番茄细菌性斑点病菌实时荧光定量PCR检测方法的建立及应用

以番茄细菌性斑点病病原菌丁香假单胞菌番茄致病变种(Pseudomonas syringae pv. tomato,Pst)的致病相关基因HrpZ为靶序列,设计了特异性引物Pst3F/Pst3R,能从Pst基因组DNA中特异性扩增出大小为161 bp的目的片段。建立的Pst实时荧光定量PCR检测技术体系的检测灵敏度比普通PCR高1 000倍。利用实时荧光定量PCR检测体系,检测模拟带菌种子中Pst的带菌量,检测下限为4.21 cfu·g^-1;检测人工接种叶片组织中Pst的带菌量,检测到1级发病叶片带菌量为4.15×102 cfu·g^-1。对田间采集的63个番茄细菌性斑点病明显症状和疑似症状样本,分别进行了实时荧光定量PCR、普通PCR和病原菌分离检测,检测到54个样本中含有Pst,3种方法检测结果一致。结果表明,建立的Pst实时荧光定量PCR检测体系具有特异性强、灵敏度高的特点,可以快速准确地定量检测番茄种子和发病组织中Pst的含量,为番茄细菌性斑点病的早期预防和流行监测提供了有效的技术手段。     

四川省丁香假单胞猕猴桃致病变种的遗传多样性分析

为明确四川省猕猴桃溃疡病菌丁香假单胞菌猕猴桃致病变种(Pseudomonas syringae pv. actinidiae,Psa)的遗传多样性及群组划分与地理来源的相关性。以四川省不同地理来源的40个Psa菌株为研究试材,用rep-PCR技术进行分子标记,NTSYS软件分析UPGMA聚类。3对rep-PCR检测引物(REP、ERIC和BOX)共获得42个位点,其中38个为多态位点,占90.5%。UPGMA聚类结果显示,以相似系数为0.70阀值时,40个Psa菌株被分为2个类群(Ⅰ、Ⅱ),其中85.0%的菌株属于第Ⅱ类群;在相似性系数为0.80时,第Ⅰ类又可分为2个亚类(Ⅰ-1、Ⅰ-2),第Ⅱ类则分为5个亚类(Ⅱ-1、Ⅱ-2、Ⅱ-3、Ⅱ-4、Ⅱ-5),菌株无明显的采集地聚类。以上结果表明,四川省各地区的猕猴桃溃疡病菌菌株具有较高的遗传多样性,但其遗传变异与菌株地理来源无明显相关性。     

Predicting the potential distribution of Pseudomonas syringae pv. actinidiae in China using ensemble models

Pseudomonas syringae pv. actinidiae (Psa) is a causal agent of kiwifruit bacterial canker worldwide, which has affected kiwifruit vines in China since 1996 and has subsequently spread to the main cultivation areas. Based on occurrence of Psa and pseudo-absences randomly generated in China, the consensus-based modelling technique was used to estimate the spatial spread of Psa epidemics within China. Environmental variables that related to Psa development were identified, and their contributions to Psa development were evaluated. Three modelling algorithms, namely generalized boosting models (GBM), random forests (RF) and classification tree analysis (CTA) within the BIOMOD2 framework, were employed to construct the model. The ensemble models weighted by the true skill statistic (TSS) value were used to predict the current habitat suitability of Psa, and were projected using the four general circulation models (GCMs) to assess range shifts under two types of representative concentration pathways (RCP 4.5 and RCP 8.5) by 2050. The results indicated that precipitation in March and mean temperature of warmest quarter were the most important limiting factors for distribution of Psa. The predictive accuracy of the ensemble model showed acceptable predictive powers (TSS = 0.852). Under future climate conditions, substantial net loss of suitability for Psa was estimated to be 3.03–12.5% under RCP 4.5 (except one GCM), and 2.46–9.89% under RCP 8.5. Shrinkage of suitable habitats was detected mainly in the areas currently infected by Psa. Special attention should be given to recent infectious regions in south and southwest China, considering the locally expanding kiwifruit commercial plantations.     

几种杀菌剂对黄瓜细菌性斑点病及大白菜软腐病的抑制作用

采用抑菌圈法和含毒介质法测定几种杀菌剂对黄瓜细菌性斑点病和大白菜软腐病的抑制作用。结果表明,抑菌效果较好的为农用链霉素、新植霉素和水合霉素;丙森锌对黄瓜细菌性斑点病较好,而对大白菜软腐病则较差;氢氧化铜、噻菌铜及中生菌素抑制作用一般;春雷霉素抑菌活性较差。     

猕猴桃溃疡病病原菌的分子鉴定和致病力测定

为明确猕猴桃细菌性溃疡病致病菌种类与特征,以从安徽岳西、陕西户县和重庆黔江等猕猴桃主产区收集到的溃疡病感染枝条和树皮为材料,采用平板划线、梯度稀释和BPA培养法分离病原物,采用烟草过敏性反应和猕猴桃健康叶片接种观察其致病性,结合特异引物扩增的16S-23SrDNA-ITS序列分析,对病原菌种类进行分子鉴定。结果表明:从收集的溃疡病感染枝条和树皮中,共分离纯化得到10株分离物;所有分离物均可使烟草叶片产生过敏反应,猕猴桃叶片接种显示同一菌株对不同品种及不同菌株对同一品种的致病力均存在差异。对特异扩增得到的280bp 16S-23SrDNA-ITS序列进行Blast比对分析,结果表明:10个菌株为同一致病菌,其DNA片段大小和序列均与丁香假单孢杆菌猕猴桃致病变种(Pseudomonas syringae pv.actinidae)完全一致。据此可以确定,试验分离所得的菌株均为丁香假单孢杆菌猕猴桃致病变种。     

不同猕猴桃品种对溃疡病抗性的评价

离体条件下,通过不同致病力溃疡病病菌、接种浓度、培养湿度、接种方式条件下接种红阳猕猴桃枝条,探讨不同条件下溃疡病病原菌对猕猴桃枝条的侵染效果。在此基础上,通过对24个猕猴桃品种(系)枝条人工接种溃疡病病原菌,评价不同猕猴桃品种对溃疡病的抗性。结果表明: 强致病力病菌、高浓度、高湿度、枝条创伤面积都能显著影响猕猴桃枝条的病情指数;不同猕猴桃品种对溃疡病的抗性存在显著差异,24个猕猴桃品种(系) 枝条中,华特和徐香表现为高抗;迷你华特、金魁、绿肉优系(G HZ201201)和毛雄表现为抗病;红阳、黄肉优系11 7、大红、早鲜、早艳和黄肉优系Y HZ201201表现为高感。通过对华特、布鲁诺和红阳3个品种猕猴桃盆栽苗接种溃疡病病原菌,并测定接种植株与对照植株的相关生理参数,评价不同猕猴桃品种对溃疡病的抗性及其相关生理参数与抗溃疡病的关系。结果表明,3个品种盆栽苗的抗性评价等级与离体枝条接种基本一致。不同抗性猕猴桃品种酶活性与其病情指数呈线性关系,健康植株韧皮部中CAT酶活性与病情指数的相关性最高(r=0999*),叶片中的CAT酶活性次之(r=0994*),健康植株韧皮部和叶片中POD酶活性与病情指数的相关性分别为r=0979和r=0969,SOD酶活性分别为r=0924和r=0938。可溶性糖、可溶性淀粉、可溶性蛋白含量与病情指数也表现为线性关系,可溶性蛋白含量与病情指数相关性最高,在健康植株韧皮部和叶片的相关性分别为r=-0972和r=-0962,可溶性糖含量与病情指数的相关性分别为r=0934和r=0940。综上所述,不同猕猴桃品种对溃疡病的抗性存在显著性差异,CAT、POD、SOD酶活性和可溶性蛋白、可溶性糖含量可以作为鉴定猕猴桃抗溃疡病的生理指标。     

<Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">cerasicola</Emphasis>, pv. nov., the Causal Agent of Bacterial Gall of Cherry Tree

Bacterial gall on trunks and twigs of cherry trees (Prunus × yedoens, Someiyoshino) was found in Miyazaki and Saga prefectures, Japan. The surface of young galls are relatively smooth and light brown, but they become rough and dark brown. Characteristics of the bacterium isolated from galls on trunks or twigs are similar to those of Pseudomonas syringae pathovars, i.e., pv. actinidiae, pv. daphniphylli, pv. dendropanacis, pv. Morsprunorum, pv. myricae, pv. rhaphiolepidis, pv. syringae and pv. tremae. This bacterium produced galls on cherry and apricot, but not on 66 other species of plants belonging to 39 families. From these results, this bacterium was classified as a new pathovar of Pseudomonas syringae, and the name Pseudomonas syringae pv. cerasicola, pv. nov., is proposed. Strain M9501(ICMP 13926) was designated as the pathotype strain. Received 10 September 1999/ Accepted in revised form 24 December 1999     

烟草品种（品系）对野火病抗性评价

采用苗期针刺接种野火病菌的方法,对收集到的烟草品种（品系）进行抗性试验,结果表明,在38个测试的品种（品系）中,大体分为中抗、中感和感病3个抗性类群,其中,感病品种（品系）4个,该类群病情指数大于70;中感品种（品系）20个,该类群病情指数介于50～70;中抗品种（品系）14个,该类群病情指数介于20～50.在所有测试品种中,十里坪烟抗性最低,可作为对野火病监测预警材料;桦甸柳叶尖抗性最高,可作为今后的抗性亲本材料.     

烟草野火病病原细菌(Pseudomonas syringaepv.tabaci)的鉴定

本实验对黑龙江省烟草叶片上引起退绿晕圈症状的病原细菌从形态学、培养性状、生理生化反应、抗菌素反应、抗血清反应及寄生范围等几方面进行了鉴定.结果表明:该菌为烟草野火病菌〔Pseudomonas syringae pv.tabaci(Wolf&Foster(1917)Young,Dye&Wilkie(1978)〕.试验菌株在细菌学性状等方面与已有的报道无显著差异,但在致病性(寄主范围)等方面则明显不同.本文还报道了该菌的几个新寄主.     

吉林省大豆细菌性斑点病菌(Pseudomonas syringae pv.glycinea)生理小种鉴定结果初报

自1989年至1992年的四年间,在吉林省的长春、公主岭、吉林、白城、通化等地区的各大豆品种上分离到大豆细菌性斑点病菌的菌株146个,接种在7个国际鉴别寄主上,即Aeme,Chippwwa,Flambeau,Harosoy,Lindarin,Merit,Norchief,观察其抗感反应。将测定结果与国际上已鉴定的9个生理小种在这7个鉴别寄主上的反应相对照,发现有98个菌株属4号生理小种,占总菌株数的67.1％,为主要菌株;4个菌株属2号生理小种;1个菌株属7号生理小种。经反复测定,发现两个新的生理小种,定名为10号和11号生理小种。