Infection process of Colletotrichum destructivum O'Gara from lucerne (Medicago sativa L.)

The infection process of Colletotrichum destructivum, a cause of anthracnose in lucerne (= alfalfa, Medicago sativa) was studied by light microscopy. At the onset of the host-pathogen interaction, the fungus produced large, multilobed, multiseptate infection vesicles with elongated neck regions. Each infection structure packed the lumen of the initially-infected epidermal cell and remained confined within its walls for 48 h. Subsequently, narrow, invasive secondary hyphae radiated from the multilobed vesicles, grew through the walls of host cells and rapidly colonized the surrounding tissues. Acervuli emerged on the surface of colonized leaves 96 h after inoculation. These observations are discussed in relation to the infection process and specificity of a genetically closely-related isolate of Colletotrichum destructivum causing anthracnose in cowpea (Vigna unguiculata).     

Subcuticular-Intracellular Hemibiotrophic and Intercellular Necrotrophic Development of Colletotrichum acutatum on Almond

ABSTRACT The early infection and colonization processes of Colletotrichum acutatum on leaves and petals of two almond cultivars with different susceptibility to anthracnose (i.e., cvs. Carmel and Nonpareil) were examined using digital image analysis of light micrographs and histological techniques. Inoculated tissue surfaces were evaluated at selected times after inoculation and incubation at 20 degrees C. Depth maps and line profiles of the digital image analysis allowed rapid depth quantification of fungal colonization in numerous tissue samples. The results showed that the early development of C. acutatum on petals was different from that on leaf tissue. On petals, conidia germinated more rapidly, germ tubes were longer, and fewer appressoria developed than on leaves. On both tissues, penetration by the pathogen occurred from appressoria and host colonization was first subcuticular and then intracellular. On petals, colonizing hyphae were first observed 24 h after inoculation and incubation at 20 degrees C, whereas on leaves they were seen 48 to 72 h after inoculation. Intercellular hyphae were formed before host cells became necrotic and macroscopic lesions developed on petals >/=48 h and on leaves >/=96 h after inoculation. Histological studies complemented data obtained by digital image analysis and showed that the fungus produced infection vesicles and broad hyphae below the cuticle and in epidermal cells. In both tissues, during the first 24 to 48 h after penetration fungal colonization was biotrophic based on the presence of healthy host cells adjacent to fungal hyphae. Later, during intercellular growth, the host-pathogen interaction became necrotrophic with collapsed host cells. Quantitative differences in appressorium formation and host colonization were found between the two almond cultivars studied. Thus, on the less susceptible cv. Nonpareil fewer appressoria developed and host colonization was reduced compared with that on cv. Carmel.     

Light and scanning electron microscopy studies on the infection process of melon leaves by Colletotrichum lagenarium

Colletotrichum lagenarium is the casual agent of anthracnose disease of melons. Light and scanning electron microscopy were used to observe the infection process of C. lagenarium on the leaves of two melon cultivars differing in susceptibility. On both cultivars conidia began germinating 12 h after inoculation (hai), forming appressoria directly or at the tips of germ-tubes. By 48 hai appressoria had melanised and direct penetration of host tissue had begun. On the susceptible cultivar, infection vesicles formed within 72 hai and developed thick, knotted primary hyphae within epidermal cells. By 96 hai C. lagenarium produced highly branched secondary hyphae that invaded underlying mesophyll cells. After 96 hai, light brown lesions appeared on the leaves, coincident with cell necrosis and invasion by secondary hyphae. While appressoria formed more quickly on the resistant cultivar, fewer germinated to develop biotrophic primary or invasive necrotrophic secondary hyphae than on the susceptible cultivar. These results confirm that C. lagenarium is a hemibiotrophic pathogen, and that resistance in melons restricts colonisation by inhibiting the development of necrotrophic secondary hyphae.     

Novel infection strategies of Colletotrichum acutatum on ripe blueberry fruit

The infection and colonization process of Colletotrichum acutatum on ripe blueberry fruit from two cultivars with different susceptibility to anthracnose were examined using light and confocal laser scanning microscopy. Ripe fruit from susceptible cv. Jersey and resistant cv. Elliott were drop-inoculated with a conidial suspension of C. acutatum, and epidermal peels were evaluated at selected times after inoculation and incubation. Results from pre-penetration studies demonstrated that there were significant differences in the rate of formation of melanized appressoria between the two cultivars, with the rate of formation being faster in the susceptible one. In both cultivars, penetration by the pathogen occurred via appressoria 48 h post-inoculation (hpi). However, in the susceptible cv. Jersey, C. acutatum then adopted an intracellular hemibiotrophic-like infection strategy, whereas in the resistant cv. Elliott subcuticular intramural-like infection occurred. In cv. Jersey by 108 hpi, intracellular growth of the pathogen led to the formation of numerous acervuli, with orange conidial masses. By 120 hpi, the conidial masses had coalesced covering the entire inoculated area. In cv. Elliott, acervuli were not seen until 144 hpi and contained few conidia. These results demonstrate for the first time the ability of C. acutatum to adopt a different infection and colonization strategy depending on the susceptibility of the host tissue being colonized.     

Histological and ultrastructural characterization of the leaf infection events of Colletotrichum fructicola on Malus domestica ‘Gala’

Glomerella leaf spot (GLS), characterized by black necrotic spots and severe defoliation, is a destructive foliar disease of apple. Widely grown cultivars such as Gala and Golden Delicious are highly susceptible to GLS. Currently, the infection biology of the causal pathogen, Colletotrichum fructicola, on apple leaves is unclear. In the present study, the penetration and colonization processes of C. fructicola were characterized on apple (cv. Gala) leaves using light and transmission electron microscopy. C. fructicola conidia produced germ tubes 4 hours post-inoculation (hpi) and appressoria at 8 hpi. In melanized appressoria, funnel-shaped appressorial cones formed around the penetration pore. At 12 hpi, C. fructicola produced secondary conidia. After penetration, C. fructicola began to develop infection vesicles at 36 hpi. At 48 hpi, the primary hyphae of C. fructicola were produced from infection vesicles within host epidermal cells; the host epidermal cell plasma membrane remained intact, indicating a biotrophic association. Subsequently, secondary hyphae penetrated epidermal cells and destroyed cell components, initiating necrotrophic colonization. C. fructicola also produced biotrophic subcuticular infection vesicles and hyphae. Together, these results demonstrate that C. fructicola forms special infection structures and colonizes apple leaves in a hemibiotrophic manner, involving intracellular as well as subcuticular colonization strategies. Detailed characterization of the infection process of C. fructicola on apple leaves will assist in the development of disease management strategies and provide a foundation for studies of the molecular mechanism of the C. fructicola–apple leaf interaction.     

Infection process of <Emphasis Type="Italic">Gilbertella persicaria</Emphasis> in papaya (<Emphasis Type="Italic">Carica papaya</Emphasis> L.) fruits

Gilbertella persicaria is a pathogenic fungus recently reported as a causative agent of soft rot in papaya fruits. Here the interactions between G. persicaria and papaya fruits was analyzed under laboratory conditions using histological techniques and optical microscopy to elucidate the process of pathogenesis. Healthy and disinfested fruits of papaya cv. Maradol were also inoculated with a suspension of sporangiospores of G. persicaria. Tissue sections were cut, which were subjected to differential staining with safranin-fast green for different times. Sporangiospores presumably adhered to the cuticle of the fruit by 3 h post inoculation (hpi) and germinated by 6 hpi; invasive intracellular hyphae were growing in host cells by 9 hpi. By 15 hpi, fruit epidermis was macerated, presumably by enzymatic activity reported for mucoral fungal species and appeared as a wet-looking lesion on the cuticle. Fruit mesocarp was colonized by 30 hpi, and asexual reproduction structures had formed by 48 hpi. This process of infection and disease development of G. persicaria in papaya fruits corresponds to that used by pathogens with a necrotrophic lifestyle.     

Effect of cold‐induced changes in physical and chemical leaf properties on the resistance of winter triticale (×Triticosecale) to the fungal pathogen Microdochium nivale

This study showed that several mechanisms of the basal resistance of winter triticale to Microdochium nivale are cultivar‐dependent and can be induced specifically during plant hardening. Experiments and microscopic observations were conducted on triticale cvs Hewo (able to develop resistance after cold treatment) and Magnat (susceptible to infection despite hardening). In cv. Hewo, cold hardening altered the physical and chemical properties of the leaf surface and prevented both adhesion of M. nivale hyphae to the leaves and direct penetration of the epidermis. Cold‐induced submicron‐ and micron‐scale roughness on the leaf epidermis resulted in superhydrophobicity, restricting fungal adhesion and growth, while the lower permeability and altered chemical composition of the host cell wall protected against tissue digestion by the fungus. The fungal strategy to access the nutrient resources of resistant hosts is the penetration of leaf tissues through stomata, followed by biotrophic intercellular growth of individual hyphae and the formation of haustoria‐like structures within mesophyll cells. In contrast, a destructive necrotrophic fungal lifestyle occurs in susceptible seedlings, despite cold hardening of the plants, with the host epidermis, mesophyll and vascular tissues being digested and becoming disorganized as a result of the low chemical and mechanical stability of the cell wall matrix. This work indicates that specific genetically encoded physical and mechanical properties of the cell wall and leaf tissues that depend on cold hardening are factors that can determine plant resistance against fungal diseases.     

Infection of apple fruit by Sphaeropsis pyriputrescens in the orchard in relation to Sphaeropsis rot in storage

Sphaeropsis rot, caused by Sphaeropsis pyriputrescens, is an important postharvest disease of apple in the United States. The objectives of this study were to determine the timing of apple fruit infection in the orchard in relation to development of Sphaeropsis rot in storage and to identify infection courts and mode of penetration by S. pyriputrescens on apple fruit. Fruit of apple cvs Red Delicious, Golden Delicious, and Fuji were inoculated in the orchard from 3 weeks after petal fall to 2 weeks before harvest at 5 to 6-week intervals in three consecutive seasons. All fruit were harvested and stored at 0?ºC to monitor decay development. Light and scanning electron microscopy were used to examine the infection courts and mode of penetration of the fungus on/in the host tissues. At harvest, the fungus was re-isolated from the stem (pedicel), sepal, anther, or filament of the inoculated fruit, but decay did not develop on fruit. Sphaeropsis rot developed on inoculated fruit during cold storage beginning 1–3 months after harvest. Stem-end rot was prevalent on cv. Golden Delicious, whereas calyx-end rot was prevalent on cv. Fuji. Both stem- and calyx-end rots were common on cv. Red Delicious. Infection also occurred at lenticels on fruit skin, particularly on cv. Golden Delicious, but at low incidence. Relationships between the incidence of Sphaeropsis rot in stored apple fruit and the timing of inoculation in the orchard varied with cultivar and year. On cv. Red Delicious apples, the incidence of Sphaeropsis rot generally increased as the timing of infection approached harvest. Histological studies indicated that infection took place through natural openings of plant organs such as stomata on stems and sepals and lenticels on fruit skin. Fungal penetration also was observed at micro-cracks on the stem and sepal and at trichome sockets where mechanical damage occurred in sepals. Direct penetration was observed on the stem and sepal of fruit, but most invasions were restricted between the cuticle and the epidermis. Our results indicate that wounding is not required for infection of apple fruit by S. pyriputrescens, though it may facilitate infections.     

Latency- and Defense-Related Ultrastructural Characteristics of Apple Fruit Tissues Infected with Botryosphaeria dothidea

ABSTRACT Apple fruit tissues infected with Botryosphaeria dothidea were examined by transmission electron microscopy using susceptible cv. Fuji and resistant cv. Jonathan. Immature (green) and mature (red) fruits of cv. Fuji with restricted or expanding lesions were also examined to reveal subcellular characteristics related with latent and restricted disease development. In infected susceptible mature fruits, cytoplasmic degeneration and organelle disruption commonly occurred, accompanying cell wall dissolution around invading hyphae. Cell wall dissolution around invading hyphae in subepidermis was rare in immature, red halo-symptomed cv. Fuji and resistant cv. Jonathan fruits. In infected immature fruits of cv. Fuji, presumably at the latent state of disease development, cellular degeneration was less severe, and invading hyphae contained prominent microbody-lipid globule complexes or the deposition of thin electron-dense outer layer around cell wall of intercellular hyphae. Both mature fruits with red halos and resistant apple fruits formed cell wall protuberances at the outside of cell walls. In addition, electron-dense extramural layers were formed in the resistant apple fruits. Aberrant hyphal structures such as intrahyphal hyphae were found only in resistant fruit tissues, indicating the physiologically altered fungal growth. These ultrastructural changes of host tissues and fungal hyphae may reflect the pathogenesis of apple white rot under varying conditions of apple fruits.     

The development of different pathotype groups of Mycosphaerella pinocles in susceptible and partially resistant pea leaves

The progress of infection by high- and low-virulence isolates of Mycosphaerella pinodes was examined in susceptible and partially resistant pea leaves. Conidia germinated with one or more germ tubes which frequently branched and formed appressorium-like structures on the leaf surface. Penetration occurred through the epidermal walls. A structure similar to an infection vesicle was formed, lying partly in the epidermal wail and partly in the cell lumen. From this structure, a penetration hypha was derived which initiated the development of the intra- and intercellularly-growing fungal colony. Infections led to rapid tissue collapse in both susceptible and resistant interactions. In resistant interactions, the formation of infection vesicles and penetration hyphae was reduced, and the development and spread of lesions was retarded.     

Structural Modifications and Programmed Cell Death of Chili Pepper Fruit Related to Resistance Responses to Colletotrichum gloeosporioides Infection

ABSTRACT Postharvest (detached) and in planta (attached) fruits of pepper plants, Capsicum annuum cv. Jejujaerae (susceptible) and Capsicum baccatum cv. PBC80 (resistant), inoculated with the anthracnose pathogen Colletotrichum gloeosporioides were examined using light, confocal laser scanning, and electron microscopy to compare the cytological differences between the compatible and incompatible interactions. In nonwound inoculation of postharvest pepper fruit, resistant pepper tissues showed a significant increase in the thickness of the cuticle layer compared with that of the susceptible and noninoculated fruit. Cytological features of programmed cell death (PCD) were observed in the resistant pepper fruit with postharvest inoculation, and these were characterized by positive responses to terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling. The oligonucleosomal fragments of DNA were confirmed electrophoretically as DNA laddering. The PCD-positive responses occurred around the inoculation sites early in in planta wound inoculation in the resistant pepper. Nuclear modifications and structural changes of hypersensitivity were also observed in the resistant fruit, including separation of the plasma membrane from the cell wall, dilation of the endoplasmic reticulum, accumulation of electron-dense inclusions in vacuoles, and cytoplasmic vacuolization accompanying fragmentation of the cytoplasm. These structural changes may also implicate PCD-like host responses. In addition, in planta wound inoculation resulted in cell enlargement and cell division during the later stages of infection to form a periderm-like boundary layer around the inoculation site.     

Pathogenic variation and virulence related responses of <Emphasis Type="Italic">Ascochyta lentis</Emphasis> on lentil

Ascochyta blight of lentil (Lens culinaris ssp. culinaris) is caused by Ascochyta lentis. The disease causes severe damage to all aerial parts of the plant and may lead to total crop loss during extremely severe epidemics. To identify qualitative differences in resistance within Australian lentil crops, variation in virulence was examined among 17 isolates of A. lentis on six differential lentil genotypes (ILL7537, ILL5588 (cv. Northfield), ILL6002, ILL5722 (cv. Digger), ILL481 (cv. Indianhead) and CIPA203 (cv. Nipper)). Six distinct virulence patterns were identified, with Pathotype I (AL4) being highly virulent, causing disease on all genotypes except ILL7537 and pathotype VI (Kewell) exhibiting low virulence on all genotypes. Histopathology studies were carried out to further understand interaction differences between isolate-host combinations and add to the knowledge of possible resistance mechanisms underlying lentil’s defence to the pathogen. The infection process was compared between lentil genotypes with different levels of resistance and isolates with different levels of virulence. Microscopic and biochemical differences were observed between compatible and incompatible interactions, which were related to time-after-inoculation, with slower responses noted in susceptible lentil genotypes. Relatively fast release of reactive oxygen species (ROS) and a subsequent hypersensitive response (HR) was central to initial defence at the point of penetration in the most resistant lentil genotypes.     

青枯菌(Pseudomonas solanacearum)在番茄抗、感病品种根部的吸附、侵入和繁殖

 采用不同的方法接种和利用扫描、透射电镜观察,研究不同致病力的青枯菌对番茄抗病及感病品种根部的吸附、侵入与繁殖。发现番茄抗病品种与感病品种的植株体内在菌体数量上有明显差异,而与青枯菌对番茄根部的吸附关系不显著。电镜观察发现青枯菌强致病力菌株菌体能以游离的形式存在于番茄感病品种根部的细胞间隙中,并能降解植株细胞壁、破坏原生质膜;青枯菌强致病力菌株菌体在抗病品种根内和青枯菌强致病力菌株在抗病及感病品种根内均被番茄植株细胞壁吸附,并且被细胞壁周围的浓密物质所包围。     

Germination and invasion by ascospores and pycnidiospores of <Emphasis Type="Italic">Leptosphaeria maculans</Emphasis> on spring-type <Emphasis Type="Italic">Brassica napus</Emphasis> canola varieties with varying susceptibility to blackleg

The infection processes of ascospores and pycnidiospores of Leptosphaeria maculans were studied on cotyledons of six cultivars of spring-type Brassica napus: one with resistance controlled by a single dominant gene (cv. Surpass 400), three with polygenic resistance (cvs. Dunkeld, Grouse, and Outback), and two susceptible cultivars (Westar and Q2). On all cultivars, ascospore germination, penetration, and development of symptoms on cotyledons were much earlier than that with pycnidiospores. At 2h after inoculation ascospores began to germinate, by 4h about 50% had germinated, and by 6–8h 85%–90% had germinated. In contrast, pycnidiospores began to germinate 1 day after inoculation (dai) and reached only 50% germination by 3 dai. Ascospores began germinating from terminal cells and then later from the interstitial cells. Pycnidiospores germinated predominantly from one end and sometimes from both ends. Germ tubes from ascospores penetrated stomata as early as 4h after inoculation, whereas those from pycnidiospores penetrated at 2 dai. Symptom development with ascospores was 2 days earlier than that with pycnidiospores. Symptoms on Surpass 400 were evident as early as 3–5 dai with ascospores and 5–7 dai with pycnidiospores. However, on other cultivars, symptoms were not evident until 10 dai with ascospores and 12 dai with pycnidiospores. This report is the first on differences in the infection processes by the two spore types. Ascospore and pycnidiospore attachment, germination, and penetration did not differ between resistant and susceptible cultivars, but there were major differences after penetration. Under high humidity, 80%–90% of stomata of susceptible Westar and Q2 had aerial hyphae emerging from stomatal pores. However, fewer stomata (5%–10%) had aerial hyphae on Surpass 400 by 10 dai with ascospores and 12 dai with pycnidiospores, but even these were usually poorly developed. Host differences in spring-type B. napus in relation to production of aerial hyphae have not previously been reported. In Surpass 400, rapid necrosis of guard cells occurred within a few hours of penetration by either type of spore, and subsequently one or a few cells immediately adjacent to the penetration site died. This necrosis then spread to the cells around the penetration site to form a hypersensitive response (in the form of a small, dark lesion) to both ascospores and pycnidiospores. This is the first detailed report on interactions between spring-type B. napus and L. maculans in relation to single dominant gene-based resistance. Neither the cultivars with polygenic resistance nor the susceptible cultivars had such a response.     

Efficacy of citronella essential oil for the management of chilli anthracnose

The potential of citronella essential oil for the management of chilli anthracnose caused by Colletotrichum acutatum was investigated. In in vitro tests, citronella essential oil inhibited mycelial growth at 0.25, 1.25 and 2.5 μL citronella/mL water, reduced conidial germination, and inhibited germ tube elongation at 1.25 μL/mL. Citronella essential oil applied as a protective or curative treatment to chilli cv. Django fruits reduced anthracnose infection. Efficacy of citronella essential oil was substantially effective to chemical fungicides and relatively superior to a biofungicide in both protective and curative tests. However, citronella essential oil at 2.5 μL/mL was phytotoxic to chilli fruits. Therefore, the most effective rate of citronella essential oil was at 1.5 μL/mL with inhibition of pathogen growth, reduction of anthracnose symptoms, and no observable phytotoxic response on chilli fruits. Citronella essential oil may be a viable alternative to chemical fungicides for the management of chilli anthracnose.     

Subcelullar localization of proteins associated with <Emphasis Type="Italic">Prune dwarf virus</Emphasis> replication

The fungus Colletotrichum lindemuthianum is the causal agent of anthracnose, one of the most severe diseases of the common bean (Phaseolus vulgaris). The infection process begins with the adhesion of conidia to the plant’s surface. Appressoria are then formed, allowing penetration of the fungus. Next, the biotrophic phase begins, followed by the necrotrophic phase. Due to the peculiar nutrition mode of the fungus, including both of the previously mentioned stages, it is of great interest to determine which genes are involved in the transition between the two phases during the infection process. To determine this, suppression subtractive hybridization (SSH) was used in association with qRT-PCR in the present study. These methods allowed for the identification of genes that were differentially expressed at each developmental stage of the fungus in the plant. This is the first report on the use of the cited techniques to evaluate the infectious cycle of the fungus. A total of 175 sequences exhibited significant identity (e?≤?10^?5) with sequences present in the sequenced genomes of P. vulgaris and C. lindemuthianum; approximately 41 % of those were determined to belong to the fungus, and 59 % were determined to belong to the plant. Of the predicted sequences, 68 % were of unknown function or were not found in the databases. Among the analyzed expressed sequence tags (ESTs), sequences were found that encode proteins related to: primary and secondary metabolism; the transport of different compounds; the degradation/modification of proteins; cell regulation and signaling; cellular stress response; and the degradation of exogenous compounds. The obtained results allowed for the identification of sequences encoding proteins that are essential for the progression of anthracnose. Furthermore, it was possible to identify new genes, the functions of which have not yet been described, and even to identify unique genes of C. lindemuthianum that are involved in the pathogenicity and virulence of this fungus.     

Characterization and pathogenicity of Colletotrichum species associated with anthracnose on chilli (Capsicum spp.) in Thailand

Fungal isolates from chilli ( Capsicum spp.) fruits in Thailand that showed typical anthracnose symptoms were identified as Colletotrichum acutatum , C . capsici and C . gloeosporioides . Phylogenetic analyses from DNA sequence data of ITS rDNA and β-tubulin ( tub 2) gene regions revealed three major clusters representing these three species. Among the morphological characters examined, colony growth rate and conidium shape in culture were directly correlated with the phylogenetic groupings. Comparison with isolates of C . gloeosporioides from mango and C . acutatum from strawberry showed that host was not important for phylogenetic grouping. Pathogenicity tests validated that all three species isolated from chilli were causal agents for chilli anthracnose when inoculated onto fruits of the susceptible Thai elite cultivar Capsicum annuum cv. Bangchang. Cross-infection potential was shown by C . acutatum isolates originating from strawberry, which produced anthracnose on Bangchang. Interestingly, only C . acutatum isolates from chilli were able to infect and produce anthracnose on PBC 932, a resistant genotype of Capsicum chinense . This result has important implications for Thai chilli breeding programmes in which PBC 932 is being hybridized with Bangchang to incorporate anthracnose resistance into chilli cultivars.     

Fine Structure of the Early Interaction of Lily Roots with Fusarium oxysporum f.sp. lilii

The early interaction of lily roots with the cortical rot pathogen Fusarium oxysporum f.sp. lilii was studied using roots of lily bulblets grown in Hoagland's solution, inoculated with the pathogen, and sampled up to 48h later. Conidia produced germ tubes within 6h, which extended towards and into the mucilage covering the root elongation zone, and along and into the anticlinal grooves and middle lamellae of epidermal cells. By 24–48h, infecting hyphae had reached the periclinal walls and intercellular spaces between the epidermis and the outermost cells of the cortex. Penetration of intercellularly growing hyphae directly across host cell walls was not observed; invasion of the cell lumen only occurred by gradual infringing of hyphae upon successive primary wall layers. Non-cellulosic wall appositions rich in vesicles and covered by a cellulosic protective-like layer were formed in response to approaching hyphae in resistant cv.Connecticut King, but rarely in susceptible cv. Esther which seemed more susceptible to plasmolysis and rot. Finger-like projections of the appositions into the host cell cytoplasm likely represent early stages of transfer cell formation.     

Histological investigation of stripe rust (Puccinia striiformis f.sp. tritici) development in resistant and susceptible wheat cultivars

The wheat cultivar Kariega expresses complete adult plant resistance against stripe rust, whereas cv. Avocet S is susceptible. Using confocal laser scanning microscopy, initial fungal penetration into flag leaves was identical in both cultivars, with directional germ-tube growth towards stomata that were penetrated without the formation of an appressorium, followed by differentiation of a substomatal vesicle, infection hyphae, haustorial mother cells and haustoria. During the following 4 days, further fungal development occurred more quickly in the resistant than in the susceptible cultivar. However, by 7 days postinoculation (dpi) the situation changed, with exponential growth of the pathogen occurring only in the susceptible line. Induced cellular lignification, a typical defence reaction of cereals, was observed at 4 dpi in the resistant cultivar, and 2 days later lignified tissue completely surrounded the fungal colonies. In the susceptible cultivar, isolated lignified host cells occurred at 6 dpi, and long, unbranched fungal hyphae outgrowing the resistance reaction were observed.