大豆茎溃疡病的研究进展及其检疫意义

本文介绍了大豆茎溃疡病的发生与分布,危害,病原菌特征,侵染循环与传播途径等,重点介绍了该病的检测方法;初步分析了该病害传入我国及在我国定殖与传播的可能性,认为该病害对我国的大豆生产具有重要的检疫意义.     

湖南省柑橘溃疡病病菌生理生化特性初步研究

通过对湖南省境内分离的10个柑橘溃疡病菌株进行8项生化特性和17项生理检测,发现10个菌株在硝酸盐还原、H2S生成、淀粉酶、氧化酶、过氧化氩酶等生化特征上完全相同,但对醋酸盐、D-果糖、a-半乳糖、蔗糖、鼠李糖等的利用能力上有差异.通过对生理检测差异性进行聚类分析,发现湖南省不同柑橘产区的A菌系间存在一定的差异性.     

柑橘溃疡病菌检测方法及其在检疫中的优选

采用常规检测法、免疫斑点法、酶联免疫检测法和PCR检测法对柑橘溃疡病菌纯培养和柑橘叶片提取液进行平行测定。结果表明,对柑橘叶片提取液的检测,4种检测法检出的样品带菌率分别为11.12%、27.22%、18.69%、20.31%;对柑橘溃疡病菌纯培养的检测,4种方法检测结果一致。本实验同时比较了几种方法的灵敏度、特异性和稳定性,以及不同方法在各级检验检疫中的适应性。     

加强对柑橘溃疡病实施检疫的几点意见

柑橘是我国南方生产的重要水果,在生产发展中,加强对柑橘溃疡病的检疫,以保护无病县(市)和疫区内无病小区的柑橘生产免受柑橘溃疡病危害是十分重要的.现就柑橘溃疡病检疫中的几个问题提出下述意见,供参考.     

柑橘斑点病菌病及其检疫重要性

本文综述了柑橘斑点病菌病(Phaeoramularia angolensis)病原学、寄主、地理分布、形态特征及鉴定、生物学特性、传播途径及其检疫重要性。     

柑橘溃疡病菌的普通LAMP及快速LAMP检测方法的建立

建立柑橘溃疡病菌的普通LAMP和快速LAMP检测方法,使其能应用于基层检验检疫部门对病害的快速检测.利用柑橘溃疡病菌基因组特有的保守区域设计LAMP引物,通过优化反应条件,建立柑橘溃疡病菌的普通LAMP检测体系;在普通LAMP引物的基础上设计一对环引物,建立柑橘溃疡病菌的快速LAMP检测体系,并以多种参比菌DNA以及健康柑橘叶片基因组DNA为模板对普通LAMP和快速LAMP检测体系的特异性进行了验证,利用柑橘溃疡病菌菌液和DNA溶液梯度稀释液对普通LAMP和快速LAMP检测体系的灵敏度进行了验证.普通LAMP检测体系菌体和DNA检测灵敏度分别达到了2.25×104 cfu和2.03×10-1 ng,快速LAMP检测体系菌体和DNA检测灵敏度分别达到了2.25 cfu和2.03×10-5ng.在特异性测试中,普通LAMP检测体系与快速LAMP检测体系均仅对柑橘溃疡病菌进行扩增,对非靶标菌和柑橘叶片基因组DNA不产生扩增,普通LAMP与快速LAMP检测体系特异性测试结果一致.快速LAMP检测体系在0.5h内就可以达到普通LAMP检测体系的扩增量,是普通LAMP检测体系反应时间的一半,大大提高了检测的效率;快速LAMP检测体系菌悬液和DNA检测灵敏度均比普通LAMP检测体系提高了10 000倍.成功地建立了柑橘溃疡病菌的普通LAMP及快速LAMP检测方法,为柑橘溃疡病菌的检测提供了一种新的简便、快速的检测手段.     

柑橘溃疡病菌EMA-PCR快速活体检测技术的建立

传统PCR方法不能诊断柑橘溃疡病菌(Xanthomonas citri subsp.citri Gabriel)的死活状态,往往导致假阳性检测结果.本研究将特异性核酸染料叠氮溴化乙锭(ethidium monoazide bromide,EMA)与PCR技术结合,旨在建立柑橘溃疡病活菌的快速检测技术.根据柑橘溃疡病菌独有的保守蛋白基因设计特异性引物扩增出278 bp的靶带,PCR反应的检测下限为25个细胞/25 μL或2.75 pg/25 μL.EMA-PCR结果表明:当卤钨灯曝光时间1 min,EMA终浓度为1.0 mg/L时,能有效抑制1.0×108 cfu/mL死菌的扩增;当EMA的浓度小于30 mg/L时,EMA对上述相同浓度活菌靶基因的扩增没有明显的抑制.EMA-PCR对死活混合菌的扩增表明,活菌数在6.875×101～6.875×105 cfu/PCR范围时,荧光强度与混合体系中活菌的对数值有线性关系.基于以上建立的EMA-PCR活体检测技术,对疑似带病柑橘材料进行检测,结果发现能降低柑橘溃疡病菌检测过程中的假阳性,有望为柑橘溃疡病的检疫检验提供更科学的技术手段.     

柑橘溃疡病田间消长规律及药剂防治技术

多年田间系统观察表明 ,柑橘溃疡病的消长 ,在春梢期 ,主要影响因子为气温。 4月下旬平均气温 2 0℃持续 1 0～ 1 5d ,田间始病 ;平均气温 2 5℃连续 3d ,田间进入病情激增期。在夏、秋梢期 ,主要影响因子为湿度和降雨。相对湿度低于 70 % ,不利于发病 ;持续阴雨或短时大雨极有利于流行。溃疡病的药剂防治应掌握在春梢展叶后 7d用药 ,夏、秋梢则萌芽后即用药。药剂品种以可杀得和百菌清最理想 ,喷药次数以 3次为好。     

柑橘溃疡病菌的侵染过程及噻森铜对溃疡病的防治效果

为高效精准防治柑橘溃疡病,本研究对不同成熟度柑橘叶片接种溃疡病菌Xanthomonas citri subsp. citri后的发病情况进行观察,并在显微镜下观察溃疡病菌侵染叶片的过程,并对不同浓度、不同施用方法下噻森铜的室内防治效果和田间防治效果进行测定,对施用噻森铜后土壤、叶片和果实中的残留进行测定。结果表明,溃疡病菌可侵染柑橘嫩叶、幼叶和未成熟叶,其在叶片上附着2～4 d既已进入寄主组织内部。15 mL/株和20 mL/株噻森铜灌根15 d接种溃疡病菌的防治效果显著,而噻森铜10 mL/株灌根的防治效果略差,对照植株严重发病。萌芽前15～20 d噻森铜灌根浓度为60、75和90 mL/株时,每株病叶数分别为3.9、3.9和3.7个,每株病斑数分别为8.4、8.5和8.5个,病果率分别为9.4%、11.5%和9.6%,3个浓度处理之间差异不显著,但均显著低于对照。萌芽前喷施500倍噻森铜处理的每株病斑数为16.1个,与灌根60 mL/株噻森铜处理的每株病斑数差异不显著,但均显著低于对照;喷施噻森铜处理的病果率为16.8%,显著高于灌根60 mL/株噻森铜处理的,且两者均显著低于对照...     

柑橘溃疡病的发生及其综合防控

柑橘溃疡病是一种为害全世界柑橘种植业最严重的细菌性病害[1],并列入国内外检疫性病害。该病主要分布于亚洲、非洲、美洲、大洋洲及马来群岛的一些国家和地区。在我     

柑桔溃疡病菌PCR快速检验检疫技术研究

 柑桔溃疡病是严重影响全世界柑桔生产的重大检疫性病害,根据柑桔溃疡病菌(Xanthomonas axonopodis pv. citri)新近公布的全基因组中独有的保守蛋白基因序列,设计筛选出一对种特异性引物(JYF5/JYR5),能专一地扩增检出柑桔组织表面所带溃疡病菌的DNA靶带(413 bp)。而柑桔叶面附生的非致病性黄单胞菌、野油菜黄单胞菌近缘种以及健康柑桔样品都不能扩增;靶细菌DNA检测下限1.56 pg/μL,靶细菌悬浮液检测下限10 cfu/μL;在不同PCR仪及各种控温方式下都能稳定地扩增出特征性靶带。这一特异、准确的柑桔溃疡病菌PCR检验技术和研制的预包被固相化PCR检测试剂盒已开始用于我国非疫生产区建设中柑桔苗木、果实的病害检疫检验。     

Effect of temperature and leaf wetness duration on infection of sweet oranges by Asiatic citrus canker

Asiatic citrus canker, caused by Xanthomonas smithii ssp. citri , formerly X. axonopodis pv. citri , is one of the most serious phytosanitary problems in Brazilian citrus crops. Experiments were conducted under controlled conditions to assess the influence of temperature and leaf wetness duration on infection and subsequent symptom development of citrus canker in sweet orange cvs Hamlin, Natal, Pera and Valencia. The quantified variables were incubation period, disease incidence, disease severity, mean lesion density and mean lesion size at temperatures of 12, 15, 20, 25, 30, 35, 40 and 42°C, and leaf wetness durations of 0, 4, 8, 12, 16, 20 and 24 h. Symptoms did not develop at 42°C. A generalized beta function showed a good fit to the temperature data, severity being highest in the range 30–35°C. The relationship between citrus canker severity and leaf wetness duration was explained by a monomolecular model, with the greatest severity occurring at 24 h of leaf wetness, with 4 h of wetness being the minimum duration sufficient to cause 100% incidence at optimal temperatures of 25–35°C. Mean lesion density behaved similarly to disease severity in relation to temperature variation and leaf wetness duration. A combined monomolecular-beta generalized model fitted disease severity, mean lesion density or lesion size as a function of both temperature and duration of leaf wetness. The estimated minimum and maximum temperatures for the occurrence of disease were 12°C and 40°C, respectively.     

A comprehensive analysis of the Asiatic citrus canker eradication programme in São Paulo state,Brazil, from 1999 to 2009

Asiatic citrus canker (ACC), caused by Xanthomonas citri subsp. citri, has been controlled by exclusion and eradication measures in São Paulo state (SP), Brazil, since it was first detected in 1957. Several protocols of eradication have been used since then, but the protocol applied from 1999 to 2009 was the most successful and well documented of these. During that time, if the incidence of trees with symptoms in a citrus block was ≤0.5%, the infected trees and the symptomless trees within a 30 m radius were removed. However, if the incidence was >0.5%, the entire block was eliminated. Subsequently, inspections were carried out to monitor the affected blocks during quarantine. Although this protocol did not eliminate the disease from SP, it suppressed the incidence of affected blocks to very low annual levels (<0.20%) for a decade. Of over 5000 foci detected, 65.1% were eliminated by removing the block, either when disease was first detected (47.8%) or at a subsequent detection (17.3%), and 28.4% were eradicated by removing trees within a 30 m radius. The latter method was not an effective stand‐alone measure for ACC eradication and the 0.5% threshold was the key factor in suppression of disease statewide. Currently, a re‐emergence of ACC is occurring in SP. The information presented here will aid the establishment of protocols for the eradication of ACC in other citrus‐growing areas with low incidence of the disease or at risk of pathogen introduction.     

脐橙溃疡病综合防控技术规范

根据江西省赣南脐橙溃疡病的发生特点,对新老病区脐橙溃疡病提出了相应的综合治理对策。     

进境柠檬样品上柑桔溃疡病菌的检测

从进境柠檬样品上分离到一株疑似柑桔溃疡病菌的分离物X206,对分离物进行了菌落形态学特征观察、16S r RNA序列测定、PCR检测、Biolog测定及致病性测试。试验结果表明分离物X206在NA培养基上形成黄色,有光泽,圆形,全缘,微隆起,粘稠状的菌落。特异引物Xac01/Xac02扩增分离物X206的DNA得到582bp的预期产物,产物序列与柑桔溃疡病菌序列的相似性为100%。其16S r RNA序列与柑桔溃疡病菌的序列完全一致。Biolog测定将分离物X206和柑桔溃疡病菌阳性菌株FZ01均鉴定为Xanthomonas campestris pv.dieffenbachiae。致病性测试显示分离物X206能导致柑桔叶片产生明显的溃疡病斑。根据试验结果将分离物X206鉴定为柑桔溃疡病菌(Xanthomonas axonopodis pv.citri)。     

Effect of soil amendments, intercropping and planting time in combination on the severity of cassava bacterial blight and yield in two ecozones of West Africa

The severity of cassava bacterial blight at two sites in the forest-savanna transition (FST) and dry savanna (DS) zones of West Africa were studied by assessing the effects of: (i) shift of planting date; (ii) potassium fertilizer application and mulching; (iii) intercropping cassava with sorghum or cowpea vs. cassava monoculture; and (iv) the combination of these measures. Disease severity of bacterial blight in two genotypes was generally reduced by 20–60% by late planting, without a negative effect on cassava root yield, in monocropping systems in most treatments in the FST zone (reduction in four treatments, and increase in two treatments, out of 19) and the DS zone (two of eight treatments in 1 year). Late planting led to crop failure in the DS zone in the second year. Intercropping cassava with sorghum reduced bacterial blight severity significantly, up to 80% at normal (all treatments) and late planting time (three out of six treatments) in the FST zone, and in some treatments (four out of seven) at normal planting in the DS zone. Intercropping of cassava with cowpea in the DS zone also reduced disease severity. Cassava-sorghum intercropping generally had no effect on root yield compared with cassava monocropping at both planting times in the FST zone and provided an additional harvest of the intercrop, while yield was affected by intercropping in the DS zone at late, and in some treatments (three out of seven) at normal, planting time. Mulching and potassium treatment had no effect on disease severity, but increased or decreased root yield in some treatments in both sites. Analysis of combined data showed that cropping system, year, site, and site combined with planting date were the highest significant determinants of variation in bacterial blight development.     

Annual and polyetic progression of citrus canker on trees protected with copper sprays

The effects of copper sprays on annual and polyetic progress of citrus canker, caused by Xanthomonas citri subsp. citri, in the presence of the Asian citrus leafminer (Phyllocnistis citrella), were evaluated in a study conducted in a commercial orchard in northwest Paraná state, Brazil, where citrus canker is endemic. Nonlinear monomolecular, logistic and Gompertz models were fitted to monthly disease incidence data (proportion of leaves with symptoms) for each treatment for three seasons. The logistic model provided the best estimate of disease progress for all years and treatments evaluated and logistic parameter estimates were used to describe polyetic disease dynamics. Although citrus canker incidence increased during each of the seasons studied, it decreased over the whole study period, more so in copper‐treated trees than in water‐sprayed controls. Copper treatment reduced disease incidence compared with controls in every year, especially 2004–2005, when incidence was ca. 10‐fold higher in controls than in treated plots (estimated asymptote values 0·82 and 0·07, respectively). Copper treatment also reduced estimated initial disease incidence and epidemic growth rates every year.     

柑橘溃疡病菌免疫荧光和生物学检测技术研究

本研究通过对7个不同柑橘品种的离体叶片接种及致病性观察筛选出了柑橘溃疡病菌敏感寄主材料，并建立了柑橘溃疡病菌的生物学检测方法；还初次尝试建立了一种简便易行的柑橘溃疡病菌免疫荧光快速鉴定方法。柑橘离体叶片接种实验结果表明：福本、红肉脐橙、纽荷尔和枳壳对柑橘溃疡病菌表现出一定的感病性，而粗柠檬和邓肯葡萄柚未表现出明显感病迹象，反而出现了坏死性应激反应。免疫荧光试验结果显示：浓度为10^3cfu／mL的菌液在30℃下用BSA封闭2h，30℃下抗体结合1h，荧光抗体浓度为4μg,／mL，室温放置40min后镜检，就可以在荧光显微镜下清晰地看到绿色的柑橘溃疡病菌体，而用该方法检测不到水稻白叶枯病菌。用免疫荧光抗体检测柑橘溃疡病菌操作简便，仅需一台荧光显微镜，整个检测过程仅需要4h。     

The Effect of Irrigation Practices on the Spatio-temporal Increase of Asiatic Citrus Canker in Simulated Nursery Plots in Reunion Island

Asiatic citrus canker is a potentially severe disease of several citrus species and cultivars in many tropical and subtropical areas. In such areas, infected nursery plants constitute an important source of primary inoculum for newly established citrus groves. The influence of overhead, drip, and mist irrigation systems on the development of Asiatic citrus canker was studied in simulated, Mexican-lime nurseries in Reunion Island. Overhead irrigation exacerbated the increase of disease incidence and severity caused by a streptomycin-resistant strain of Xanthomonas axonopodis pv. citri. The temporal development of Asiatic citrus canker for overhead irrigated nursery plots was best described by an exponential model, because disease incidence in these plots did not come close to an asymptote during the experimental period. This can be explained by the continuous production of new growth, susceptible to infection by Xanthomonas axonopodis pv. citri, and splash dispersal of Xanthomonas axonopodis pv. citri associated with overhead irrigation. Based on spatial correlation and spatio-temporal analyses, aggregated disease patterns were found irrespective of the irrigation system. In overhead-irrigated plots, the spread of Xanthomonas axonopodis pv. citri lacked directionnality. Rainstorms of short duration and high intensity were apparently associated with disease increase in drip-irrigated plots. There is a need to improve cultivation practices in Reunion Island citrus nurseries to minimize Asiatic citrus canker incidence in nurseries and to minimize the introduction of Xanthomonas axonopodis pv. citri to new groves.