Cauliflower Mosaic Virus Isolate NY8153 Breaks Resistance in Arabidopsis Ecotype En-2

ABSTRACT Arabidopsis thaliana ecotype En-2 was previously shown to be resistant to cauliflower mosaic caulimovirus (CaMV) isolate CM4-184. In this study, En-2 plants were screened with eight other isolates of CaMV to identify viruses capable of overcoming resistance and to determine if the mechanism of resistance was the same for each virus. En-2 resistance to most CaMV isolates was mediated by the same mechanism, i.e., preventing virus long-distance movement. One CaMV isolate, NY8153, was found that produced a severe systemic infection on En-2 plants. In addition, the CM1841 isolate was able to spread systemically through En-2 plants, to a limited extent, without producing visible symptoms. These data indicate that the resistance shown by En-2 plants is not an all-or-none phenomenon. En-2 plants were susceptible to turnip mosaic potyvirus, suggesting that resistance is specific to CaMV.     

Introgression of a Tombusvirus Resistance Locus from Nicotiana edwardsonii var. Columbia to N. clevelandii

ABSTRACT A new variety of Nicotiana, N. edwardsonii var. Columbia, was evaluated for its capacity to serve as a new source for virus resistance genes. Columbia was developed from a hybridization between N. glutinosa and N. clevelandii, the same parents used for the formation of the original N. edwardsonii. However, in contrast to the original N. edwardsonii, crosses between Columbia and either of its parents are fertile. Thus, the inheritance of virus resistance genes present in N. glutinosa could be characterized by using Columbia as a bridge plant in crosses with the susceptible parent, N. clevelandii. To determine how virus resistance genes would segregate in interspecific crosses between Columbia and N. clevelandii, we followed the fate of the N gene, a single dominant gene that specifies resistance to Tobacco mosaic virus (TMV). Our genetic evidence indicated that the entire chromosome containing the N gene was introgressed into N. clevelandii to create an addition line, designated N. clevelandii line 19. Although line 19 was homozygous for resistance to TMV, it remained susceptible to Tomato bushy stunt virus (TBSV) and Cauliflower mosaic virus (CaMV) strain W260, indicating that resistance to these viruses must reside on other N. glutinosa chromosomes. We also developed a second addition line, N. clevelandii line 36, which was homozygous for resistance to TBSV. Line 36 was susceptible to TMV and CaMV strain W260, but was resistant to other tombusviruses, including Cucumber necrosis virus, Cymbidium ringspot virus, Lettuce necrotic stunt virus, and Carnation Italian ringspot virus.     

Phenotype and spectrum of action of the Pvr4 resistance in pepper against potyviruses, and selection for virulent variants

The dominant Pvr4 gene identified in Capsicum annuum cv. Criollo de Morelos 334 (CM334) is frequently used in pepper cultivars because it possesses one of the largest spectra of action among plant virus resistance genes. This gene was previously shown to confer efficient resistance to all known Potato virus Y isolates, to Pepper mottle virus , to Pepper yellow mosaic virus and to Ecuadorian rocoto virus. This study showed that the W4 line, derived from CM334 and carrying Pvr4 , was also resistant to Peru tomato mosaic virus and Pepper severe mosaic virus , but not to Pepper veinal mottle virus , Chilli veinal mottle virus or Tobacco etch virus . It was noticed that the phenotype of the resistance was atypical since, in the W4 line, hypersensitive reaction or extreme resistance could be observed, depending on virus isolates and inoculated organs. Despite the large deployment of Pvr4 in hybrid cultivars, the numerous tests performed in controlled conditions and the use of W4 serial back-inoculations with potyvirus isolates controlled by this line, no virulent variant isolates were obtained. However, it was shown that the use of graft inoculation experiments allow PVY virulent variants to be selected.     

A Single Locus Leads to Resistance of Arabidopsis thaliana to Bacterial Wilt Caused by Ralstonia solanacearum Through a Hypersensitive-like Response

ABSTRACT Strains of Ralstonia solanacearum have been shown to cause bacterial wilt in some, but not all, ecotypes of Arabidopsis thaliana. We demonstrate here that after inoculation of the leaves of resistant ecotype S96 with R. solanacearum strain Ps95 necrosis around the inoculation site rapidly appeared and no further symptoms developed in the plant. Leaves of susceptible ecotype N913 completely wilted 7 days after inoculation with Ps95, and symptoms spread systemically throughout the whole plant within 2 weeks after inoculation. These results suggest that the resistance of Arabidopsis S96 to R. solanacearum is due to a response similar to the hypersensitive response (HR) observed in other plant diseases. Northern blot analysis of the expression of defense-related genes, known to be differentially induced during the HR in Arabidopsis, indicated that pathogenesis-related protein PR-1, glutathione S-transferase (GST1), and Cu, Zn superoxide dismutase (SOD) mRNAs increased significantly in S96 leaves between 3 to 12 h after infiltration with Ps95. The induction of these genes in susceptible ecotype N913 by Ps95 was clearly delayed. Genetic analysis of crosses between resistant ecotype S96 and susceptible ecotype N913 indicated that resistance to Ps95 is due to a single dominant locus.     

Inheritance of Resistance to Watermelon mosaic virus in Cucumis melo that Impairs Virus Accumulation, Symptom Expression, and Aphid Transmission

ABSTRACT The Cucumis melo accession TGR-1551 was found to be resistant to Watermelon mosaic virus (WMV, genus Potyvirus, family Potyviridae). The resistance resulted in a drastic and significant reduction of virus titer and infected plants were asymptomatic or exhibited mild disease symptoms. The same gene or closely linked genes restricted virus accumulation and ameliorated symptom expression. No effect was observed on virus accumulation in inoculated leaves, which suggested that the initial phases of infection were not affected. The resistance was effective against a range of WMV isolates from Spanish melon production areas. Using aphid inoculations, resistant plants showed a lower propensity for infection by WMV and for serving as virus sources for secondary spread once infected. Resistance was determined to be under recessive genetic control.     

Identification and characterization of resistance to rhizomania in an ecotype of Beta vulgaris subsp. maritima

An ecotype (S023) of Beta vulgaris subsp. maritima was identified which was highly resistant to beet necrotic yellow vein virus (BNYVV) but susceptible to its fungal vector, Polymyxa betae . A comparative study of viral development in the roots showed that the resistance was effective from the early stages of infection. Mechanical inoculation experiments suggested that the resistance was not due to a lack of transmission of BNYVV by its fungal vector. Immunogold-silver labelling of the viral coat protein in root sections confirmed the existence of BNYVV transmission in the resistant ecotype. Virus replicated in isolated root protoplasts of both susceptible and resistant genotypes. Viral particles localized by immunogold-silver labelling diffused over short distances from the primary infected root cells in both resistant and susceptible plants. Evidence for inhibition of long-distance movement in the resistant ecotype requires further investigation.     

小麦丛矮病研究之二关于小麦丛矮病的病原病毒问题

在田间的小麦丛矮病非典型病株申常常分离到北方禾谷花叶病毒(NCMV)及玉米粗缩病毒(MRDV)两种病原物。单独用前者接种,仅表现小麦丛矮症,单独用后者接种,仅表现小麦蓝矮症,即植株矮化、暗绿、分蘖减少,叶坚挺。两者混合侵染时,往往表现两者的混合症状,但以何种病毒占优势而倾向于优势病毒的症状。典型丛矮症植株中电镜检出弹状病毒粒体,40—70×270—400nm尺度与日本报导的稍异。典型丛矮病株中不存在玉米粗缩病毒的粒体。小麦丛矮病毒很可能是北方禾谷花叶病毒的一种地方性株系。     

A recessive resistance gene of alfalfa to alfalfa mosaic virus and impact of high ambient temperature on virus resistance

Twenty plants of alfalfa cv. Beaver were screened for resistance to alfalfa mosaic virus (AIMV) severe strain A-515. ELISA screening on both inoculated and apical leaves at fixed temperature (20°C day, 16°C night) suggested the following three types of clonal response to AIMV infection: extremely resistant, AIMV not normally detected from either inoculated or apical leaves; resistant, AIMV detected from inoculated leaves only and did not spread systemically; susceptible, AIMV detected from both inoculated and apical leaves. When plants in the second category were maintained at high temperature (30°C), AIMV was detected from inoculated and apical leaves 6 and 12 days after inoculation, but was not detectable in the apical leaves thereafter. Plants in the first category remained extremely resistant at all temperatures tested. The results of comparative tests using progeny plants of extremely resistant, resistant and susceptible plants, and of their hybrids, suggested that the resistance to AIMV A-515 was controlled by a temperature-dependent recessive gene.     

Response of Hard Red Winter Wheat to Soilborne wheat mosaic virus Using Novel Inoculation Methods

ABSTRACT Soilborne wheat mosaic virus (SBWMV) is an agronomically important pathogen of wheat that is transmitted by the soilborne plasmodiophorid vector Polymyxa graminis. In the laboratory, attempts to generate SBWMV-infected plants are often hampered by poor infectivity of the virus. To analyze the mechanism for virus resistance in wheat cultivars, we developed novel inoculation techniques. A new technique for foliar inoculation of SBWMV was developed that eliminated wound-induced necrosis normally associated with rub inoculating virus to wheat leaves. This new technique is important because we can now uniformly inoculate plants in the laboratory for studies of host resistance mechanisms in the inoculated leaf. Additionally, wheat plants were grown hydroponically in seed germination pouches and their roots were inoculated with SBWMV either by placing P. graminis-infested root material in the pouch or by mechanically inoculating the roots with purified virus. The susceptibility of one SBWMV susceptible and three field resistant wheat cultivars were analyzed following inoculation of plants using these novel inoculation techniques or the conventional inoculation technique of growing plants in P. graminis-infested soil. The results presented in this study suggest that virus resistance in wheat likely functions in the roots to block virus infection.     

A single amino Acid change in viral genome-associated protein of potato virus y correlates with resistance breaking in 'virgin a mutant' tobacco

ABSTRACT Tobacco cultivar Virgin A Mutant (VAM) is reported to have the recessive potyvirus resistance gene va. Varied levels of resistance were observed in VAM plants inoculated with Japanese potato virus Y (PVY) isolates. VAM was highly resistant to most of the PVY isolates tested and tolerant to three necrotic strain isolates of PVY-T. Based on data obtained from tissue printing and press blotting, the resistance appeared to be mainly at the level of cell-to-cell movement. PVY replicated in VAM proto-plasts, but the replication was 30% lower than in susceptible tobacco, suggesting that impairment of replication also contributes to resistance. To identify the viral gene product or products involved in VAM resistance, we isolated spontaneous resistance-breaking mutants by passing vein-banding (O strain) isolates several times through VAM plants. By comparing the amino acid sequences of the mutants with their original isolates, we identified a single amino acid substitution in the viral genome-associated protein (VPg) domain that is correlated with VAM resistance breaking. Together, these results suggest that, in addition to its role in replication, VPg plays an important role in the cell-to-cell movement of PVY.     

Near immunity to rice tungro spherical virus achieved in rice by a replicase-mediated resistance strategy

ABSTRACT Rice tungro disease is caused by rice tungro bacilliform virus (RTBV), which is responsible for the symptoms, and rice tungro spherical virus (RTSV), which assists transmission of both viruses by leafhoppers. Transgenic japonica rice plants (Oryza sativa) were produced containing the RTSV replicase (Rep) gene in the sense or antisense orientation. Over 70% of the plants contained one to five copies of the Rep gene, with integration occurring at a single locus in most cases. Plants producing antisense sequences exhibited significant but moderate resistance to RTSV (60%); accumulation of antisense RNA was substantial, indicating that the protection was not of the homology-dependent type. Plants expressing the full-length Rep gene, as well as a truncated Rep gene, in the (+)-sense orientation were 100% resistant to RTSV even when challenged with a high level of inoculum. Accumulation of viral RNA was low, leading us to conclude that RTSV Rep-mediated resistance is not protein-mediated but is of the cosuppression type. Resistance was effective against geographically distinct RTSV isolates. In addition, RTSV-resistant transgenic rice plants were unable to assist transmission of RTBV. Such transgenic plants could be used in an epidemiological approach to combat the spread of the tungro disease.     

Emergence of Resistance-breaking Isolates of Rice yellow mottle virus during Serial Inoculations

Two sources of resistance to Rice yellow mottle virus were challenged in host passage experiments. Pronounced changes in pathogenicity occurred over serial passages of virus isolates inoculated to partially or highly resistant cultivars. The changes encompassed the known existing pathogenic variability of field isolates. Ultimately, the high resistance of the Oryza indica cv. Gigante was overcome and the partial resistance of the O. sativa japonica cv. Azucena broke down. The effect was resistance-specific as different isolates overcame partial and high resistance, and may also be allele-specific as different isolates overcame the resistance of cultivars carrying the same resistance gene. The ability of isolates to break resistance was not linked to a high initial pathogenicity of the isolates and did not result in higher virus content in the infected plants. Implications for resistance breeding and deployment are discussed.     

草莓伪轻型黄边病毒的鉴定

 从6省13市约18品种草莓样本、分离纯化并研究鉴定,初步证实辽宁、哈尔滨及南京一些栽培田,有草莓伪轻型黄边病毒(SPMYEV)的发生分布。在春香、宝交早生、丹东鸡心,诺宾卡及80-3.1等品种上发病。病样先以小叶嫁接技术,接于草莓UC-4、EMC及"Alpine",约3~5周显症、下部叶片呈现黄色斑块,接UC-5、EMB不太敏感仅于老叶上现轻斑驳或褪绿斑,一种草莓中瘤钉毛蚜(Chaetosiphon sp.)及棉蚜(Aphis gossypii)作半持久性传播本病毒,桃蚜(Myzus persicae)不传播。UC-4对本病毒的繁殖、保存和嫁接鉴别最为适合,提纯病毒电镜观察其粒子线状,大小约630~680×12~13nm,县典型核蛋白吸收光谱,其最大吸收值位于2.63nm处;最小吸收值位于243nm处。A260/280=1.24。经SDS-聚丙烯酰胺凝胶电泳,测其外壳蛋白分子量约为37,000d。病叶制做超薄切片也观察到线形晶状病毒粒子,分散或聚集于寄主的薄壁组织细胞质中。本病毒系国内首报。     

Transgenic resistance to cucumber mosaic virus in tomato: blocking of long-distance movement of the virus in lines harboring a defective viral replicase gene

ABSTRACT Tomato breeding lines were transformed with a defective replicase gene from RNA 2 of cucumber mosaic virus (CMV). A total of 63 transformants from five tomato genotypes were evaluated for resistance to CMV strains. The responses of R1 transgenic offspring fit into three categories: fully susceptible lines (44%), fully resistant lines (8%), and an intermediate-type mixture of susceptible and resistant seedlings in variable proportions (48%). Further characterization of the response of two highly resistant lines was performed by mechanical inoculation, aphid transmission, or grafting experiments. No virus was detected in noninoculated leaves from these lines, although a low level of virus accumulated initially in the inoculated leaf. The homozygous R2 plants and further generations that were evaluated (up to R5) showed resistance to the Fny-CMV strain, two Israeli isolates tentatively classified as subgroup IA, and K-CMV (a representative of subgroup IB). These lines were partially resistant to LS-CMV (a representative of subgroup II) when a high-virus-titer inoculum was used. Expression of the viral transgene was verified in these lines; however, the expected translation product was not detectable. In grafting experiments, we demonstrated that CMV virions were blocked in their ability to move from infected rootstocks of nontransformed tomato or tobacco into the transgenic scions. Interestingly, virions could not move through a transgenic intersection into the upper scion. These results provide an additional indication that replicase-mediated resistance affects long-distance movement.     

利用RNA介导的抗病性获得抗2种病毒的转基因烟草

 RNA介导的病毒抗性(RMVR)是近年发展起来的一种新的植物抗病毒基因工程策略,具有抗病性强、抗性持久、生物安全性高等特点。利用该策略培育多抗病毒植株具有广阔的应用前景和重要的实践意义。本研究将非翻译的马铃薯X病毒的衣壳蛋白(PVX-CP)基因和非翻译的马铃薯Y病毒的衣壳蛋白(PVY-CP)基因组成嵌合基因,构建植物表达载体pROKXY,利用农杆菌介导法转化烟草NC89,获得6株对PVX和PVY的混合侵染表现为免疫的转基因植株。分子分析表明,这种抗性为RNA介导的病毒抗性。这一研究结果为利用RMVR进行植物多抗病毒育种提供了重要数据和资料。     

EMS and UV irradiation induce unstable resistance against CAA fungicides in <Emphasis Type="Italic">Bremia lactucae</Emphasis>

Wild type (WT) field isolates of Bremia lactucae failed to germinate in vitro or infect lettuce leaves in the presence of CAA (carboxylic acid amide) fungicides. Minimal inhibitory concentrations (MIC) for mandipropamid, dimethomorph, benthiavalicarb and iprovalicarb were 0.005, 0.5, 0.5 and 5 μg ml⁻¹, respectively. Mutagenesis experiments showed that spores exposed to EMS (ethyl methane sulphonate) or UV irradiation (254 nm) could infect lettuce leaves in the presence of up to 100 μg ml⁻¹ CAA. The proportion of infected leaves relative to the number of spores inoculated (infection frequency) was inversely related to the concentration of CAA used, ranging between 0 and 160 per 1 × 10⁶ spores. Resistant mutants (RM) lost their resistance within 1–14 reproduction cycles on CAA-treated plants. Crosses were made between RMxWT isolates and RMxRM isolates with an attempt to obtain stable homozygous resistant off-springs. Such crosses yielded few resistant but unstable progeny isolates. Mutagenic treatments given to hybrid isolates also failed to produce stable resistance. Previous gene sequencing data showed that stable resistance to CAAs is based on a single SNP in the cellulose synthase 3 (CesA3) gene of Plasmopara viticola. Therefore, we sequenced a 582 bp DNA fragment of Ces3A of WT, RM and hybrid isolates of B.lactucae. No mutation in this gene fragment was found. We conclude that mutagenic agents like EMS or UV may induce resistance to CAA in Bremia lactucae but this resistance is not stable and not linked to mutations in CesA3 gene.     

A Laboratory Assay for Phytophthora infestans Resistance in Various Solanum Species Reflects the Field Situation

Physiological and molecular research on resistance responses of Solanum tuberosum cultivars and partially resistant Solanum species to Phytophthora infestans requires a reliable resistance test that can be used in the laboratory. Laboratory tests performed on detached leaves and intact plants were compared with field tests for similarity of late blight reactions. Detached leaves from field-grown plants were as resistant as detached leaves from climate chamber-grown plants when challenged with P. infestans. However, detached leaves incubated in covered trays at high relative humidity were more susceptible than detached leaves kept in open trays or leaves on intact plants. The incubation conditions of detached leaves in covered trays rather than detachment itself appeared to affect the resistance expression. Detached leaves of some wild Solanum genotypes became partially infected, whereas intact plants were completely resistant when inoculated. Inoculation of leaves on intact plants, however, resulted in lower infection efficiencies. These limitations should be taken into account when choosing the appropriate inoculation method for specific purposes. For resistance screening, laboratory tests proved to be a good alternative for field tests. The ranking of resistance levels for twenty plant genotypes was similar under laboratory and field conditions.