Effect of Host Plant Resistance on Haustorium Formation in Cereal Rust Fungi

Haustoria of Puccinia triticina (wheat leaf rust fungus) and P. hordei (barley leaf rust fungus) were isolated from susceptible and partially resistant wheat lines, and susceptible, hypersensitive and partially resistant barley lines. Haustoria were counted and measured. The size of haustoria was similar in the partially resistant and susceptible genotypes but haustoria were smaller in the hypersensitive barley line L94+Pa7. The number of haustoria was reduced in both partially and hypersensitive lines when compared with susceptible ones. Therefore it seems that the reduction in the number of haustoria is a consequence of the resistance that can be attributable either to early abortion of infection units or reduced colony growth. The reduction of the number of haustoria was more pronounced in the adult plant stage.     

Investigations on Adult Plant Resistance of Barley Against Erysiphe graminis f.sp. hordei

Spring barley cultivars and lines were tested for 3 years in field studies for adult plant resistance against Erysiphe graminis f.sp. hordei. The cultivars Osiris and Asse were selected for further detailed cytological studies and compared with the susceptible cultivar Peruvian. Under controlled greenhouse conditions, the percentage of conidia that had formed a functional haustorium and secondary hyphae (infection efficiency) was reduced in fifth leaves of the adult plant resistant cultivars. On fifth and flag leaves of adult plant resistant cultivars, papillae were formed more frequently under primary germ tubes and appressoria, and fungal penetration was prevented more often than on the susceptible cultivar Peruvian. In ultrastructural studies various types of papillae were observed, but could not be strictly correlated with penetration success or failure of the fungus.     

Seedling resistance of wheat to the powdery mildew fungus,Erysiphe graminis f. sp. tritici. I. Resistance of first leaves

During primary infection by conidia ofErysiphe graminis f. sp.tritici, three mechanisms of resistance operate in first leaves of 8-day-old seedlings of both resistant and susceptible wheats. The first mechanism, operating at the penetration site, is responsible for the failure of penetrations attempted by primary germ tubes (PGT). The second mechanism is concerned with the abortion of haustoria in normal-appearing host cells. The third mechanism relates to the abortion of haustoria and the hypersensitivity of the penetrated host cells.With the inoculum-level of 19–24 conidia/mm², the three mechanisms together prevented 89.3 % of the attempted penetrations by PGT from producing normal haustoria in resistant wheat Purdue 5752C1-7-5-1 and 37.4 % in the susceptible wheat Vermillion. The first mechanism accounted for the prevention of 73.3 % of the attempted PGT penetrations on Purdue 5752C1-7-5-1 and 36 % on Vermillion. The second mechanism was responsible for stopping 19 % of all the successful penetrations in Purdue 5752C1-7-5-1 and 0.8 % in Vermillion. The third mechanism accounted for the failure of 41 % of all the successful penetrations in Purdue 5752C1-7-5-1 and 1.4% in Vermillion. Thirty-six hours after inoculation, 10.7% of all the attempted PGT penetrations appeared to be developing normally in first leaves of 8-day-old seedlings of resistant wheat Purdue 5752C1-7-5-1 as compared to 62.6 % in the susceptible wheat Vermillion.This appears to be the first report showing the relative effectiveness of various mechanisms of resistance concerning any powdery mildew fungus.     

Untersuchungen zur Anfälligkeit und Resistenz von Kopfsalat (Lactuca sativa) gegen falschen Mehltau (Bremia lactucae): II. Licht- und elektronenmikroskopische Untersuchungen zur Morphologie des Wirt-Parasit-Verhältnisses

Investigations on the susceptibility and resistance of head lettuce (Lactuca sativa) to downy mildew (Bremia lactucae) II. Light and electron microscopic examinations of the host-parasite interface Infected leaves of lettuce varieties susceptible and incompletely resistant to Bremia lactucae were observed by light and electron microscopy. Primary infection structures in the epidermal cells as well as intercellular hyphae with the adjacent haustoria could be seen by differential interference contrast microscopy. The haustoria in host cells of susceptible varieties collapsed before degeneration of the invaded host cell. On the contrary, host cells of incompletely resistant varieties died before the haustoria in these cells showed any sign of degeneration. Electron microscopic investigations confirmed the observations with light microscopy. In incompletely resistant varieties, an electron transparent sheath enveloped the haustorium. In the sheath fragments of membranes are localized. These membrane particles as seen by using the goniometer in electron microscopic work were flat faced. The sheath material consists of transformed host cell wall material and involves fragments of the host plasmalemma as well as fragments of the unit membrane separating the sheath from extrahaustorial matrix. The sheath has an important role as a special filter to prevent the passage of nutrients from the host cell into the haustorium. Thus the incomplete resistance is based not only on an impeded penetration of the parasite into the epidermal cells and their hypersensitive reactions in case of a successful penetration but also on hypersensitivity of mesophyll cells which does not necessarily lead to death of the parasite but does impede the absorption of nutrients.     

Isolation of haustoria ofPuccinia graminis f.sp.tritici from wheat leaves

Haustoria of the wheat stem rust fungus,Puccinia graminis f.sp.tritici, race 32, were isolated from two different infected wheat cultivars. Yield of haustoria from the highly susceptible wheat cv. “Little Club” was 10 times higher than that of the resistant wheat line “Prelude × Eagle” carrying the Sr26 gene for resistance toP. graminis f.sp.tritici. Tests for the integrity of haustoria using the stain methylene blue showed that up to 91% of the haustoria were undamaged.     

The early stages of mildew colony development on susceptible oats

The chronology of primary infection by Erysiphe graminis f. sp. avenae on detached leaves of a susceptible host, and growth patterns of the primary haustorium and secondary hyphae, proved similar to those of wheat and barley mildew found by other workers. The timing of formation and subsequent growth of the primary haustorium was not affected by the light regime, but the formation of subsequent haustoria was highly synchronous under an alternating dark/light cycle, and far less so under continuous illumination. Five days after inoculation almost twice as many secondary and tertiary haustoria were formed per colony under the dark/light treatment than under continuous light. Because of the synchronous formation of haustoria subsequent to the primary, haustoria selected at random from leaves of susceptible host cultivars showed a bimodal distribution in length, the less well developed tertiary haustoria being distinguished from earlier formed primary and secondary haustoria. There was also a significant positive correlation between length and the number of digitate processes/haustorium. The energy required to produce one secondary haustorium was calculated to be equivalent to that required to produce approximately 4–7 hyphal cells.     

Combined Gene Effects on Resistance against Erysiphe graminis DC. f. sp. hordei Marchal Analysed by Means of Double Mutants1)

The effect of genes for resistance against powdery mildew of barley was determined microscopically in stained leaf segments 44 h after inoculation and compared with the visual was established between the number of haustoria and the visual disease symptoms. Deviations were expressed after C17 Am inoculation of the mutant B682, which exhibited low infection grade although high numbers of haustoria were formed; thus the induced resistance gene of B682 is expressed only in later stages of the infection process. On the other hand, after inoculation of SR1 with the ml-o virulent isolate HL3 high infection grade was measured following low haustoria numbers; obivously, this isolate is able to grow more intensively after first haustoria have been formed.Consequently, the close relation between haustoria numbers and infection grade is reestablished in the double mutants under HL3 infection. The implications of this finding for resistance breeding are discussed.     

马铃薯对晚疫病水平抗性的组织细胞学观察

以马铃薯晚疫病水平抗性品种LBr-12和感病品种费乌瑞它为材料,采用普通光学和电子显微镜技术,系统研究了马铃薯与晚疫病菌(致病疫霉)互作的组织细胞学反应特征。观察结果显示:(1)接种后,水平抗性材料LBr-12出现过敏反应,病菌被限制在侵染点的几个细胞中,菌丝产生较少的分支和吸器。(2)感病品种费乌瑞它被侵染细胞呈蔓延趋势,菌丝产生较多的分支和吸器。(3)电镜观察发现,抗病品种上病菌的胞间菌丝、吸器母细胞、吸器在细胞和亚细胞水平均发生了一系列异常变化,包括原生质的电子致密度加深、液泡增多变大、菌丝细胞壁不规则增厚、细胞器排列紊乱及解体、吸器母细胞及吸器形态异常、病菌最终畸形坏死,同时发现抗病品种受病菌侵染时可迅速产生结构防卫反应,形成的细胞壁沉积物使胞壁极度增厚或细胞膜上产生乳突状结构。     

Ultrastructural and Histochemical Studies on Mildew of Barley (Erysiphe graminis DC. f. sp. hordei Marchal)

Plants of the mildew susceptible barley cultivar Peruvian and the adult plant resistant cultivar Osiris were inoculated with Erysiphe graminis f. sp. hordei at the first and fifth leaf stages. Samples taken at 32 hr after inoculation were examined by electron microscopy to compare papillae associated either with penetration failure or with successful penetration of the fungus into the epidermal cell and haustorium formation. Four types of papillae with ultrastructural differences could, be classified. Although their definite association with fungal ingress or failure is not possible, our data suggest that papillae with larger, more compacted and amorphous or globular structures may be more effective as penetration barriers than others, with more or less uniform distribution of irregular, smaller electrondense structures.     

Production of branches from unsuccessful primary germ tubes of conidia of Erysiphe graminis f. sp. tritici on wheat leaves

Confusion exists regarding nomenclature and possible significance of outgrowths produced by conidial primary germ tubes (PGT) of Erysiphe graminis f. sp. tritici Em. Marchai on wheat leaves. Production of these outgrowths has been taken as a criterion of successful host-parasite relationship. However, such outgrowths have been observed to have developed from 12.3 % of the unsuccessful PGT on a resistant wheat and 9.2 % of the unsuccessful PGT on a susceptible wheat.It is proposed that these outgrowths may be called PGT branches which may develop whether the penetration by PGT is successful or not. In case the PGT penetration is successful, then such a PGT branch may be called a secondary hypha. These branches from unsuccessful PGT may also produce host responses in the form of halos in the host cell-walls or may also penetrate the host cells successfully and produce apparently healthy (normal) haustoria. Both PGT branches and the secondary hyphae may be called functional when they produce either host responses in the form of halos or penetrate successfully and produce haustoria, or produce both halos and haustoria.     

Resistance to powdery mildew in rose

Resistance and susceptibility of rose species and varieties to the powdery mildew fungus Sphaerotheca pannosa were tested on detached leaflets in Petri plates. Strains of the pathogen from greenhouse plants were maintained under aseptic conditions on rose shoots growing in agar medium in glass vials. Biological specialization in S. pannosa was confirmed; the strain on Rosa virginiana did not infect common rose varieties. Variation in susceptibility of commonly cultivated rose varieties to another strain of mildew was demonstrated, and resistance increased with age of leaf. Germination and prepenetration development of the fungus were not related to disease resistance. Fewer fungus haustoria developed normally in epidermal cells of resistant than of susceptible rose varieties. The relative importance of morphological barriers and internal resistance factors is discussed. Attempts failed to infect freshly isolated or long established callus tissue cultures of several rose varieties.     

Inheritance of powdery mildew (Erysiphe polygoni DC) resistance in mungbean (Vigna radiata L. Wilczek)

Detached mungbean (Vigna radiata L.Wilczek) leaves were inoculated with a conidial suspension of a local isolate (TI-1) of the powdery mildew pathogen (Erysiphe polygoni DC) under controlled environment conditions. Based on the latent period and severity of the infection, a rating scale of 0–5 was used to classify the host pathogen interactions. Reactions 0, 1 and 2 were considered resistant and referred to as R0, R1 and R2 while 3, 4 and 5 were classified as susceptible (S). RUM lines (resistant to powdery mildew) and their derivatives are crossed with several susceptible (reaction types 3–5) genotypes and the inheritance of the resistance was studied in the F₁, F₂ and F₃ generations. The results showed that powdery mildew resistance in mungbean is governed by two dominant genes designated as Pm-1 and Pm-2. When both Pm-1 and Pm-2 were present, an R0 reaction was observed after inoculation with TI-1. The resistant reaction was R1 when only Pm-1 was present and R2 in the presence of Pm-2. In the absence of both Pm-1 and Pm-2, susceptible reactions 3, 4 and 5 were observed.     

Common Bean Reaction to Fusarium oxysporum f. sp. Phaseoli, the Cause of Severe Vascular Wilt in Central Africa

During the September‐December season of 1990, severe symptoms of Fusarium wilt were for the first time observed on a popular climbing bean (Phaseolus vulgaris L.) cultivar. G 2333. introduced within the previous 5 years. Seventy‐three bean genotypes were screened for resistance lo the disease, using artificial inoculation. The effect of inoculation density on the reaction of four selected genotypes was also investigated. Of the 29 climbing bean genotypes evaluated, 19 were resistant, including 11 of the 15 pre‐release or released cultivars. Of the 44 bush bean cultivars evaluated, 28 were resistant, five were intermediate and 11 were susceptible. All susceptible cultivars showed vascular discoloration. In both susceptible and resistant genotypes, the fungus spread almost equally from the entry points in inoculated roots to the base of the plants, but colonization and vertical spread within the vascular system were markedly less in resistant than in susceptible cultivars. At 20 and 30 cm above soil level, the fungus was only recovered from susceptible cultivars. Increasing inoculum density from 10² to 10⁷ conidia/ml did not affect the resistance of cultivars RWR 950 and G 685 but. in the susceptible cultivars G 2333 and MLB‐48‐89 A. it resulted in early appearance, high incidence and severity of the disease.     

籽用美洲南瓜叶片对白粉病菌的抗逆生理响应

以籽用美洲南瓜(Cucurbita pepo L.)白粉病抗病品系F2和感病品系M3为试材,在人工气候箱内接种白粉病生理小种2US孢子悬浮液,考察在接种白粉病菌后南瓜幼苗植株与白粉病菌的互作、叶片活性氧代谢及保护酶活性的变化,探讨南瓜抵御白粉病的生理机制。结果表明：(1)与感病品系M3相比,接种白粉病菌后,抗病品系F2叶片上病原菌发育缓慢,较难侵染叶片。(2)抗病品系F2在感病初期叶片H₂O₂、O₂^-·含量迅速升高后逐渐下降,而感病品系在感病初期H₂O₂、O₂^-·含量上升缓慢,在达最大值后始终保持较高水平,且感病品系叶片MDA含量始终高于抗病品系;组织化学染色分析发现,抗病品系叶片着色比感病品系快,之后着色面积有所减少并趋于较低水平。(3)抗病品系F2和感病品系M3叶片抗氧化酶CAT、SOD、POD活性及PAL、PPO活性在接种白粉病菌后均显著增加,但抗病品系的活性及其增幅均高于感病品系。研究发现,籽用美洲南瓜抗病品系叶片上白粉病菌发育缓慢,较难受到侵染,生成菌丝体后叶片上粉状斑点较小;抗病品系在被白粉病菌侵染初期依靠活性氧的增加抵御病原菌的入侵,随着活性氧含量增加抗病品系通过迅速增加自身抗氧化酶活性来防止氧化胁迫;与感病品系相比,抗病品系在受病原菌侵染后能迅速增加PAL、PPO活性以抵御病原菌侵染。     

Identification of the determinants of host resistantce and pathogenicity in interactions between Alternaria linicola Groves & Skolko and Linum Usitatissiumum L. accessions using multivariate analyses

Multivariate analysis of variance (MANOVA) and canonical variates analysis (CVA) were used to examine differences in host plant resistance and pathogen behaviour in interactions between Altemaria linicola and three genotypes of Linum usitatissimum, previously identified as susceptible, moderately resistant and resistant to the pathogen. Significant differences in pathogen development were found among the Linum accessions at 18, 24, and 40 h after inoculation. At 18 h after inoculation attempted penetration by the pathogen was relatively rare on all three accessions and canonical variates analysis revealed that overall differences among accessions resulted from large differences with respect to a small number of variables associated with successful penetration on the most susceptible accession. At later times after inoculation, when attempted penetration was more common, overall differences among accessions were found to result from smaller absolute differences among a group of variables which characterised the early colonisation of the host tissue. The results from these investigations are discussed in relation to recent research on the ecology of the pathogen and the importance of the timing of host responses to infection in determining host plant resistance.     

Variation for Resistance to <i>Alternaria tenuissima</i> and Potential Structural Mechanism among Different Cultivars of <i>Chrysanthemum morifolium</i>

Black spot disease, caused by the necrotrophic fungus Alternaria tenuissima (Fr.) Wiltsh (A. tenuissima), is considered a highly destructive disease of Chrysanthemum (Chrysanthemum morifolium Ramat.). A set of 17 accessions of commercial chrysanthemum cultivars were evaluated for resistance to A. tenuissima by seedling artificial inoculation. It was found that the reaction of the accessions to artificial inoculation ranged from resistant to highly susceptible. Five varieties of chrysanthemum (‘Zhongshan Taogui’, ‘Jinba’, ‘Zhongshan Jinguan’, ‘Jinling Wanhuang’ and ‘Jinling Yangguang’) were resistant; two varieties of chrysanthemum (‘Zhongshan Xinggui’ and ‘Zhongshan Jinkui’) were moderately resistant; and others were susceptible to various degrees, four varieties of chrysanthemum (‘Zhongshan Zihe’, ‘Zhongshan Jiuhong’, ‘Zaoyihong’ and ‘Jinling Jiaohuang’) were highly susceptible, especially. Some leaf morphological features of two resistant and two highly susceptible cultivars were further researched. Trichome density, length, height, gland size and stomata density were found to be associated with plant passive resistance. Resistant varieties that were identified in present study will be promising germplasm for exploitation of breeding programmes aimed at developing A. tenuissima-resistant cultivars and increasing genetic diversity.     

Dynamic reorganization of microtubules and microfilaments in flax cells during the resistance response to flax rust infection

The cytoskeleton in plant cells is a dynamic structure that can rapidly respond to extracellular stimuli. Alteration of the organization of microtubules and actin microfilaments was examined in mesophyll cells of flax, Linum usitatissimum L., during attempted infection by the flax rust fungus, Melampsora lini (Ehrenb.) Lev. Flax leaves that had been inoculated with either a compatible (yielding a susceptible reaction) or an incompatible (yielding a resistant reaction) strain of M. lini were embedded in butyl-methylmethacrylate resin; sections of this material were immunofluorescently labelled with anti-tubulin or anti-actin and examined using confocal laser scanning microscopy. In uninfected leaves, microtubules in the mesophyll cells formed a transverse array in the cell cortex. Microfilaments radiated through the cytoplasm from the nucleus. In an incompatible interaction, microtubules and microfilaments were extensively reorganized in mesophyll cells that were in contact with fungal infection hyphae or haustorial mother cells before penetration of the cell by the infection peg. After the initiation of haustorium development, microtubules disappeared from the infected cells, and growth of the haustoria ceased. In an incompatible interaction, hypersensitive cell death occurred in more than 70% of infected cells but occurred in less than 20% of cells in compatible interactions. After the infected cell had undergone hypersensitive cell death, the cytoskeleton in neighbouring cells became focused on the walls shared with the necrotic cell. In compatible interactions, reorganization of the cytoskeleton was either not observed at all or was observed much less frequently up to 48 h after inoculation.Abbreviations FITC fluorescein isothiocyanate - WGA wheatgerm agglutinin We thank Dr. G.J. Lawrence for providing valuable discussions and materials.     

Cell wall alterations in the leaves of fusariosis-resistant and susceptible pineapple cultivars

Fusariosis, caused by the fungus Fusarium subglutinans f. sp. ananas (Syn. F. guttiforme), is one of the main phytosanitary threats to pineapple (Ananas comosus var. comosus). Identification of plant cell responses to pathogens is important in understanding the plant–pathogen relationship and establishing strategies to improve and select resistant cultivars. Studies of the structural properties and phenolic content of cell walls in resistant (Vitoria) and susceptible (Perola) pineapple cultivars, related to resistance to the fungus, were performed. The non-chlorophyll base of physiologically mature leaves was inoculated with a conidia suspension. Analyses were performed post-inoculation by light, atomic force, scanning and transmission electron microscopy, and measurement of cell wall-bound phenolic compounds. Non-inoculated leaves were used as controls to define the constitutive tissue characteristics. Analyses indicated that morphological differences, such as cell wall thickness, cicatrization process and lignification, were related to resistance to the pathogen. Atomic force microscopy indicated a considerable difference in the mechanical properties of the resistant and susceptible cultivars, with more structural integrity, associated with higher levels of cell wall-bound phenolics, found in the resistant cultivar. p-Coumaric and ferulic acids were shown to be the major phenolics bound to the cell walls and were found in higher amounts in the resistant cultivar. Leaves of the resistant cultivar had reduced fungal penetration and a faster and more effective cicatrization response compared to the susceptible cultivar.     

18份广东香蕉种质对枯萎病的抗性评价

【背景】香蕉枯萎病是世界性的香蕉毁灭性病害,尚无有效药剂防控,筛选抗病品种是目前理想的防治方法。【方法】采用组培苗伤根接种法,研究了18份香蕉种质对香蕉枯萎病菌4号生理小种的抗性水平,并根据病情指数进行抗性分级。【结果】在供试的18份香蕉种质中,2份(东莞大蕉、抗枯5号)高抗,2份(碧盛、大丰)抗病,3份(抗枯1号、粉杂、农科1号)中抗,7份(粤优抗1号、广东-741、泰国B9、大蕉、台湾8号、海贡蕉、威廉斯8818)感病,4份(巴西、广东2号、广粉1号、粉蕉)高感。【结论与意义】不同香蕉种质对香蕉枯萎病菌4号生理小种的抗病性存在较大差异,本研究初步筛选出7份抗枯萎病的香蕉种质,为香蕉枯萎病抗病育种提供了依据,为病区种植香蕉品种提供了有效参考。     

Bioassay for in vitro differentiation of pineapple cultivar resistance levels to heart rot disease

Summary An in vitro bioassay to differentiate pineapple plant resistance levels to Phytophthora nicotianae var. parasitica (heart rot disease) is deseribed here. Conditions to cause death of in vitro-cultured plants were defined using a cultivar previously found to be susceptible to this fungus in our Field-Grown Pineapple Germplasm Bank (ev. Smooth Cayenne Serrana). The effects of zoospore concentration, inoculation technique, and disease progress during the course of time after infection were evaluated. The highest rates of plant death were observed with the use of 10⁸ zoospores ml⁻¹, and the inoculation technique of needle-mediated leaf base wound. One hundred percent plant death was observed at 144h after infection. Different susceptible varieties along with a resistant pineapple relative were additionally compared. In vitro results confirmed previous observations obtained under field conditions. The protocol described here may be used for early selection (in vitro) of new pineapple genotypes showing resistance to this fungus. At present, this protocol is extensively used in the Biotechnology-assisted Cuban Program for Pineapple Breeding.