The Twin-Arginine Translocation （Tat） Pathway Is Essential for Virulence,Flagellation, and Chemotaxis in Xanthomonas oryzae pv. oryzicola Strain RsGD42

Bacterial leaf streak, caused by Xanthomonas oryzae pv. oryzicola, is an important disease of rice （Oryza sativa）. Genetic determinants （tatABC genes） of the twin-arginine translocation （Tat） pathway from X. oryzae pv. oryzicola strain RsGD42 were cloned and characterized, meanwhile, a tatC disruption mutant was generated. The tatC mutant lacked detectable flagella and was highly impaired in motility and chemotaxis. Furthermore, it was observed that the tatC mutant exhibited a reduced production of extracellular polysaccharide （EPS） and a significant reduction of virulence on adult rice plants compared to wild type strain. However, the tatC mutation in X. oryzae pv. oryzieola strain RsGD42 did not affect the growth rate and the ability to induce hypersensitive response （HR） in nonhost tobacco （Nicotiana tabacum L. cv. Samsun）. In conclusion, the data indicated that the Tat pathway significantly contributed to the virulence of X. oryzae pv. oryzicola.     

Identification of a Resistance Gene bls1 to Bacterial Leaf Streak in Wild Rice Oryza rufipogon Griff.

Bacterial leaf streak (BLS) of rice, caused by Xanthomonas oryzae pv. oryzicola (Xoc) is a worldwide destructive disease. Development of resistant varieties is considered to be one of the most effective and eco-friendly ways to control the disease. However, only a few genes/QTLs having resistance to BLS have been identified in rice until now. In the present study, we have identified and primarily mapped a BLS-resistance gene, bls1, from a rice line DP3, derived from the wild rice species Oryza rufipogon Griff. A BC₂F₂ (9311/DP3//9311) population was constructed to map BLS-resistance gene in the rice line DP3. The segregation of the resistant and susceptible plants in BC₂F₂ in 1:3 ratio (χ²=0.009, χ²_{0.05, 1}=3.84, P>0.05), suggested that a recessive gene confers BLS resistance in DP3. In bulked segregant analysis (BSA), two SSR markers RM8116 and RM584 were identified to be polymorphic in resistant and susceptible DNA bulks. For further mapping the resistance gene, six polymorphic markers around the target region were applied to analyze the genotypes of the BC₂F₂ individuals. As a result, the BLS-resistant gene, designated as bls1, was mapped in a 4.0-cM region flanked by RM587 and RM510 on chromosome 6.     

超表达OsPR1A增强了Xa21介导的水稻对白叶枯病的抗性反应

【背景】前期研究发现,水稻病程相关蛋白质OsPR1A的表达受上游抗病基因Xa21调控,接菌后早期启动Xa21介导的OsPR1A较高水平表达对水稻抵抗白叶枯病菌至关重要。同时OsPR1A也受到水稻白叶枯病菌（Xanthomonas oryzae pv. oryzae,Xoo）的诱导表达。对于OsPR1A的研究绝大部分是作为抗性反应发生的标志基因佐证其他基因或途径在抗性中的作用,缺乏直接的证据证实OsPR1A本身的生物学功能。【目的】通过获得OsPR1a-OX超表达转基因植株,调查其表型及农艺性状,并明确OsPR1A蛋白质表达与抗性的关系,为鉴定OsPR1A功能提供依据。【方法】通过农杆菌介导法,将构建的OsPR1a-OX转化载体转入到水稻受体4021中,利用PCR和免疫印迹（western blot,WB）技术分别在基因水平和蛋白质水平上筛选并鉴定OsPR1A超表达阳性纯合株系。在成熟期,调查OsPR1A超表达转基因植株的表型及农艺性状（株高、穗长、分蘖数、结实率和籽粒大小等）。在31℃条件下,将生长2周的水稻幼苗TP309、4021和OsPR1A超表达转基因植株接种水稻白叶枯病菌,并在接菌0、2、4、6、8、10和12 d时测量病斑长度。在接菌0、4和6 d时,收集TP309、4021和OsPR1A超表达转基因植株的水稻叶片,提取蛋白质,利用WB技术检测OsPR1A的表达特征。【结果】构建了OsPR1a-OX转化载体,并转入到受体4021中,筛选并鉴定到2个OsPR1A超表达转基因纯合株系（#704和#709）。调查了OsPR1A超表达转基因植株在成熟期的表型及农艺性状,与对照4021相比,#704和#709的株高较矮、穗长较短、分蘖数减少、结实率降低,但籽粒稍大,可能与结实率低有关。在31℃条件下,OsPR1A超表达转基因植株的病斑长度与对照4021相比明显缩短,结果具有显著性差异（P<0.05）。在接菌0、4和6 d的材料中,超表达转基因植株#704和#709中OsPR1A始终有较高水平的表达丰度,从而提高了对白叶枯病菌的抗性。【结论】采用农杆菌介导法,获得OsPR1A超表达转基因植株;超表达OsPR1A影响到水稻的正常发育过程;超表达OsPR1A后增强了Xa21介导的水稻对白叶枯病的抗性。     

Identification and Genetic Analysis of a Novel Rice Spotted-Leaf Mutant with Broad-Spectrum Resistance to Xanthomonas oryzae pv. oryzae

A spotted-leaf mutant of rice HM143 was isolated from an EMS-induced IR64 mutant bank. Brown lesions randomly distributed on leaf blades were observed about 3 wk after sowing. The symptom lasted for the whole plant growth duration. Histochemical analysis indicated that cell death occurred in and around the site of necrotic lesions accompanied with accumulation of hydrogen hyperoxide. Agronomic traits were largely similar to the wild type IR64 except seed setting rate and 1 000-grain weight which were significantly decreased in the mutant. Disease resistance of the mutant to multiple races of Xanthomonas oryzae pv. oryzae was significantly enhanced. Genetic analysis showed that the mutation was controlled by a single recessive gene, tentatively termed splHM143. In addition, using molecular markers and 1023 mutant type individuals from an F2 segregating population derived from the cross HM143/R9308, the spotted-leaf gene was finally delimited to an interval of 149 kb between markers XX25 and ID40 on the long arm of chromosome 4. splHM143 is likely a novel rice spotted-leaf gene since no other similar genes have been identified near the chromosomal region.     

水稻OsBBR1基因的白叶枯病抗性研究

水稻Y73是疣粒野生稻和栽培稻‘大粒香’经不对称体细胞杂交产生的后代,它对水稻白叶枯病表现为高抗。Y73在接种白叶枯病菌株P10 (PXO124) 后,一个NBS-LRR类基因的表达量显著上调。为了研究该基因的水稻白叶枯病抗性,采用RT-PCR技术从Y73中克隆了该基因的CDS片段 (2 631 bp),将该基因命名为OsBBR1(Bacterial Blight Resistance-related gene 1),并且利用pCAMBIA1300双元载体分别构建了OsBBR1基因超表达及干涉载体,采用农杆菌介导的转基因方法获得了转基因水稻植株。接菌试验显示,抑制OsBBR1基因的表达,能减弱抗病材料Y73的抗性;而提高OsBBR1基因的表达水平,则能增强感病材料的耐病性。结果表明OsBBR1基因与水稻白叶枯病抗性直接相关。此外,OsBBR1基因亚细胞定位结果还显示,OsBBR1编码产物主要定位于水稻细胞膜和细胞核,表明该基因很可能与信号转导和下游基因转录密切相关。     

Induction of Tolerance to Fusarium Wilt and Defense-Related Mechanisms in the Plantlets of Susceptible Berangan Banana Pre-lnoculated with Pseudomonas sp. （UPMP3） and Burkholderia sp. （UPMB3）

This study is aimed at assessing the ability of two endophytic bacteria originally isolated from healthy oil palm roots, Pseudomonas sp. （UPMP3） and Burkholderia sp. （UPMB3） to induce resistance in susceptible Berangan banana against Fusarium oxysporum race 4 （FocR4）. Increased accumulation of resistance-related enzymes such as peroxidase （PO）, phenylalanine ammonia lyase （PAL）, lignithioglycolic acid （LTGA）, and pathogenesis-related （PR） proteins （ehitinase and β-1,3-glucanase） has been observed in plantlets treated with endophytic bacteria UPMP3 and UPMB3 singly or as mixture under glasshouse conditions. Pre-inoculation of banana plantlets with UPMP3 showed a significant reduction in Fusarium wilt incidence 72 d after challenged inoculation with FocR4. UPMB3 was less effective in suppressing Fusarium wilt compared to UPMP3, whereas, the mixture of both endophytes showed an intermediate effect. Based on these results, it is concluded that UPMP3 could be a promising biological control agent that can trigger resistance against Fusarium wilt in susceptible Berangan banana.     

水稻白叶枯病菌新鉴别品种的筛选及其菌系的研究

 利用云南、日本和国际水稻研究所(IRRI)共19个稻白叶枯病鉴别品种,测定了来自云南不同地区的76个稻白叶枯病菌菌株。结果证明稻白叶枯病菌菌株与品种间具有特异性的交叉互作关系。.菌株间致病力的差异表现为质的差异。试验结果,初步以IR₂₆、Heen-Dikwee-1、珍白18、早生爱国3号、Tetep、窄叶青8号、IR₈和南粳33这8个品种作为新筛选的1套鉴别品种,其特点是品种与菌株具有明显的互作,鉴别能力强,可用来研究云南稻白叶枯病菌系。可将云南省36个稻白叶枯病菌株区分为22个菌系。     

Potassium sulphate induces resistance of rice against the root-knot nematode Meloidogynegraminicola

Potassium (K), an important nutrient element, can improve the stress resistance/tolerance of crops. The application of K in resisting plant-parasitic nematodes shows that the K treatment can reduce the occurrence of nematode diseases and increase crop yield. However, data on K₂SO₄ induced rice resistance against the root-knot nematode Meloidogynegraminicola are still lacking. In this work, K₂SO₄ treatment reduced galls and nematodes in rice plants and delayed the development of nematodes. Rather than affecting the attractiveness of roots to nematodes and the morphological phenotype of giant cells at feeding sites, such an effect is achieved by rapidly priming hydrogen peroxide (H₂O₂) accumulation and increasing callose deposition. Meanwhile, galls and nematodes in rice roots were more in the potassium channel OsAKT1 and transporter OsHAK5 gene-deficient plants than in wild-type, while the K₂SO₄-induced resistance showed weaker in the defective plants. In addition, during the process of nematode infection, the expression of jasmonic acid (JA)/ethylene (ET)/brassinolide (BR) signaling pathway-related genes and pathogenesis-related (PR) genes OsPR1a/OsPR1b was up-regulated in rice after K₂SO₄ treatment. In conclusion, K₂SO₄ induced rice resistance against M. graminicola. The mechanism of inducing resistance was to prime the basal defense and required the participation of the K⁺ channel and transporter in rice. These laid a foundation for further study on the mechanism of rice defense against nematodes and the rational use of potassium fertilizer on improving rice resistance against nematodes in the field.     

Reactive oxygen species are involved in cell death in wheat roots against powdery mildew

Inoculation of wheat(Triticum aestivum L.) leaves with wheat powdery mildew fungus(Blumeria graminis f. sp. tritici) induces the cell death in adventitious roots. Reactive oxygen species(ROS) play a key role in respond to biotic stress in plants. To study the involvement of ROS and the degree of cell death in the wheat roots following inoculation, ROS levels and microstructure of root cells were analyzed in two wheat cultivars that are susceptible(Huamai 8) and resistant(Shenmai 8) to powdery mildew fungus. At 18 d after powdery mildew fungus inoculation, only Huamai 8 displayed the leaf lesions, while root cell death occurred in both varieties. Huamai 8 had a high level of ROS accumulation, which is associated with increased root cell degradation, while in Shenmai 8, there was little ROS accumulation correlating with slight root cell degradation. The molecular study about the expression levels of ROS scavenging genes(MnSOD and CAT) in wheat roots showed that these genes expression decreased after the leaves of wheat was inoculated. The difference between Huamai 8 and Shenmai 8 on subcellular localization of H2 O2 and O2–· was corresponded with the different down-regulation of the genes encoding for superoxide dismutase and catalase in two wheat cultivars. These results suggested that ROS were involved in the process by which powdery mildew fungus induced cell death in wheat roots.     

杜仲内生拮抗细菌DZSY21诱导玉米抗病基因表达变化的转录组学研究

杜仲内生拮抗细菌DZSY21可在玉米中稳定定殖并增强玉米植株的抗病能力。试验从基因水平上对内生拮抗细菌DZSY21诱导玉米产生抗病性的分子机制进行预测,利用转录组测序技术对内生拮抗细菌DZSY21处理玉米后不同生长时间段叶片的总mRNA进行差异表达分析,以Fold change≥2且FDR<0.01为标准,挑选差异表达基因(DEGs)。结果显示:以处理0 h作为对照,处理12 h时有2 413个差异表达基因,其中上调基因1 278个,下调基因1 135个;处理24 h时有737个差异表达基因,其中上调基因538个,下调基因199个。在此基础上,根据DEGs功能注释分类,在差异表达基因中筛选与抗病相关的基因,最终获得267个抗病基因。处理12 h筛选得到218个基因,主要涉及脂质转移蛋白、MATE转运蛋白及LysM受体蛋白激酶等26个抗病途径;处理24 h获得71个基因,主要涉及异黄酮还原酶、纤维素合成酶、苯丙氨酸解氨酶、L-抗坏血酸过氧化物酶、GLK转录因子及ACC氧化酶等30个抗病途径,其中不同处理时间段内重复基因23个。上述结果表明,将杜仲内生拮抗细菌DZSY21引入玉米,可调节玉米叶片中抗病相关基因的表达,为进一步寻找抗病基因及其功能鉴定奠定了基础。     

玉米幼根接种禾谷镰刀菌后的转录组分析

为研究禾谷镰刀菌侵染后玉米幼根内转录组的变化情况,利用RNAseq对禾谷镰刀菌接种后6和18h的感病玉米自交系Y331分别进行转录组测序,分析接种后玉米幼根内差异表达的基因。结果表明:接种禾谷镰刀菌后,玉米幼根内共有5 153个基因差异表达;在接种后上调表达的聚类中,多种抗病过程相关的基因显著富集,包括苯丙烷类次生代谢物质合成过程的基因以及植物激素水杨酸(Salicylic acid,SA)、茉莉酸(Jasmonic acid,JA)和乙烯(Ethylene,ET)合成、响应及信号介导途径的基因;在下调表达的聚类中,植物生长发育、基础物质和能量代谢相关过程的基因显著富集。进一步分析发现:ET相关过程的基因在聚类1和聚类6中显著富集,均在接种后6h前上调表达;苯丙烷类次生代谢物合成途径相关基因和镰刀烯醇毒素(Deoxynivalenol,DON)解毒相关基因在接种后上调表达。研究表明,SA、JA/ET、苯丙烷类次生代谢物和DON解毒基因在玉米和禾谷镰刀菌互作中发挥重要作用。     

Genome-wide identification and expression analysis of GDSL esterase/lipase genes in tomato

The GDSL esterase/lipase family contains many functional genes that perform important biological functions in growth and development, morphogenesis, seed oil synthesis, and defense responses in plants. The expression of GDSL esterase/lipase genes can respond to biotic and abiotic stresses. Although GDSL esterase/lipase family genes have been identified and studied in other plants, they have not been identified and their functions remain unclear in tomato. This study is the first to identify 80 G...     

SlGH9-15 regulates tomato fruit cracking with hormonal and abiotic stress responsiveness cis-elements

Fruit cracking occurs easily during the late period of fruit development when plants encounter an unsuitable environment, dramatically affecting fruit production and marketing. This study conducted the bulked segregant RNA-Seq (BSR) to identify the key regulatory gene of fruit cracking in tomatoes. BSR-Seq analysis illustrated that two regions associated with irregularly cracking were located on chromosomes 9 and 11, containing 127 candidate genes. Further, through differentially expression analysis and qRT-PCR in cracking-susceptible and cracking-resistant genotypes, the candidate gene SlGH9-15 (Solyc09g010210) with significantly differential expression levels was screened. Bioinformatics analysis of the GH9 gene family revealed that 20 SlGH9 genes were divided into three groups. The phylogenetic analysis showed that SlGH9-15 was closely related to cell wall construction-associated genes AtGH9B1, AtGH9B6, OsGH9B1, and OsGH9B3. The cis-acting elements analysis revealed that SlGH9-15 was activated by various hormones (ethylene and ABA) and abiotic stresses. The expression pattern indicated that 13 SlGH9 genes, especially SlGH9-15, were highly expressed in the cracking-susceptible genotype. Its expression level gradually increased during fruit development and achieved maximum value at the red ripe stage. Additionally, the cracking-susceptible tomato showed higher cellulase activity and lower cellulose content than the cracking-resistant tomato, particularly at the red ripe stage. This study identified SlGH9-15 as a key gene associated with fruit cracking in tomatoes for the first time and gives new insights for understanding the molecular mechanism and complex regulatory network of fruit cracking.