Adhesive knob formation by conidia of the nematophagous fungusDrechmeria coniospora

We studied conidiogenesis and adhesive knob formation (maturation) by newly developed conidia of the nematophagous fungusDrechmeria coniospora. Upon conidiogenesis on infected nematodes or during saprophytic growth of the fungus in axenic cultures compact clusters of conidia developed. Less than 10% of such clustered conidia matured; mature conidia were invariably located on the periphery of the clusters.The kinetics and rate of maturation of conidia were studied inin vitro systems and in soil. In both cases adhesive knobs were formed; the rate at which knobs were formed appeared to be determined by the age of the conidia, the temperature and the soil moisture. In addition, knob formation was suppressed at increasing conidial densities. Under favorable conditions, however, over 90% of the conidia matured within a period of 3 days. The rate of knob formation was neither influenced by the presence of nematodes nor by that of exogenous nutrients, which suggests that maturation is an autonomous process. Electron-microscopical analysis indicated that budding of the conidia at the initial stage of maturation occurred simultaneously with the deposition of the sticky, adhesive layer around the wall of the developing knob.The ecological significance of the time- and spatially separated maturation of conidia after conidiogenesis is discussed with respect to survival of the conidia.     

Surface interactions of Fusarium graminearum on barley

The filamentous fungus Fusarium graminearum, a devastating pathogen of barley (Hordeum vulgare L.), produces mycotoxins that pose a health hazard. To investigate the surface interactions of F. graminearum on barley, we focused on barley florets, as the most important infection site leading to grain contamination. The fungus interacted with silica‐accumulating cells (trichomes and silica/cork cell pairs) on the host surface. We identified variation in trichome‐type cells between two‐row and six‐row barley, and in the role of specific epidermal cells in the ingress of F. graminearum into barley florets. Prickle‐type trichomes functioned to trap conidia and were sites of fungal penetration. Infections of more mature florets supported the spread of hyphae into the vascular bundles, whereas younger florets did not show this spread. These differences related directly to the timing and location of increases in silica content during maturation. Focal accumulation of cellulose in infected paleae of two‐row and six‐row barley indicated that the response is in part linked to trichome type. Overall, silica‐accumulating epidermal cells had an expanded role in barley, serving to trap conidia, provide sites for fungal ingress and initiate resistance responses, suggesting a role for silica in pathogen establishment.     

Occurrence of Phomopsis longicolla β Conidia in Naturally Infected Soybean

Although Phomopsis longicolla is primarily known as a seedborne pathogen, it can be isolated from all parts of the plant. The disease lesions observed on the basal parts of soybean stems were slightly sunken with irregular shapes and sizes, bordered by a thin black margin. Within the lesions themselves, large and diffusely distributed pycnidia with α and β conidia, typical of the genus Phomopsis, were observed. The percentages of the two types of conidia varied considerably, but β conidia were predominant in most of the pycnidia. The presence of these reproductive organs indicated that the symptoms could have been caused by Phomopsis sojae. However, after isolation on a nutritive medium, all cultural and morphological characteristics clearly indicated that the isolated fungus was P. longicolla, whose identification was subsequently confirmed by sequencing three genomic regions. Monosporic isolates, with different ratios of α and β conidia, exhibited a high level of pathogenicity on soybean, after artificial inoculation. Both types of conidia were observed on the stems of the inoculated soybean plants. Beta conidia also formed quickly on medium made of soybean seeds and mature stems after exposure to low temperatures (?10°C). This study suggests that P. longicolla is capable of a massive production of β conidia, not only in old fungal cultures as it had until now been believed, but also in infected soybean plants in the field.     

Taka-Amylase A in the Conidia of Aspergillus oryzae RIB40

A study of Taka-amylase A of conidia from Aspergillus oryzae RIB40 was done. During the research, proteins from conidia and germinated conidia were analyzed using SDS–PAGE, 2-D gel electrophoresis, Western blot analysis, MALDI-TOF Mass spectrometry, and native-PAGE combined with activity staining of TAA. The results showed that TAA exists not only in germinated conidia but also in conidia. Some bands representing degraded products of TAA were detected. Conidia, which formed on starch (SCYA), glucose (DCYA), and glycerol (GCYA) plates, contained mature TAA. Only one active band of TAA was detected after native-PAGE activity staining. In addition, TAA activity was detected in cell extracts of conidia using 0.5 M acetate buffer, pH 5.2, as extraction buffer, but was not detected in whole conidia or cell debris. The results indicate that TAA exists in conidia in active form even when starch, glucose, or glycerol is used as carbon source. TAA might belong to a set of basal proteins inside conidia, which helps in imbibition and germination of conidia.     

Perilipin LDP1 coordinates lipid droplets formation and utilization for appressorium-mediated infection in Magnaporthe oryzae

Lipid droplets (LDs) serve as one of the major reservoirs in conidia of Magnaporthe oryzae and are quickly utilized during appressorium formation. Here, we identified a gene, LDP1, encoding a perilipin that is important for LD formation and utilization during appressorium maturation. LDP1 is highly expressed in conidium and immature appressorium. Disruption mutants of LDP1 were significantly reduced in virulence, due to appressorial turgor reduction and difficulty in penetration. LDs were significantly reduced in the Δldp1 mutant, indicating LDP1 was required for LDs formation. LDP1 was colocalized with the LDs in conidium and immature appressorium but was gradually separated during appressorium maturation. A typical intracellular triacylglycerol lipase, TGL1-2, was clearly separated with LDs in conidium and immature appressorium but was well colocalized with LDs during appressorium maturation. The subcellular localization of TGL1-2 was affected by LDP1. These data suggested that LDP1 was bound to LDs for protecting from utilization in conidia and at the early appressorium stage but was separated from LDs for lipase entering and degradation. LDP1 was phosphorylated by CPKA at Thr96, which was essential for its localization and functions. These data indicate perilipin LDP1 can coordinate LD formation and utilization for appressorium-mediated infection of M. oryzae.     

Woronin bodies in arthrinium genus

Woronin bodies are present near all septal pores and in conidia of Arthrinium strains and may regulate cytoplasmic flow in both injured and actively growing healthy colonies. They vary in size and frequency, the central one plugs the septal pores in actively developing colonies and in mature conidia. The septa are thinner in the Woronin-body region.     

FINE STRUCTURE OF ANNELLOPHORES. I. SCOPULARIOPSIS BREVICAULIS AND S. KONINGII

Fine-structure observations of annelloconidium production in filamentous Hyphomycetes are reported for the first time. The difference in conidium morphology between Scopulariopsis brevicaulis and S. koningii was quite distinct. In S. brevicaulis, verrucosities appeared early in conidium ontogeny and formed an integral part of the primary wall layer of mature conidia. In S. koningii, verrucosities were absent. In S. brevicaulis, annellations did not invariably result on conidiophore necks with the production of each conidium in the basipetal sequence, but alternatively could be left on subapical regions of subsequently formed conidia. In S. koningii, annellations were more distinct, and the position of a conidium-delimiting septum was variable. If a septum were formed at a position proximal to previous septa, a portion of the annellophore neck remained attached to the base f the seceding conidium. In both species, a spherical electron-dense body, perhaps analogous to septal pore plugs in vegetative hyphae, plugged the pore between conidia and conidiophores and remained embedded in the base of seceded conidia.     

中国东部暖温带地区土壤中的皮司霉属（丝孢纲）真菌

报道从中国东部暖温带地区诸省（市）土壤中分离到的皮司霉属 Pithomyces 真菌的两个新种：长棒孢皮司霉 Pithomyces longiclavisporus和淡色皮司霉Pithomyces pallidus,及两个中国新记录种：卡罗皮司霉 Pithomyces karoo和帕夫皮司霉Pithomyces pavgii;对另外三个已知种亦作了分离地点和生境的引证。研究过的标本（干制培养物）与活菌种保存在山东农业大学植物病理学标本室（HSAUP）。     

Ultrastructural studies on zygomycotan fungi in the Zoopagaceae and Cochlonemataceae

The morphology of fungi in the Zoopagaceae and Cochlonemataceae (Zoopagales, Zoopagomycotina, Zygomycota) is reviewed, and some new ultrastructural information is added on conidia and zygospores, as well as haustoria in the former family and vegetative thalli in the latter. The cell wall of the conidia of Acaulopage dichotoma, Ac. tetraceros, Stylopage cephalote, Zoophagus insidians, and Zph. tentaclum (Zoopagaceae), and of Cochlonema odontosperma and Endocochlus gigas (Cochlonemataceae), is known to be composed of outer electron-dense and inner less dense layers in ultrathin sections, and no additional cell walls were found on the conidial cell wall. Although two nuclei were found in the zygosporangium before maturation to the zygospore in Acaulopage rhaphidospora (Zoopagaceae), more than one nucleus had never been observed previously in a zygospore in either of these families in ultrathin sections.     

Conidium differentiation in Aspergillus nidulans wild-type and wet-white (wetA) mutant strains

Conidium (asexual spore) differentiation in wild-type and the wet-white (wetA) mutant of Aspergillus nidulans was compared in intact chains of successively older conidia. Carbohydrate cytochemistry helped define three stages (Stages I, II, and III) of wild-type conidium maturation on the basis of changes in the ultrastructure and composition of the conidium wall. Conidia of the wetA6 mutant strain formed normally but failed to mature during Stages II and III. Specifically, the inner wall layer of wetA6 conidia did not condense during Stage II and two wall layers that stained for carbohydrates did not form during the transition to Stage III. Concomitantly, wetA6 conidia formed large cytoplasmic vacuoles and underwent lysis. The wetA gene appears to have a conidium-specific function for the modification of the conidium wall during Stages II and III. These modifications of the conidium wall are essential for the stability of mature, dormant conidia.     

Necrotrophic Mycoparasitism of Chalara elegans (Thielaviopsis basicola) by a Sterile Basidiomycete

Interactions between Chalara elegans and a sterile hyaline basidiomycete (SHB) were studied by light, scanning and transmission electron microscopy. Young colonies of C. elegans were quickly overgrown by the mycoparasite, with lysis of, hyphae and conidia. Direct penetration of host hyphae and conidia was observed, but simple coiling around these also occurred. Chlamydospore development was inhibited, but 12.5% of mature chlamydospores were able to survive attack for 120 days. Chlamydospore germination was apparently stimulated by the SHB, whereupon lysis followed. Evidence of invasion of intact chlamydospores is also presented.     

Collection of highly germinative pseudochain conidia of Oidium neolycopersici from conidiophores by electrostatic attraction

A population of simultaneously germinating conidia is an ideal inoculum of the powdery mildew pathogen, Oidium neolycopersici. In conditions of no or low wind velocity, O. neolycopersici successively stacks mature conidia on conidiophores in a chain formation (pseudochain), without releasing the precedent mature conidia. These pseudochain conidia represent a perfect inoculum, in which all conidia used for inoculation germinate simultaneously. However, we found that conidia must be collected before they fall to the leaf surface, because the germination rate was lower among conidia deposited on the leaf surface. We used an electrostatic spore collector to collect the pseudochain conidia, and their high germination rate was not affected by this treatment. The spore collector consisted of an electrified insulator probe, which created an electrostatic field around its pointed tip, and attracted conidia within its electric field. The attractive force created by the probe tip was directly proportional to voltage, and was inversely proportional to the distance between the tip and a target colony on a leaf. Pseudochain conidia were successfully collected by bringing the electrified probe tip close to target colonies on leaves. In this way, conidia were collected from colonies at 3-d intervals. This effectively collected all conidia from conidiophores before they dropped to the leaf surface. A high germination rate was observed among conidia attracted to the probe tip (95.5 ± 0.6 %). Conidia were easily suspended in water with added surfactant, and retained their germination ability. These conidia were infective and produced conidia in pseudochains on conidiophores after inoculation. The electrostatic spore collection method can be used to collect conidia as they form on conidiophores, thus obtaining an inoculum population in which all of the conidia germinate simultaneously.     

Effects of temperature and rainfall on date of release of ascospores of Leptosphaeria maculans (phoma stem canker) from winter oilseed rape (Brassica napus) debris in the UK

Data from a controlled environment experiment investigating effects of temperature on maturation of Leptosphaeria maculans pseudothecia were used to derive equations describing the times until 30% or 50% of pseudothecia were mature as a function of temperature. A wetness sensor was developed to estimate the oilseed rape debris wetness and operated with debris exposed in natural conditions in 2000 and 2001. The maturation of L. maculans pseudothecia on debris and concentrations of airborne L. maculans ascospores were observed from 1999 to 2004. There were considerable differences between years, with the first mature pseudothecia observed in September in most years. There were linear relationships between the first date when 10% of maximum ascospore release was observed and the dates when 30% or 50% of pseudothecia were mature. By summing the daily temperature‐dependent rate of pseudothecial maturation for days after 1 August with rainfall >0.5 mm, the dates when 30% or 50% of pseudothecia were mature were predicted. There was good agreement between predicted and observed dates when 30% or 50% of pseudothecia were mature. These equations for predicting the timing of L. maculans ascospore release could be incorporated into schemes for forecasting, in autumn, the severity of phoma stem canker epidemics in the following spring/summer in the UK.     

Chemical treatment of soil to prevent transmission of tobacco rattle virus to potatoes by Trichodorus spp

Alternaria longipes (Ell. &Ev.) Mason survived on autoclaved maize stems for 6 months without losing its pathogenicity, but rapidly lost viability on non-autoclaved stems and could not be re-isolated 4 months after inoculation. In laboratory tests it infected both living and dead maize leaves. Some Alternaria isolates from non-solanaceous hosts infected tobacco leaves kept at high humidities for 10 days after inoculation, but not when this incubation period was reduced to 48 h. In the field, perennation on plants other than tobacco is unlikely to be important as a source of inoculum. Pathogenicity of Alternaria isolates was maintained from one season to the next when stored as conidia in sterile soil, or as dried, infected tobacco leaves; some isolates maintained on agar slopes under oil were still pathogenic after 5 years. Alternaria conidia collected from the surface of tobacco seedlings, and isolates from apparently healthy seedling leaves were pathogenic to mature tobacco. In the field conidia were detected on tobacco leaves soon after these emerged, and epiphytic colonies were occasionally found well in advance of symptoms. Many latent infections were also detected up to 5 weeks in advance of symptoms. Visual development of latent infections closely coincided with the end of leaf expansion.     

How to enter the state of dormancy? A suggestion by Trichoderma atroviride conidia

It is generally accepted that conidia, propagules of filamentous fungi, exist in the state of dormancy. This state is defined mostly phenomenologically, e.g., by germination requirements. Its molecular characteristics are scarce and are concentrated on the water or osmolyte content, and/or respiration. However, a question of whether conidia are metabolic or ametabolic forms of life cannot be answered on the basis of available experimental data. In other words, are mature conidia open thermodynamic systems as are mycelia, or do they become closed upon the transition to the dormant state? In this article, we present observations which may help to define the transition of freshly formed conidia to the putative dormant forms using measurements of selected enzyme activities, ¹H- and ¹³C-NMR and LC-MS-metabolomes, and ¹⁴C-bicarbonate or ⁴⁵Ca²⁺ inward transport. We have found that Trichoderma atroviride and Aspergillus niger conidia arrest the ⁴⁵Ca²⁺ uptake during the development stopping thereby the cyclic (i.e., bidirectional) Ca²⁺ flow existing in vegetative mycelia and conidia of T. atroviride across the cytoplasmic membrane. Furthermore, we have found that the activity of α-ketoglutarate dehydrogenase was rendered completely inactive after 3 weeks from the conidia formation unlike of other central carbon metabolism enzymes. This may explain the loss of conidial respiration. Finally, we found that conidia take up the H¹⁴CO₃^- and convert it into few stable compounds within 80 d of maturation, with minor quantitative differences in the extent of this process. The uptake of H¹³CO₃^- confirmed these observation and demonstrated the incorporation of H¹³CO₃^- even in the absence of exogenous substrates. These results suggest that T. atroviride conidia remain metabolically active during first ten weeks of maturation. Under these circumstances, their metabolism displays features similar to those of chemoautotrophic microorganisms. However, the Ca²⁺ homeostasis changed from the open to the closed thermodynamic state during the early period of conidial maturation. These results may be helpful for studying the conidial ageing and/or maturation, and for defining the conidial dormant state in biochemical terms.     

MybA,a transcription factor involved in conidiation and conidial viability of the human pathogen Aspergillus fumigatus

Aspergillus fumigatus, a ubiquitous human fungal pathogen, produces asexual spores (conidia), which are the main mode of propagation, survival and infection of this human pathogen. In this study, we present the molecular characterization of a novel regulator of conidiogenesis and conidial survival called MybA because the predicted protein contains a Myb DNA binding motif. Cellular localization of the MybA::Gfp fusion and immunoprecipitation of the MybA::Gfp or MybA::3xHa protein showed that MybA is localized to the nucleus. RNA sequencing data and a uidA reporter assay indicated that the MybA protein functions upstream of wetA, vosA and velB, the key regulators involved in conidial maturation. The deletion of mybA resulted in a very significant reduction in the number and viability of conidia. As a consequence, the ΔmybA strain has a reduced virulence in an experimental murine model of aspergillosis. RNA‐sequencing and biochemical studies of the ΔmybA strain suggested that MybA protein controls the expression of enzymes involved in trehalose biosynthesis as well as other cell wall and membrane‐associated proteins and ROS scavenging enzymes. In summary, MybA protein is a new key regulator of conidiogenesis and conidial maturation and survival, and plays a crucial role in propagation and virulence of A. fumigatus.     

A serological investigation of hyphal growth in Fusarium culmorum

Summary Apical growth of hyphae of Fusarium culmorum was demonstrated using an immunofluorescent labelling technique. An antiserum prepared against hyphal tips contained a series of antibodies, detected by immunodiffusion, not present in antisera against mature hyphae or conidia. Absorption of the tip antiserum with hyphae allowed a specific immunofluorescence reaction with hyphal tips only. The antiserum against mature hyphae gave non-fluorescent tips to the hyphae.     

Factors affecting the development of Botrytis cinerea (grey mould) on linseed (Linum usitatissimum) buds, flowers and capsules

A key was produced to describe 10 stages of development of linseed buds, flowers and capsules. Botrytis cinerea conidia germinated more rapidly and germ tubes grew longer on linseed stigmas, petals and mature senescing capsules than on green leaves, sepals and immature capsules. The proportion of conidia which germinated increased and the germ tubes continued growing for longer in the presence of linseed pollen and flower petal extracts. In controlled environment and field experiments, the response of buds, flowers and capsules to inoculation with B. cinerea changed with stage of development; few pre‐flowering buds developed symptoms (brown lesions, then grey mould), but high proportions of flowering and post‐flowering buds did so. Few immature green capsules developed symptoms and the proportion of capsules which developed symptoms increased as they matured. The presence of linseed pollen decreased the incubation period from inoculation with spore suspensions to appearance of B. cinerea symptoms on buds. A disease cycle was produced to suggest the changes in susceptibility of linseed to infection by B. cinerea conidia during bud, flower and capsule development.