Silicon enhances the accumulation of diterpenoid phytoalexins in rice: a potential mechanism for blast resistance

ABSTRACT Although several reports underscore the importance of silicon (Si) in controlling Magnaporthe grisea on rice, no study has associated this beneficial effect with specific mechanisms of host defense responses against this fungal attack. In this study, however, we provide evidence that higher levels of momilactone phytoalexins were found in leaf extracts from plants inoculated with M. grisea and amended with silicon (Si(+)) than in leaf extracts from inoculated plants not amended with silicon (Si(-) ) or noninoculated Si(+) and Si(-) plants. On this basis, the more efficient stimulation of the terpenoid pathway in Si(+) plants and, consequently, the increase in the levels of momilactones appears to be a factor contributing to enhanced rice resistance to blast. This may explain the lower level of blast severity observed on leaves of Si(+) plants at 96 h after inoculation with M. grisea. The results of this study strongly suggest that Si plays an active role in the resistance of rice to blast rather than the formation of a physical barrier to penetration by M. grisea.     

The role of silicon in preventing appressorial penetration by the rice blast fungus

To test the hypothesis that silicon (Si) confers resistance against appressorial penetration of the rice blast fungus, the proportion of appressorial penetration into the leaf epidermis to total appressoria formed was compared among rice plants amended with various rates of silica gel to those plants nonamended. The amounts of Si in the youngest leaves were consistent with the amounts of silica gel applied to the rice plants. Relative Si levels on the adaxial surface of leaves as detected by energy dispersive X-ray analysis also increased with the amounts of silica gel applied. Based on light microscopic observation of the adaxial surface of rice leaves, the proportion of appressorial penetration was reduced by increasing amounts of silica gel applied and increased with the length of period after spray inoculation. Consequently, these results strongly support the hypothesis and suggest that Si in the leaf epidermis may confer resistance against appressorial penetration. Meanwhile, the number of lesions per leaf also decreased with the amount of Si applied, while only a certain part of penetrated appressoria could become sporulating susceptible lesions. This suggests that Si also confers physiological resistance against blast infection after the penetration.     

Wheat leaf rust uredospore production and carbon and nitrogen export in relation to lesion size and density

ABSTRACT To develop mechanistic yield loss models for biotrophic fungi, we need better account for the export of dry matter, carbon, and nitrogen from the leaf into the spores. Three experiments in controlled environment chambers were performed to study the dynamics of uredospores production of Puccinia triticina on seedling leaves of wheat in relation to time, lesion density, and sporulating surface area. The detrimental effect of lesion density on the sporulation capacity of brown rust lesions was confirmed. When lesion density increased, spores production per lesion strongly decreased. However, our results showed that increasing lesion density also greatly reduces lesion size. A model was developed to summarize these relationships. Our main conclusion is that the density effect on spore production per lesion is accounted for by lesion size. When sporulation was related to the sporulating surface area, it became independent of density. As well, carbon and nitrogen contents of the spores were independent of lesion density. Our data suggest that when nitrogen available in the host is limiting, spore production is reduced but nitrogen content of spores tend to remain stable.     

Ultrastructural and cytochemical aspects of silicon-mediated rice blast resistance

ABSTRACT Although exogenous application of silicon (Si) confers efficient control of rice blast, the probable hypothesis underlying this phenomenon has been confined to that of a mechanical barrier resulting from Si polymerization in planta. However, in this study, we provide the first cytological evidence that Si-mediated resistance to Magnaporthe grisea in rice correlates with specific leaf cell reaction that interfered with the development of the fungus. Accumulation of an amorphous material that stained densely with toluidine blue and reacted positively to osmium tetroxide was a typical feature of cell reaction to infection by M. grisea in samples from Si+ plants. As a result, the extent of fungal colonization was markedly reduced in samples from Si+ plants. In samples from Si- plants, M. grisea grew actively and colonized all leaf tissues. Cytochemi-cal labeling of chitin revealed no difference in the pattern of chitin localization over fungal cell walls of either Si+ or Si- plants at 96 h after inoculation, indicating limited production of chitinases by the rice plant as a mechanism of defense response. On the other hand, the occurrence of empty fungal hyphae, surrounded or trapped in amorphous material, in samples from Si+ plants suggests that phenolic-like compounds or phytoalexins played a primary role in rice defense response against infection by M. grisea. This finding brings new insights into the complex role played by Si in the nature of rice blast resistance.     

施氮量对水稻叶瘟菌侵染循环中主要组分的影响

在人工气候室内进行了施氮量对稻瘟侵染过程主要环节影响的定量试验。结果表明,施氮量对稻瘟菌侵染、病斑扩展及产孢均有不同程度的影响,其中,侵染几率所受影响最大,其次为病斑扩展及产孢,潜育期基本上不受影响。试验还发现,稻瘟侵染、产孢和病斑扩展各组分对氮量均有最适要求,当施氮量超出它们的最适范围后,将不再有利于各组分的发展。以上结果均用数学模型进行了描述,揭示了侵染循环各组分与施氮量间关系及警戒氮量在稻瘟流行学上的意义。     

Wheat Leaf Rust Uredospore Production on Adult Plants: Influence of Leaf Nitrogen Content and Septoria tritici Blotch

ABSTRACT Leaf rust uredospore production and lesion size were measured on flag leaves of adult wheat plants in a glasshouse for different lesion densities. We estimated the spore weight produced per square centimeter of infected leaf, per lesion, and per unit of sporulating area. Three levels of fertilization were applied to the plants to obtain different nitrogen content for the inoculated leaves. In a fourth treatment, we evaluated the effect of Septoria tritici blotch on leaf rust uredospore production. The nitrogen and carbon content of the spores was unaffected or marginally affected by lesion density, host leaf nitrogen content, or the presence of Mycosphaerella graminicola on the same leaf. In leaves with a low-nitrogen content, spore production per lesion was reduced, but lesion size was unaffected. A threshold effect of leaf nitrogen content in spore production was however, evident, since production was similar in the medium- and high-fertilizer treatments. In leaves inoculated with M. graminicola and Puccinia triticina, the rust lesions were smaller and produced fewer spores. The relationships among rust lesion density, lesion size, and uredospore production were fitted to a model. We determined that the density effect on spore production resulted mainly from a reduction in lesion size, the spore production per unit of sporulating surface being largely independent of lesion density. These results are consistent with those obtained previously on wheat seedlings. The main difference was that the sporulation period lasted longer in adult leaves.     

Effect of host genotype on leaf rust (Puccinia triticina) lesion development and urediniospore production in wheat seedlings

Experiments were performed in controlled conditions to compare several aggressiveness components of an isolate of Puccinia triticina on different wheat ( Triticum aestivum ) genotypes. Latency period, urediniospore production per lesion and per unit of sporulating surface, lesion size, and nitrogen and carbon content of the spores were measured on four host genotypes. Three of the host cultivars (Soissons, Altria and Isengrain) were susceptible and the fourth (Trémie) was resistant to the isolate used in the experiments, producing a mesothetic infection type. In all the analyses, lesion density was used as a covariable. The latent period was identical on the susceptible host cultivars, and there were no marked differences in the urediniospore production capacity when related to the sporulating surface area. In cv. Isengrain, differences in lesion size relative to the other susceptible cultivars resulted in lower urediniospore production per lesion. Urediniospore production per lesion and lesion size were density dependent and decreased exponentially with increasing lesion density. Urediniospore production per unit of sporulating surface was either independent of lesion density or marginally dependent (decreased linearly with a limited rate). On the resistant cultivar, Trémie, the latent period was longer and of a much greater variance compared with the susceptible cultivars. Urediniospore production per lesion was considerably reduced, but this reduction was fully accounted for by a reduction in lesion size, the urediniospore production capacity of the diseased tissue remaining equivalent to that observed in the susceptible cultivars. In all cultivars the C and N contents of the urediniospores were considered unchanged, even if minor differences were detected.     

Quantification of Magnaporthe grisea During Infection of Rice Plants Using Real-Time Polymerase Chain Reaction and Northern Blot/Phosphoimaging Analyses

ABSTRACT Rice blast, caused by Magnaporthe grisea, is a serious fungal disease of rice worldwide. Currently, evaluation of the fungal pathogenicity and host resistance is mainly based on a disease rating or measurement of blast lesion number and size. However, these methods only provide visual estimation rather than accurate measurement of fungal growth in rice plants. In this study, DNA-based real-time polymerase chain reaction (PCR) and RNA-based northern blot/phosphoimaging analyses were evaluated to quantify M. grisea. Both methods were sensitive, specific, and reproducible and could accurately measure the relative growth and absolute biomass of M. grisea. The real-time PCR analysis showed that the growth of M. grisea in seedling leaves of susceptible cultivars (M201 and Wells) was approximately 46 to 80 times higher than that of a resistant cultivar (Drew) at 4 and 6 days after inoculation. The data obtained from the real-time PCR assays also were consistent with that from northern blot/ phosphoimaging analysis. However, the real-time PCR approach was much faster and more convenient in most cases. Therefore, it is an excellent tool for in planta quantification of M. grisea and can be used for reliable assessment of fungal pathogenicity and host resistance.     

稻瘟病菌在水稻CO39近等基因系上的生命表和繁殖表

 本文选用以CO39为背景的水稻近等基因系与广东省稻瘟病菌3个优势生理小种(ZC₁₃、ZB₁和ZB₅)构成12个亲和性和非亲和性互作组合,组建各组合稻瘟病菌的生命表和繁殖表,分析在亲和性互作中稻瘟病菌的生存和繁殖状况对寄主发病的影响,并探讨在非亲和性互作中抗性基因在抵御病菌侵染过程中的作用。结果表明,在亲和性和非亲和性互作中,稻瘟病菌生长繁殖各阶段均参与了感抗反应并发挥了一定的作用:在CO39近等基因系上,同一小种的孢子萌发率或菌丝形成率无明显差别,而小种间则表现一定差异;病菌的定殖和繁殖是亲和与非亲和互作的关键阶段,定殖率、病斑形成率低,则寄主表现抗病,反之则表现感病。在繁殖表众多表示种群增长的参数中,世代平均产孢量F'、净增殖率R₀、最大相对增长率r_max和种群趋势指数Ⅰ等4个指标适合于反映稻瘟病菌繁殖状况与感抗病反应的关系。     

持续低温对水稻稻瘟病抗性的影响

研究了在持续低温作用下,具有不同抗性的品种汕优多系１号和汕优６３对稻瘟病菌的抗性变化。结果表明：（１）持续低温处理比未经处理的对照稻瘟病病斑要早出现２－３ｄ;低温处理３－５ｄ出现病斑数目最多,对病害发生最为有利;但处理时间过长,则不利于病害;（２）持续低温处理３－５ｄ对病斑扩展的影响显著。     

水稻品种对稻瘟病菌慢瘟抗性的研究

1985—1987年,以HA-4、Ba-90-2、红突31、HA84494、1952、V/38、837及早选4号为供试品种,研究了水稻品种的慢瘟抗性。结果表明,病斑大小、单位病斑的产孢量、最终百分病叶面积及病害进展曲线下的面积是慢瘟抗性的4个有效组分。4个组分问彼此高度相关(r=0.8082-0.9741)。就4个慢瘟组分对8个品种作聚类分析,可将供试品种划为8类。HA-4、Ba-90-2、红突31、HA84494及1952为慢瘟品种,V/38为中慢品种,837及早选4号为速瘟品种。以ZC_(15)为代表菌研究的慢瘟品种,对ZC_(13)、ZE_3、ZF_1及ZG_1 4个生理小种呈类似的慢瘟反应,在田间自然诱发试验里,也皆表现为抗病。室内外慢瘟性研究结果高度相关(r=0.9972)。     

小麦慢白粉品种的抗性组分研究

 2003~2004年度和2004~2005年度在田间测定了16个小麦品种(系)对小麦白粉菌E20菌株的抗性组分,包括潜育期、侵染几率、产孢量、病斑扩展和传染期。结果表明,小麦慢白粉品种(系)与高感对照品种相比,表现为潜育期长、侵染几率低、产孢量低、病斑扩展慢的特点,且这4个抗性组分与对照的差异都达到了显著和极显著水平,但慢白粉品种的传染期变化无一定规律。参试慢白粉品种的抗性组分参数与病情曲线下面积(AUDPC)的相关分析表明,潜育期与AUDPC呈负相关,但相关性不显著;产孢量和病斑扩展与AUDPC呈正相关,两年数据相关性均达到显著水平;侵染几率与AUDPC呈正相关,但2003-2004年度相关性不显著,而2004~2005年度相关性显著;传染期与AUDPC相关关系不明显。由此可见,在慢白粉品种的病害流行中起主要作用的是产孢量、病斑扩展,而侵染几率、潜育期和传染期作用相对较小。     

稻瘟病为害损失研究

采用人工接种诱发和药剂控制相结合的方法,造成小区间稻叶瘟病和稻穗瘟的发病梯度系统监测病情,收获期结合病情程度进行单株考上区测产,并分别建立了依据稻叶瘟病,稻穗瘟病和二者综合的损失估计模型。就稻叶瘟病地水稻有效穗、穗粒数、千粒重和精米率的影响作了定量研究。另外,地稻穗瘟病在不同品种,不同侵染发病时间以及不同发病部位的为害影响进行了初探。结果表明,稻穗瘟病的为害程度在不同品种上存在一定差异,与侵染发病     

水稻旱晚播“活体病株”接种法的研究及应用

作者针对水稻抗瘟育种中“旱地稻瘟病圃”易受气候生态等条件的影响,导致年际间发病的稳定性和能否发病的安全性,改进以往喷雾接种或利用病区自然诱发法,采用活体病株持续接种,和人为小气候控制等措施,完善成一套发病系数高且稳定,方法简便实用,病圃容量大,鉴定结果可靠性强的综合技术措施,在连续几年试验中鉴定筛选出一批利用价值较高的种质资源和抗病性较好的系统材料。     

Magnaporthe oryzae conidia on basal wheat leaves as a potential source of wheat blast inoculum

Wheat blast caused by Magnaporthe oryzae Triticum causes significant losses on wheat during outbreak years in several South American countries. Despite reports of wheat blast leaf lesions on some wheat cultivars, the importance of inoculum originating from leaves in severely affected commercial fields is disputed. It is generally considered that leaf lesions and/or sporulation on leaves do not usually appear before the occurrence of spike blast in wheat. The purpose of this study was to (i) determine the occurrence of wheat blast on basal leaves, (ii) estimate the number of conidia produced on these leaves, and (iii) determine the impact of current fungicide application practices on inoculum produced from sporulating lesions on basal wheat leaves. Inoculations at the three‐leaf stage showed that certain cultivar and isolate combinations caused more disease on old wheat leaves than young expanding leaves. Under optimum conditions, M. oryzae had the potential to produce tens to hundreds of thousands of conidia on small amounts of wheat basal leaves. A mean of 1 669 000 conidia were produced on 1 g dry basal leaves of a highly susceptible cultivar under optimum conditions for sporulation. Conidia production on leaves coincided with spike emergence under both greenhouse and field conditions. When field studies were conducted under natural epidemic conditions, foliar fungicide applications reduced the amount of M. oryzae conidia on basal leaves by 62–77% compared to non‐sprayed controls. An earlier application of foliar fungicides might reduce inoculum if conidia from basal leaves contribute to wheat spike blast development.     

A comparative assessment of potential components of partial disease resistance to <Emphasis Type="Italic">Fusarium</Emphasis> head blight using a detached leaf assay of wheat,barley and oats

The relative resistance of 15 winter barley, three winter wheat and three winter oat cultivars on the UK recommended list 2003 and two spring wheat cultivars on the Irish 2003 recommended list were evaluated using Microdochium nivale in detached leaf assays to further understand components of partial disease resistance (PDR) and Fusarium head blight (FHB) resistance across cereal species. Barley cultivars showed incubation periods comparable to, and latent periods longer than the most FHB resistant Irish and UK wheat cultivars evaluated. In addition, lesions on barley differed from those on wheat as they were not visibly chlorotic when placed over a light box until sporulation occurred, in contrast to wheat cultivars where chlorosis of the infected area occurred when lesions first developed. The pattern of delayed chlorosis of the infected leaf tissue and longer latent periods indicate that resistances are expressed in barley after the incubation period is observed, and that these temporarily arrest the development of mycelium and sporulation. Incubation periods were longer for oats compared to barley or wheat cultivars. However, oat cultivars differed from both wheat and barley in that mycelial growth was observed before obvious tissue damage was detected under macroscopic examination, indicating tolerance of infection rather than inhibition of pathogen development, and morphology of sporodochia differed, appearing less well developed and being much less abundant. Longer latent periods have previously been related to greater FHB resistance in wheat. The present results suggest the longer latent periods of barley and oat cultivars, than wheat, are likely to play a role in overall FHB resistance if under the same genetic control as PDR components expressed in the head. However the limited range of incubation and latent periods observed within barley and oat cultivars evaluated was in contrast with wheat where incubation and latent periods were shorter and more variable among genotypes. The significance of the various combinations of PDR components detected in the detached leaf assay as components of FHB resistance in each crop requires further investigation, particularly with regard to the apparent tolerance of infection in oats and necrosis in barley, after the incubation period is observed, associated with retardation of mycelial growth and sporulation.     

Expression Analysis of Rice Defense-Related Genes in Turfgrass in Response to Magnaporthe grisea

ABSTRACT Magnaporthe grisea (anamorph = Pyricularia grisea) causes blast on rice (Oryza sativa) and gray leaf spot on turfgrass. Gray leaf spot is a serious disease on St. Augustinegrass (Stenotaphrum secundatum), perennial ryegrass (Lolium perenne), and tall fescue (Festuca arundinacea). Virulence assays performed in this study revealed that M. grisea collected from rice could also cause disease on St. Augustinegrass and tall fescue. One rice isolate, Che86061, caused similar disease reactions on susceptible cultivars of rice and St. Augustinegrass and an incompatible interaction on resistant cultivars of both species. To explore whether similar defense-related genes are expressed in rice and St. Augustinegrass, a rice cDNA library was screened using pooled cDNAs derived from M. grisea-infected St. Augustinegrass. Thirty rice EST (expressed sequence tag) clones showing differential expression in St. Augustinegrass following M. grisea inoculation were identified and classified into six putative functional groups. Northern blot analyses of seven EST clones that collectively represented each putative functional group confirmed that the expression of five out of seven EST clones was similar in both rice and St. Augustinegrass. This study represents one of the first attempts to use a broad-scale genomic approach and resources of a model monocot system to study defense gene expression in St. Augustinegrass following M. grisea infection.     

Seasonal variation in the components of partial resistance of seedlings of winter wheat cultivars Maris Huntsman and Maris Ranger to Septoria nodorum

Components of partial resistance of two winter wheat cultivars, Maris Huntsman and Maris Ranger, to Septoria nodorum were measured on potted seedlings exposed in the open during the spring and early summer of 1978. The number of lesions 10 days after inoculation was always greater for Maris Ranger than for Maris Huntsman but the relative differences changed during the season: about 10:1 in March, April and May, but only 3:1 in June. The time from infection to first sporulation (latent period) on Maris Ranger was always shorter, especially in March. The percentage leaf area covered by lesions at the end of the latent period was usually greater on Maris Ranger, but variation in the percentage leaf area necrotic at this time did not follow a distinct seasonal trend, nor did cultivars differ in this respect. Twice as many spores per unit area diseased were produced on Maris Ranger as on Maris Huntsman. Seasonal variation in these components could partially be interpreted in terms of the changing weather conditions to which the plants were exposed.     

The effect of temperature and light intensity on growth and sporulation of Puccinia striiformis on wheat

Urediniospore production by Puccinia striiformis on wheat per unit leaf area infected was much lower at low light intensities than at high light intensities. The number of pustules per unit area of infected leaf and the daily sporulation rate per pustule increased linearly with increasing light over the range 10–50 W/m². Increasing temperature between 7 and 20°C shortened latent period and reduced the longevity of sporulating leaves. Colonization rate and the frequency of pustules per unit area of infected leaf increased between 7 and 15°C but declined markedly at 20°C. Spore production reached its peak earlier and declined more rapidly with increasing temperature between 7 and 15°C. this decline being less marked in the highly susceptible cultivar Maris Beacon than in the more resistant Maris Nimrod and Maris Huntsman.     

稻瘟菌水平抗性稳定性的研究——品种与小种适合度的分析

在温室条件下,对9个无垂直抗性基因粳稻品种×8个日本标准稻瘟菌株组合测定了侵染几率(IP)、病斑型比例(PB)、病斑扩展速率(LR)和产孢能力(SPC)组分,并据此确定了品种与小种的适合度。方差分析结果表明,除SPC的互作项外,其它组分及类型,方差均达到显著或极显著水平。但品种和小种主效应所占方差更大,从而证实了品种的抗性类型为水平抗性。某些品种在IP、PB及LR组分上反映了小种的专化作用,但因为不同品种在各组分上的抗性反应不同,因此,品种水平抗性的稳定性不能由组分的小种专化作用确定,而须由各组分组成的适合度的小种专化性确定。对品种X小种的适合度进行了互作的多重比较,未发现专化性互作,说明供试品种的水平抗性是稳定的。