Characterization of pathotypes among isolates of Xanthomonas axonopodis pv. manihotis in Colombia

Cassava bacterial blight, caused by Xanthomonas axonopodis pv. manihotis ( Xam ) is a destructive disease occurring in most cassava growing-areas. Although Colombian isolates of Xam differ in DNA polymorphism and pathogenicity, no suitable host differentials have been identified to demonstrate physiological specialization. A set of 26 Xam isolates from three edaphoclimatic zones (ECZs) in Colombia was selected for inoculation on a set of 17 potential cassava differentials. Leaf inoculation and stem puncture were used in order to detect possible specific interactions between cultivars and isolates. Cultivar × isolate interaction was highly significant ( P  < 0·001) after stem inoculation, but not after leaf inoculation. The stem inoculation technique was selected as a method for resistance screening of cassava cultivars for bacterial blight resistance. A highly significant interaction was also detected when cultivar behaviour was rated as area under the disease progress curve (AUDPC) after stem inoculation. Different pathotypes were defined among the 26 isolates and differential cultivars were proposed to define the pathotypic composition of Xam populations in three ECZs in Colombia. The results should help to improve selection of sources of resistance to cassava bacterial blight.     

水稻上三种条斑病细菌DNA的多态性初析

 试用40个引物对我国水稻条斑病菌、"稻短条斑病菌"和李氏禾条斑病菌等14个代表菌株进行RAPD分析,其中11个引物的扩增产物表现明显的多态性,共扩增出158条谱带,多态性为89.74%。聚类分析结果显示14个菌株可区分为3个类群,第1群包括LLS2、LLS3、LLS4、RS05、R1008、TAS和TAX;来自不同稻区水稻条斑病菌的群体结构差异明显,分属于第2群(如RS-Hai等)和第3群(如RS105等)。菌株DNA-RAPD指纹分析和致病性测定结果证明:李氏禾条斑病菌、"稻短条斑病菌"在水稻和李氏禾上不仅可相互侵染,而且遗传背景的相似性较高,确认是同一种病菌,其与小麦黑颖病菌亲缘关系较近,但与水稻细菌性条斑病菌具有明显差异。     

水稻细菌性条斑病种子带菌检测技术研究——Ⅰ.免疫放射分析法

免疫放射分析法(IRMA)集放射性同位素的灵敏性与抗原-抗体反应的特异性为一体。该方法应用于水稻细菌性条斑病种子带菌检测,经三年试验表明这一方法具有灵敏度高(最小检测浓度为10~3细菌/ml),特异性强,稳定性好,快速(4～5小时内可得出结果)。通过402份样品的检测,经与常规的浓缩接种法比较,证明该方法应用于带菌稻种的检测前景广阔。     

稻白叶枯病菌在人工培养基上毒性降低因素研究（英文）

 稻白叶枯病菌在人工培养基上毒性降低受多种因素的影响。试验结果表明,营养因素对菌的毒性降低比菌的不同保存方法更为明显,在供试的6种常用培养基中,NA 和 PSA 培养基对菌的毒性降低影响较小。较短的移植间隔期和频繁的移植次数也会影响菌的毒性降低。菌对低浓度氧的敏感性可因菌系而有差异。因此认为冰冻真空干燥和低温保存对抑制菌的毒性降低是较为理想的保存办法。试验还表明粘多糖的产生与菌的毒性之间不存在一定的相关性。     

稻白叶枯病菌外泌蛋白激发子的活性测定及其纯化

采用(NH4)2SO4分步沉淀法,从水稻白叶枯病菌的培养液中分离到具有激发子活性的物质。该蛋白处理水稻,可使水稻体内与抗病性相关的苯丙氨酸解氨酶和过氧化物酶活性显著提高;在田间喷洒水稻叶片,可以显著提高水稻对白叶枯病菌的抗性,病斑长度明显降低。经滚动式等电聚焦电泳和离子交换层析分离,生测结果表明第II吸收峰具有生物活性;再经SDS PAGE电泳和考马斯亮蓝染色,第II吸收峰收集物有明显的一条带,相对分子质量为47 9kD。     

水稻白叶枯病菌群体感应系统对T3SS基因表达的调控作用分析

 为了阐明水稻白叶枯病菌(Xanthomonas oryzae pv. oryzae, Xoo)利用DSF信号群体感应(QS)系统对毒性表达进行调控的机理,本研究对编码QS的rpf基因进行了分子鉴定, 分析了Δrpf基因突变体在水稻叶组织中的种群量变化, 利用RT-qPCR方法定量测定了编码T3SS的hrp基因转录本以及rpf基因自身表达。结果表明,rpfF、rpfC和rpfG基因与几种主要植物病原黄单胞菌同源序列高度保守;RpfF是烯脂酰辅酶A水合酶家族成员之一,RpfC含有组氨酸磷酸激酶结构域(HisKA)和磷酸受体结构域(REC),RpfG含有REC和HD-GYP结构域;Δrpf突变体在水稻叶组织中的种群量及其扩展能力均明显下降; hrp基因转录显著地受到QS调控;rpf基因表达与Xoo种群密度密切相关。因此,Xoo QS系统显著地调控了hrp基因的表达,在QS与T3SS表达之间存在一个信号通路。     

Isolation and Sequence of a Gene, celD Xo Involved in Cellobiose Utilization in Xanthomonas oryzae pv. oryzae

A genomic library of Xanthomonas oryzae pv. oryzae (X. o. pv. oryzae) T7174 was screened for 4-methylumbelliferyl β-D-glucoside-hydrolyzing (MUGase) activity. In subcloning of one of the MUGase-positive clones, an approximately 4.2-kb SacI-SphI fragment conferred not only MUGase activity but also 4-methylumbelliferyl β-D-cellobioside-hydrolyzing (MUCase) activity. Sequence analysis showed that the fragment contained an ORF of 2951 bp. The conceptual ORF product was significantly homologous with 1,4-β-D-glucan glucohydrolase D (CELD) from Pseudomonas fluorescens subsp. cellulosa, and was named CELD_Xo. Cell fractionation experiments suggested that CELD_Xo is localized in the cell-envelope fraction. We constructed a CELD_Xo-deficient mutant (74ΔCELD) from X. o. pv. oryzae. Little MUCase activity was detected in the cell-envelope fraction prepared from the mutant. The mutant 74ΔCELD did not grow in synthetic medium containing cellobiose as the sole sugar source. On the other hand, growth in rice leaves and pathogenicity of the mutant and the parental strain did not differ. These results suggested that CELD_Xo is involved in cellobiose utilization of X. o. pv. oryzae but that the gene is not required for bacterial growth in rice leaves. Received 16 February 2001/ Accepted in revised form 11 April 2001     

赣南脐橙溃疡病菌遗传多样性分析

本文根据已报道亚洲柑橘溃疡病菌基因组上的14个SSR位点,以分离自赣南18县（市）的508份柑橘溃疡病菌样品为材料分析赣南地区柑橘溃疡病菌遗传多样性。结果共选出7个位点用于赣南地区溃疡病菌种群多样性研究,Nei’s基因多样性指数介于0.66—0.83,表明赣南地区柑橘溃疡病菌具有丰富的遗传多样性。通过这7个位点分析赣南18县（市）不同地理来源的样品,发现大余县样品多态性较高,而定南县样品多态性较低。Structure 3.25和PowerMarker 2.3.4软件均发现赣南18县（市）的柑橘溃疡病菌样品分成两个组群,且其中某些县、区群体结构构成单一,信丰、定南和章贡区群体构成一组群,兴国、会昌、龙南和宁都群体构成另一组群,其他县区样品为两个组群菌株不同比例混合构成。本研究通过SSR分子标记的方法探索赣南地区柑橘溃疡病菌遗传多样性,为研究柑橘溃疡病的发生和流行规律提供有价值的线索。     

Cloning of a Genomic DNA Encoding Caffeoyl-coenzyme A 3-<Emphasis Type="Italic">O</Emphasis>-methyltransferase of Citrus

In recent studies, we found that Apll (a PthA homologue) bound to three citrus proteins. Amino acid sequence analysis revealed that one of the target proteins was homologous to that of S-adenosyl-l-methionine : trans-caffeoyl-coenzyme A 3-O-methyltransferase (CCoAMT), an enzyme which is specific for the substrate trans-caffeoyl-CoA and catalyzes the synthesis of trans-feruloyl-CoA. From the consensus nucleotide sequences of CCoAMT genes, primers were chosen for PCR amplification of this gene from citrus total DNA. Two selected DNA fragments of 1.0 kb and 2.0 kb were obtained. These fragments were used as the probe to screen a citrus library. One clone, pCCl00, contained a 1.0-kb SalI fragment that hybridized to the probes. The nucleotide sequence of this fragment was determined in both directions. In this fragment, there was an open reading frame of 232 amino acids interrupted by an intron of 106 nt, and the deduced amino acid sequence had 95.9% homology to tobacco CCoAMT. Southern blot analysis of total citrus DNA showed that four EcoRI fragments hybridized to the probes, suggesting the presence of more than one copy of CCoAMT in the citrus DNA. Received 4 November 1999/ Accepted in revised form 26 November 1999