 in: Proc. 13th Conf. of the Intl. Organization of Citrus Virologists. J. V. da Graça, P. Moreno, and R. K.Yokomi, eds. Univ. of California, Riverside.3. Murray, R. G. E., and Stackebrandt, E. 1995. Taxonomic Note: Implementation of the provisional status Candidatus for incompletely described procaryotes. Int. J. Syst. Bacteriol. 45:186-187.4. Murray, M. G., and Thompson, W. F. 1980. Rapid isolation of high molecular weight plant DNA. Nucleic Acids Res. 8:4321-4324.5. Teixeira, Ddo C., Luc Danet, J., Eveillard, S., Cristina Martins, E., de Jesus Junior, W. C., Takao Yamamoto, P., Aparecido Lopes, S., Beozzo Bassanezi, R., Juliano Ayres, A., Saillard, C., and Bove, J. M. 2005. Citrus huanglongbing in Sao Paulo State, Brazil: PCR detection of the 'Candidatus ' Liberibacter species associated with the disease. Mol. Cell. Probes. 19:173-179.Fig. 2. Detection of Candidatus Liberibacter asiaticus by non-nested (usingprimer set OI1/OI2c-only) and nested PCR (using the general 16S rDNA primer set fDl/rD1 and then primer set OI1/OI2c) and results from Xba I digestion ofPCR amplicons.。