Serological detection of Fiji disease virus antigens in the planthopper Perkinsiella saccharicida and its inefficient ability to transmit the virus

Individual adult Perkinsiella saccharicida reared on sugarcane infected with Fiji disease virus (FDV) were confined on healthy plants for 7 days to test for virus transmission and were then sacrificed and assayed for the presence of FDV-specific structures by immunoelectron microscopy. Only 15% of the insects contained FDV antigens and 6% transmitted the virus. FDV-specific antigens were detected in individual planthoppers by immuno-osmophoretic and enzyme-linked immunosorbent assays about as efficiently as by immunoelectron microscopy. The possible reasons for the inefficient acquisition and transmission of FDV by P. saccharicida are discussed.     

Improved method for assaying maize plant resistance to maize rough dwarf disease by artificial inoculation and real-time RT-PCR

The study of maize rough dwarf disease (MRDD) and breeding for resistance requires inoculation of maize plants by means of planthoppers. The plant age, insect density and inoculation duration are main factors in the success of maize rough dwarf disease inoculation. These parameters were tested using a susceptible maize inbred line Ye478. Using one or two-leaf plants, 15 planthoppers per plant and a five day inoculation duration, the line Ye478 was the most susceptible with 100% diseased plants; F112132 was moderately susceptible with 60% diseased plants and 90110 and F022411 were resistant without any disease. The results were consistent with those from six years of field studies. Using enzyme-linked immunosorbent assay (ELISA) and real-time quantitative RT-PCR, rice black-streaked dwarf virus was detected in severely diseased plants. The plants were rated from 0 to 3 according to their symptoms at the time of flowering. Plants scoring 0, 1 and 2 could not be distinguished by ELISA, only by real-time quantitative RT-PCR. All of the plants with a score of 3 were positive by ELISA and real-time quantitative RT-PCR. The significant differences in the average viral contents in plants with different symptom ratings could be distinguished by using real-time RT-PCR.     

低纬高原甘蔗白叶病植原体传播方式测定分析

甘蔗白叶病(Sugarcane white leaf,SCWL)是由16SrXI组植原体引起的甘蔗重要病害[1],主要通过带病蔗种传播,还可在田间通过叶蝉自然传播[2,3].该病1954年首次在泰国发现[4],现已在印度、巴基斯坦、斯里兰卡、菲律宾、缅甸、越南等国普遍发生,给当地蔗糖业造成巨大经济损失[5,6].201...     

Movement of aphid-transmitted Sugarcane yellow leaf virus (ScYLV) within and between sugarcane plants

Sugarcane yellow leaf virus (ScYLV) is distributed worldwide and has been shown to be the cause of the disease sugarcane yellow leaf syndrome (YLS). This study was an investigation of the transmission and spread of ScYLV in Hawaii. Several aphids are known to transmit the virus, but investigation of infestation and transmission efficiency showed Melanaphis sacchari to be the only vector important for field spread of the disease. The initial multiplication of ScYLV in a virus-free plant occurred exclusively in very young sink tissues. When a single leaf was inoculated on a plant, that leaf and all older leaves remained virus-free, based on tissue-blot immunoassay, whereas meristems and all subsequently formed new leaves became infected. Therefore, only after those leaves which had already developed before inoculation had been shed, did the complete plant contain ScYLV. Spread of the viral infection to neighbouring plants in the plantation fields via aphids was relatively slow and in the range of a few metres per year. No indication of long-distance transfer could be seen. This indicates that it may be possible to produce and use virus-free seed cane for planting of high-yielding but YLS-susceptible cultivars.     

甘蔗病毒病及其防治研究进展

甘蔗是最重要的糖料作物,由于其栽培过程中采用种茎无性繁殖,病毒病发生逐年加重.已知侵染甘蔗的病毒种类有甘蔗花叶病毒(Sugarcane mosaic virus,SCMV)、高粱花叶病毒(Sorghummosaic virus,SrMV)、甘蔗线条花叶病毒(Sugarcane streakmosaic virus,SCSMV)、甘蔗黄叶病毒(Sugarcane yellow leaf virus,SCYLV)、甘蔗斐济病病毒(Sugarcane Fiji disease virus,SFDV)、甘蔗线.条病毒(Sugarcane streak virus,SSV)和甘蔗杆状病毒(Sugarcane bacilliform virus,SCBV).文中简要介绍上述几种病毒的基本特性及其所致病害的发生特点,对目前甘蔗病毒病防治技术进行了评述,提出了我国甘蔗病毒研究中需要关注的若干问题.     

广东甘蔗黄叶病田间调查及病原病毒的分子检测

 广东省粤北和粤西蔗区多个县市的田间甘蔗上观察到甘蔗黄叶病(Sugarcane yellow leaf disease,SYLD)典型症状,目前该病仅局部分布,但部分田块病株率为5%~80%,发病品种有青皮果蔗、黑皮果蔗、新台糖系列品种、粤糖79/177和粤糖93/159等。采集发病田间显症叶片、无症叶片和在病叶上取食的甘蔗绵蚜(Ceratovacuna lanigera)样品,抽提总RNA,以基于甘蔗黄叶病毒(Sugarcane yellow leaf virus,SCYLV) CP基因序列的特异引物进行一步RT-PCR和巢式PCR扩增,并对扩增产物进行核苷酸序列测定和BLAST比对。结果显示,RT-PCR及巢式PCR产物核苷酸序列与分离自巴西的SCYLV B1株系相应区段同一率为100%;一步RT-PCR可从约70%的显症叶片样品中检测到SCYLV,而病田中的无症叶片样品以及在病叶上取食的单头甘蔗绵蚜样品需经巢式PCR扩增方可检测到SCYLV,阳性率分别为1%~5%和83%。本研究表明,广东省栽培甘蔗已受到SCYLV侵染,甘蔗绵蚜携带SCYLV。     

Aerial Contamination of Sugarcane in Guadeloupe by Two Strains of Xanthomonas albilineans

Two sugarcane plots were set up in Guadeloupe with disease-free tissue cultured plants in a banana growing location distant from sugarcane fields. Thirteen weeks after planting sugarcane in the field, a Xanthomonas albilineans strain belonging to serotype 3 (strain XaS3) was detected in water sampled at sunrise on the leaves in the first plot. This strain randomly invaded the sugarcane canopy. Seven weeks later, a new strain belonging to serotype 1 (strain XaS1) appeared on leaves and populations of strain XaS1 progressively increased on the leaf surface, whereas populations of strain XaS3 progressively decreased. Leaf scald symptoms were first noted 26 weeks after sugarcane planting. However, only strain XaS1 was isolated from leaves and a few sugarcane stalks showing symptoms. Both strains also colonized the second field plot, which was studied at the end of the experiment to avoid human interference of aerial contamination of sugarcane. After inoculation of three sugarcane cultivars by the decapitation technique, strain XaS1 was as virulent or more virulent than five other strains of X. albilineans isolated from diseased sugarcane plants in Guadeloupe. Although strain XaS3 colonized a few stalks, it failed to produce any symptoms and was the least virulent strain. Leaf surface colonization by X. albilineans was reproduced in a greenhouse trial by spraying the pathogen on sugarcane foliage. After 8 weeks, the pathogen was isolated from disinfected leaf blades. Although the leaf scald pathogen is thought to be mainly transmitted by infected cuttings, aerial transmission of X. albilineans is also known to occur. These results indicate the importance of sugarcane phyllosphere colonization by virulent strains in the epidemiological cycle of leaf scald disease in Guadeloupe.     

Spread of <Emphasis Type="Italic">Sugarcane yellow leaf virus</Emphasis> in initially disease-free sugarcane is linked to rainfall and host resistance in the humid tropical environment of Guadeloupe

Sugarcane yellow leaf virus, the causal agent of yellow leaf, is transmitted from plant to plant by aphids. Understanding and evaluating the epidemic risks due to spread of yellow leaf by aphids is an important feature for sugarcane production. Four distinct sugarcane trials were set up with disease-free plants to study the relationship between spread of yellow leaf, the vector dynamics and environmental conditions that may favour yellow leaf epidemics. The study was performed by surveys of vector populations and determination of plant infections. Sugarcane cultivar SP71-6163, highly susceptible to yellow leaf, was analyzed spatially at different dates in all four trials and compared to commercial cultivars in two of the four trials. These surveys allowed us to identify a correlation between the aphid dynamics in the field and yellow leaf progress. Additionally, a negative correlation was found between rainfall during the first weeks after transferring sugarcane plants to the field and aphid dispersal within the field. This later result revealed an impact of rainfall on aphid invasion and subsequent plant infection by SCYLV. If aphids are the key factor for disease spread, plant response varied also according to cultivar resistance with high variation depending on rain conditions.     

A stable Leifsonia xyli subsp. xyli GFP‐tagged strain reveals a new colonization niche in sugarcane tissues

Ratoon stunting disease (RSD), caused by the bacterium Leifsonia xyli subsp. xyli (Lxx), is one of the most economically important diseases of sugarcane worldwide. Because knowledge on the interaction of Lxx with its host at the microscopic level is limited, the development of tools to monitor Lxx during the colonization process could shed new light on the processes that control disease development. In this investigation, a transformation protocol was optimized and a mutant Lxx strain engineered that stably expressed the gfp gene in sugarcane tissues. In vitro, the growth of the mutant did not differ from that of the wild type. Also, plants inoculated with both strains showed comparable growth and development when analysed 180 days after inoculation (dai). Fluorescence microscopy of roots, stalks, meristems and leaf tissues of Lxx‐GFP‐inoculated plants was performed at 180 dai. In the leaves, Lxx‐tagged cells were observed within the xylem vessels as has been described before but, in addition, they were found in a new niche within the host tissues, in the mesophyll and in the bundle sheath cells surrounding the vascular system. This finding indicates that Lxx is able to move from the xylem to the parenchyma of the leaf cells. This first report of an Lxx mutant expressing a heterologous gene revealed that colonization of sugarcane by this pathogen is not limited to the xylem vessels as commonly reported.     

大麦黄矮病毒-GAV在燕麦植株体内运动规律的初步研究

 利用RT-PCR方法研究了大麦黄矮病毒-GAV在燕麦植株内的移动规律。先将介体麦二叉蚜(Schizaphis graminum)在BYDV-GAV新鲜病叶上饲毒,再将获毒蚜虫放置到二叶期的健康燕麦植株接种48h,随后分期提取接种植株的第1~6片叶和根组织的总RNA,利用特异引物扩增BYDV-GAV的外壳蛋白(CP)基因以检测病毒在燕麦植株内的复制和移动。结果表明,在接种5d后,接种叶片(第2片叶)呈现阳性,接种7d后,植株新生的第4片叶被侵染,接种9d后,部分的第3片叶呈现阳性,至接种16d,几乎所有的叶片均呈现阳性。仅在接种的第5、7和9d收集的根组织呈现阳性,而所有的第1片叶均为阴性,可能是由于这些组织内病毒含量太低所致。本研究初步揭示了BYDV-GAV长距离运动的规律并且发现该病毒在燕麦根部从接种到系统发病都没有进行大量增殖,为今后进一步研究病毒运动机制选取适当的植物材料提供了基本信息。     

Transmission of a sugarcane yellow leaf phytoplasma by the delphacid planthopper Saccharosydne saccharivora, a new vector of sugarcane yellow leaf syndrome

During surveys of sugarcane fields in western and central Cuba from December 2001 to March 2003, the delphacid planthopper Saccharosydne saccharivora was the most prevalent of the Auchenorrhyncha fauna surveyed. Individuals of S. saccharivora collected tested positive for the sugarcane yellow leaf phytoplasma (SCYLP). Saccharosydne saccharivora were reared in cages and used for experimental transmission studies of SCYLP. The S. saccharivora were given acquisition-access feeds of 72 h on SCYLP-infected canes collected from the field followed by an inoculation-access period of 15 days on healthy sugarcane seedlings. Symptoms of yellow leaf syndrome developed on 24 out of 36 plants, 7–12 months postinoculation. None of the 36 healthy seedlings that were inoculated with S. saccharivora fed on phytoplasma-free sugarcane developed symptoms. All phytoplasma-positive sugarcane and S. saccharivora samples showed identical RFLP patterns and had 99·89% similarity in their 16S/23S spacer-region sequences, but only 92·6–93·6% similarity with other phytoplasmas. Sequences were deposited with GenBank [accession numbers: AY725237 ( S. saccharivora ) and AY257548 (sugarcane)]. Phylogenetic analysis suggested that the phytoplasmas from sugarcane and S. saccharivora are putative members of a new 16Sr phytoplasma group. This is the first report of vector transmission of a phytoplasma associated with sugarcane yellow leaf syndrome and the first time that S. saccharivora has been shown to vector a phytoplasma.     

玉米粗缩病毒及抗病策略

玉米粗缩病毒属于植物呼肠孤病毒组的斐济病毒属，是一种有双层衣壳的球状病毒。该病毒基因组包含10条线状的双链RNA片段，近年来已经完成了其中4条RNA片段的测序工作。玉米粗缩病毒对我国玉米生产的危害最为严重。该病毒主要由介体昆虫传播，侵染的植株表现严重矮化和叶片浓绿等病症。玉米对该病毒的抗性表现出多基因控制的数量性状遗传的特征。作者描述了玉米粗缩病病原、病症及侵染循环的特点，对玉米粗缩病毒基因组的研究进展进行了评述，并对抗病策略进行了讨论。     

Elimination of Sugarcane yellow leaf virus from infected sugarcane plants by meristem tip culture visualized by tissue blot immunoassay

Sugarcane yellow leaf virus (SCYLV), a member of the Luteoviridae , is implicated in the sugarcane disease known as yellow leaf syndrome (YLS), which is characterized by yellowing of the leaf midrib followed by leaf necrosis and possible growth suppression. YLS is distributed worldwide and susceptible cultivars are commonly infected with SCYLV. However, not all cultivars infected with SCYLV show symptoms of YLS and some cultivars that show symptoms do so sporadically. Since it is difficult to obtain virus-free plants of susceptible cultivars, it has not been possible to study the factors involved in SCYLV infection nor the effects of infection on plant growth and yield. A tissue blot immunoassay was used to visualize in vivo presence of the virus so that virus-infected and virus-free plants could be distinguished. Meristem tip cultures were used to produce virus-free plantings of six SCYLV-susceptible sugarcane cultivars. Nearly all of the regenerated sugarcane lines remained virus-free over a period of up to 4 years, whether grown in isolated fields or in the glasshouse. Experimental re-infection of the virus-free plants by viruliferous aphids demonstrated that meristem tip culture did not affect susceptibility of sugarcane to SCYLV. Improved diagnosis and production of virus-free plants of SCYLV-susceptible cultivars will facilitate research to quantify the effect of the virus on yield and to analyse the processes involved in disease development.     

Transmission efficiency of Papaya ringspot virus by three aphid species

The transmission efficiency of Papaya ringspot virus (PRSV) by three aphid vectors (i.e., Aphis gossypii, A. craccivora, and Myzus persicae) was studied. Efficiency was measured by single-aphid inoculation, group inoculation (using five aphids), duration of virus retention, and the number of plants following a single acquisition access period (AAP) to which the aphids could successfully transmit the virus. Single-aphid inoculation studies indicated that M. persicae (56%) and A. gossypii (53%) were significantly more efficient in transmitting PRSV than A. craccivora (38%). Further, in the former two species, the time required for initiation of the first probe on the inoculation test plant was significantly shorter compared to A. craccivora. PRSV transmission efficiency was 100% in all three species when a group of five aphids were used per plant. There was a perceptible decline in transmission efficiency as the sequestration period increased, although M. persicae successfully transmitted PRSV after 30 min of sequestration. A simple leaf-disk assay technique was employed for evaluating the transmission efficiency of three species of aphids. The results of leaf-disk assays also indicated that A. gossypii (48%) and M. persicae (56%) were more efficient PRSV vectors than A. craccivora. Using leaf-disk assays, the ability of individual aphids to inoculate PRSV serially to a number of plants was studied. Following a single AAP on an infected leaf, M. persicae was more efficient than the other two species with 52.5% transmission after the first inoculation access period (IAP). However, its inoculation efficiency significantly decreased with the second and subsequent IAPs. A. gossypii was able to transmit PRSV sequentially up to four successive leaf disks, but with significantly declining efficiency. Since A. gossypii is reported to be the numerically dominant vector in south India in addition to being a more efficient vector capable of inoculating PRSV to multiple plants, it should be the target vector for control strategies.     

Detection, identification and significance of phytoplasmas in grasses in northern Australia

Sugarcane yields have been severely reduced by white leaf and grassy shoot phytoplasma diseases in many parts of Asia. Australian sugarcane crops are not known to be affected by these diseases, but plant pathogenic phytoplasmas found in other introduced and native grasses in northern Australia could pose a serious threat to the Australian sugarcane industry. To further evaluate this threat, leaves from plants of 20 grass species, with and without symptoms, were collected during field surveys in northern Australia and tested to determine whether phytoplasmas were present and whether symptoms were reliable indicators of phytoplasma presence. Molecular tools were used to detect and characterize phytoplasmas. Four different phytoplasmas were found in seven grass species known to grow near healthy sugarcane crops. All the phytoplasmas were closely related to sugarcane white leaf phytoplasma (SCWL), one of the phytoplasmas that causes disease in sugarcane in Asia. Four of the host plant species and two of the phytoplasmas were new records. The relationship between symptoms and phytoplasma presence was poor. Because some plants with symptoms tested negative for phytoplasmas, a series of surveys was carried out in which flowers, leaves, roots and stems of two known host plant species, Whiteochloa cymbiformis and Sorghum stipoideum, were tested separately on nine occasions during two wet seasons. This was done to investigate the distribution of phytoplasmas within plants over time. Results showed that spatial and temporal variation of phytoplasmas occurred in these two host plant species. Hence, evaluation of disease distribution within a region requires repeated testing of all plant parts from plants without symptoms, as well as those with symptoms. To date, there is no report of a vector capable of transmitting to Australian sugarcane the phytoplasmas found in grasses in this study. If one is present, or occurs in the future, then native and introduced grasses could constitute a large reservoir of phytoplasma for vectors to draw on. This work provides an early warning for the sugarcane industry that the potential for infection exists.     

Transmission Efficiency of 'Candidatus Liberibacter solanacearum' and Potato Zebra Chip Disease Progress in Relation to Pathogen Titer, Vector Numbers, and Feeding Sites

ABSTRACT With diseases caused by vector-borne plant pathogens, acquisition and inoculation are two primary stages of the transmission, which can determine vector efficiency in spreading the pathogen. The present study was initiated to quantify acquisition and inoculation successes of 'Candidatus Liberibacter solanacearum', the etiological agent of zebra chip disease of potato, by its psyllid vector, Bactericera cockerelli (Hemiptera: Triozidae). Acquisition success was evaluated in relation to feeding site on the host plant as well as the acquisition access period. Inoculation success was evaluated in relation to vector number (1 and 4) on the plants. Acquisition success was influenced by the feeding site on the plant. The highest acquisition success occurred when insects had access to the whole plant. The results of the inoculation study indicated that the rate of successfully inoculated plants increased with the vector number. Plants inoculated with multiple psyllids had higher bacterial titer at the point of inoculation. Although disease incubation period was significantly shorter in plants inoculated with multiple psyllids, this effect was heterogeneous across experimental blocks, and was independent of pathogen quantity detected in the leaflets 3 days postinoculation. Disease progress was not affected by bacterial quantity injected or psyllid numbers.     

甘蔗重要病害分子检测技术

快速有效地对甘蔗重要病害病原进行诊断检测,明确监测病害的病原是科学有效防控甘蔗病害的基础和关键。云南省农业科学院甘蔗研究所通过探索研究、改进创新、优化建立了甘蔗黑穗病、锈病、白条病、宿根矮化病、赤条病、花叶病、斐济病、黄叶病、杆状病毒病和白叶病等10种重要病害13种病原的分子快速检测技术,为甘蔗病害的有效诊断和防控、脱毒健康种苗检测及引种检疫提供了技术支撑。     

Foliar application of manganese increases sugarcane resistance to orange rust

Orange rust (Puccinia kuehnii) is a serious disease compromising the sustainability of sugarcane (Saccharum spp.) production. The objective of this research was to study whether supplemental manganese (Mn) supplied through foliar sprays ameliorates negative effects of orange rust on sugarcane and, if so, to reveal the underlying mechanisms. The experiment was conducted using a sugarcane variety susceptible to the disease; a single spray of Mn at 5 g L⁻¹ (Mn_0.5%) or 10 g L⁻¹ (Mn_1%), plus a control (Mn_0%) was performed before pathogen inoculation. Symptom severity, antioxidant metabolism, lignin deposition and anatomical organization were evaluated. Photosynthesis was also measured in newly expanded leaves and plants were harvested to estimate growth responses. The percentage diseased leaf area was reduced from 15% under Mn_0% to 2.2% and 0.9% under Mn_0.5% and Mn_1%, respectively. This decrease was accompanied by increases in biomass production in the plants. Scanning and light microscopy images revealed that Mn treatment caused direct damage to the fungal spores and improved lignin deposition in the mesophyll. In the presence of the disease, Mn-sprayed leaves exhibited lower levels of oxidative stress, in addition to improved structural organization of xylem and phloem vessels compared to the untreated control. The negative effects of orange rust on gas exchange and photochemistry were also ameliorated by Mn application. The results give insight into the mechanisms underlying augmented sugarcane resistance to orange rust under supplementary foliar Mn spray and contribute to the development of sustainable crop production systems by offering alternatives for reduction of disease damage.