水稻白叶枯病菌的母株与单细胞系的遗传与致病性差异的分析

由水稻白叶枯病菌(Xanthomonas oryxae pv.oryxae)引发的稻白叶枯病是水稻生产上的重要病害.水稻白叶枯菌自然群体是由包括基本无毒性的弱毒菌在内的不同致病型组成的混合群体,代表自然群体的原始菌株的致病力与其毒力结构紧密相关.     

中国、日本和菲律宾水稻白叶枯病菌遗传多样性比较分析

用IS-PCR和Rep-PCR对19个来自中国、日本和菲律宾的水稻白叶枯病菌(Xanthomonas oryzae pv. oryzae)群体进行遗传多样性分析。4个专化引物中的IS1113和ERIC能较好的区分三国水稻白叶枯病菌。UPGMA聚类结果表明, 三国菌株主要呈现第2簇和第3簇遗传型;中国和菲律宾菌株在第2簇和第3簇遗传型基础上有各自的特异性分化。病原菌的遗传分簇与致病群之间没有相关性。     

中国、日本和菲律宾水稻白叶枯病菌遗传多样性比较分析

用IS-PCR和Rep-PCR对19个来自中国、日本和菲律宾的水稻白叶枯病菌(Xanthomonas oryzae pv.oryzae)群体进行遗传多样性分析.4个专化引物中的IS1113和ERIC能较好的区分三国水稻白叶枯病菌.UPGMA聚类结果表明,三国菌株主要呈现第2簇和第3簇遗传型;中国和菲律宾菌株在第2簇和笫3簇遗传型基础上有各自的特异性分化.病原菌的遗传分簇与致病群之间没有相关性.     

水稻白叶枯病菌基因XOO2193的突变体构建及其毒力和胞外多糖分析

水稻白叶枯病菌是一种引起水稻白叶枯病的植物病原细菌,水稻白叶枯病是世界水稻生产中最严重的细菌性病害之一.本研究采用携带同源序列的自杀质粒pK18MobGⅡ整合的办法构建了水稻白叶枯病菌中国菌株13751编码6-磷酸葡糖酸内酯酶的基因XOO2193的非极性突变体GNM2193.对突变体的表型分析发现其毒力在杂交水稻品种特优63上显著减弱,突变体在非寄主植物蓖麻上不能引起过敏反应.此外,突变体胞外多糖的产量是野生型的43.4%.用一段含有XOO2193基因的DNA 片段对GNM2193进行功能互补,互补菌株在水稻上的毒力、引起过敏反应的能力和胞外多糖产量恢复到野生型水平.说明XOO2193基因与病菌的毒力和胞外多糖的产生有关.     

水稻与白叶枯病菌互作机制研究进展

水稻白叶枯病由Xanthomonas oryzae pv.Oryzae(Xoo)致病菌引起,为水稻三大病害之一,对世界水稻生产造成了严重危害.水稻与Xoo互作符合“基因对基因”假说,是研究植物与细菌互作的典型模式系统.水稻基因组以及Xoo基因组测序的完成,极大地推动了水稻-Xoo互作分子机理的研究.就有关水稻与Xoo互作机制的最新研究进展作一概述.     

水稻白叶枯病菌和水稻细菌性条斑病菌的实时荧光PCR快速检测鉴定

成功建立了水稻白叶枯菌与水稻细菌性条斑病菌快速检测鉴定的实时荧光PCR方法。根据含铁细胞接受子基因设计两菌的通用引物PSRGF/PSRGR（扩增一个152bpDNA片段）和特异性探针（Baiprobe和Tiaoprobe）,并对13种细菌和1种植原体进行实时荧光PCR。结果表明,两个特异性探针能分别特异性检测到目标病原菌产生荧光信号而其它参考菌不产生荧光信号。检测的绝对灵敏度是30.6fg/μL质粒DNA和103CFU/mL的菌悬浮液,相当于1个细菌细胞的基因,比常规PCR电泳检测高约100倍,相对灵敏度为105CFU/mL。整个检测过程只需2h,完全闭管,降低了污染的机会,无需PCR后处理。 用这两个特异性探针分别对自然感染白叶枯菌和条斑菌的叶片DNA提取液和种子浸泡液进行实时荧光PCR,结果均可特异性检测到目标菌的存在并完全可将两种病原细菌区分开来,且只需03g叶片和10g种子。     

水稻白叶枯病菌ahpC基因转录单元分析和原核表达

从水稻白叶枯病菌(Xanthomnas oryzae pv.Oryzae,Xoo)菌株PXO99A中克隆了H2O2降解基因ahpC,发现其编码的烷基过氧化氢酶AhpC在所测定的不同种病原细菌中的蛋白序列高度保守;采用RT-PCR方法分析了基因的转录结构特征,发现ahpC基因与酶电子供体基因ahpF组成了同一个转录单元;通过对ahpCp-lacZ活性检测,发现该启动子活性显著地受转录调控因子OxyR的正调控.此外,利用表达载体pET-28a(+)对ahpC基因进行了原核表达,经诱导后获得了可溶性的靶蛋白,可用于后续的生物学功能的分析.     

抗水稻白叶枯病菌植物内生真菌的筛选、鉴定

从多种植物叶片内分离、纯化出31株植物内生真菌,活性筛选表明分离自健康水稻叶片内生真菌Bo-1菌的乙酸乙酯粗提物对水稻白叶枯病菌具有较强的抑菌活性。当供试浓度为100μg/mL时,Bo-1菌乙酸乙酯粗提物对水稻白叶枯病菌抑菌圈为22.4 mm,与同浓度的氯霉素对水稻白叶枯病菌的抑菌效果相当。通过形态学特征观察和ITS rDNA测序分析,Bo-1菌被鉴定为淡色生赤壳菌(Bionectria ochroleuca)。抗菌谱实验表明,Bo-1菌对多种细菌类病原体具有较强的拮抗能力。     

水稻白叶枯病菌受寄主诱导启动子的克隆

用一个具链霉素（Ｓｍ）抗性,含无启动子ＣＡＴ基因的广宿主启动子探针质粒ｐＩＪ３１００．用鸟枪法克隆水稻白叶枯病菌染色体ＤＮＡ的ＢａｍＨ１酶切片段,重组质粒转化大肠杆菌ＥＤ＿（８７６７）,在含链霉素和氯霉素的ＬＢ平板上筛选含有水稻白叶枯病菌启动子活性片段的转化子。在帮助质粒ｐＲＫ２０１３的帮助下,通过三亲交配,将转化子中重组质粒转移进野生型的水稻白叶枯病菌中去,在分别含有链霉素和氯霉素的ＰＳＡ对照平板上筛选对热氯霉素敏感的接合子。随机挑取２００个该接合子,接种用氯霉素处理的水稻感病品种金南风,得到１５个比对     

白叶枯病菌侵染水稻体细胞无性系变异植株叶片的扫描电镜观察

选用离体筛选的抗白叶枯病水稻体细胞无性变异系后代植株的种子,收获后播种。对其功能叶用白叶枯病菌菌株浙１７３剪叶接种７２ｈ后,对叶片进行扫描电镜观察。结果发现,筛选的抗性品种的水孔周围病菌较少,而感病品种的水孔周围病菌则分布较多。证明了白叶枯病菌的侵染途径首先是通过水孔进入植物叶片内部的;另外也证明了筛选的水稻体细胞无性变异系后代植株已具有成株抗性。     

Status of Streptomycin Resistance Development in Xanthomonas oryzae pv. oryzae and Xanthomonas oryzae pv. oryzicola in China and their Resistance Characters

Rice leaves with bacterial blight or bacterial leaf streak symptoms were collected in southern China in 2007 and 2008. Five hundred and thirty‐four single‐colony isolates of Xanthomonas oryzae pv. oryzae and 827 single‐colony isolates of Xanthomonas oryzae pv. oryzicola were obtained and tested on plates for sensitivity to streptomycin. Four strains (0.75%) of X. oryzae pv. oryzae isolated from the same county of Province Yunnan were resistant to streptomycin, and the resistance factor (the ratio of the mean median effective concentration inhibiting growth of resistant isolates to that of sensitive isolates) was approximately 226. The resistant isolate also showed streptomycin resistance in vivo. In addition to resistant isolates, isolates of less sensitivity were also present in the population of X. oryzae pv. oryzae from Province Yunnan. However, no isolates with decreased streptomycin‐sensitivity were obtained from the population of X. oryzae pv. oryzicola. Mutations in the rpsL (encoding S12 protein) and rrs genes (encoding 16S rRNA) and the presence of the strA gene accounting for streptomycin resistance in other phytopathogens or animal and human pathogenic bacteria were examined on sensitive and resistant strains of X. oryzae pv. oryzae by polymerase chain reaction amplification and sequencing. Neither the presence of the strA gene nor mutations in the rpsL or rrs were found, suggesting that different resistance mechanisms are involved in the resistant isolates of X. oryzae pv. oryzae.     

Pathotypic variation of Xanthomonas oryzae pv. oryzae in Bangladesh

Bacterial Blight (BB) caused by Xanthomonas oryzae pv. oryzae (Xoo), a destructive disease of rice. Altogether, 96 isolates of Xoo were collected from 19 rice growing districts of Bangladesh in irrigated and rainfed seasons during 2014 to assess pathotypic variation. Pathotypic analyses on a set of 12 Near Isogenic Lines (NILs) of rice containing resistance genes viz. Xa1, Xa2, Xa3, Xa4, Xa5, Xa7, Xa8, Xa10, Xa11, Xa13, Xa14 and Xa21 and two check varieties IR24 and TN1 by leaf clip-inoculation technique. A total of 24 pathotypes were identified based on their virulence patterns on NILs tested. Among these, pathotypes VII, XII, and XIV considered as major, containing maximum number of isolates, (9.38% each) frequently distributed in North to Mid-Eastern districts of Bangladesh. Most virulent pathotype I recorded from Habiganj and Brahmanbaria. This pathotypic variation explained the pathogenic relatedness of X. oryzae pv. oryzae populations from diverse geographic areas in Bangladesh.     

Involvement of Gluconeogenic Pathway in Virulence of Xanthomonas oryzae pv. oryzae

Xanthomonas oryzae pv. oryzae causes bacterial leaf blight, one of the most widespread and destructive bacterial diseases of rice. A phosphoenolpyruvate synthase (ppsA)‐disrupted mutant OSPAM was generated by homologous suicide plasmid integration. The mutant was unable to grow in medium with pyruvate or C₄‐dicarboxylates as the sole carbon source, compared with the wild‐type, indicating a disruption in ppsA function. The mutant showed a reduction in virulence on rice but still induced a hypersensitive response in tobacco. When the mutant was complemented, the response was recovered to wild‐type. These results suggested that X. oryzae pv. oryzae possesses only PPSA route in gluconeogenesis, which is necessary for virulence.     

High-quality mutant libraries of Xanthomonas oryzae pv. oryzae and X. campestris pv. campestris generated by an efficient transposon mutagenesis system

A novel transposon mutagenesis system for the phytopathogenic bacteria Xanthomonas oryzae pv. oryzae (Xoo) and X. campestris pv. campestris (Xcc) was developed using a Tn5-based transposome. A highly efficient transformation up to 10(6) transformants per microg transposon DNA was obtained. Southern blot and thermal asymmetric interlaced polymerase chain reaction analyses of Tn5 insertion sites suggested a random mode of transposition. The transposition was stable in the transformants for 20 subcultures. Eighteen thousand and 17000 transformants for Xoo and Xcc, respectively, were generated, corresponding to 4X ORF coverage of the genomes. The libraries will facilitate the identification of pathogenicity-related genes as well as functional genomic analysis in Xoo and Xcc.     

Rapid inhibition of protein histidine phosphorylation by UV-irradiation in Xanthomonas oryzae pv. oryzae

Abstract Exposure of Xanthomonas oryzae pv. oryzae cells to 254 nm UV radiation resulted in an alteration of protein phosphorylation. Labelling of the phosphohistidine-containing proteins with molecular masses of 81 and 32 kDa, named p81 and p32, was rapidly reduced following UV irradiation in the early exponential cells, but the decrease was not detected in mid-exponential cells. Mitomycin C, a DNA replication inhibitor, and rifampicin, a drug generally used to inhibit RNA synthesis and DNA replication, were also found to reduce the histidyl phosphorylation. However, this alteration of protein phosphorylation was not hindered by chloramphenicol treatment. A possible role for these histidyl phosphopfoteins in sensing UV light is proposed.