Scanning Electron Microscopy on the Perithecial Development of Phyllactinia corylea on Mulberry-II. Sexual Stage

Development of perithecia of Phyllactinia corylea (Pers.) Karst. on mulberry (Morus spp.) leaves was examined by scanning electron microscopy. Two short specialized structures, antheridium and ascogonium, emerged from two separate hyphae, were fused with each other and developed into an egg‐shaped perithecial primordia. These primordia later developed into globose immature perithecia, which covered with protruded wall cells with clear margins. A large number of hyphae emerged from near the base of globose perithecia, which radiated on the leaf surface and thus helped the perithecia to fix to the surface. Specific characteristic penicillate cells and acicular appendages originated from the immature perithecia. The penicillate cells developed with apical sterigma‐like projections from the wall cells of the upper part of immature perithecia, whereas the acicular appendages originated from the shrunken wall cells at the perithecial equatorial planes. On maturation of perithecia, the acicular appendages bent down and pushed the perithecia above the substrate and thus helped them to liberate out. The sterigma‐like projections were covered with paste‐like granular substance, which help the dispersed perithecia to attach to mulberry leaves and branches.     

Scanning electron microscopy of surface and internal features of developing perithecia of Neurospora crassa.

Stages in the development of perithecia of Neurospora crassa, designated by the time elapsed after crossing, were investigated with the scanning electron microscope, from protoperithecia through perithecia. The usual examination of external features of whole specimens with this instrument was augmented by a freeze-fracture technique which allowed the viewing of development internally as well. Rapid increases in perithecial size soon after crossing were followed by the appearance, in section, of a centrum, at first undifferentiated but subsequently developing ascogenous hyphae. The perithecial beak appeared as a compact mass easily distinguishable in whole specimens from the surrounding hyphae by means of texture as well as shape. Two ascospores were photographed during emergence from an ostiole, but ostioles were found more frequently closed than open.     

PERITHECIAL FORMATION IN GELASINOSPORA RETICULISPORA. I. EFFECTS OF LIGHT AT TWO DIFFERENT GROWTH STATES

Hyphae of Gelasinospora reticulispora were cultured on corn meal agar in a growth tube at 25 ± 0.4°C under different light conditions. While the hyphal tip was growing, perithecia were not formed under continuous white light (ca. 2000 ergs cm^?2 sec^?1), but some perithecia were initiated in total darkness. However, when white light was given after a dark period, perithecial formation was greatly promoted. In these cases, perithecial formation occurred in older portion of the culture (the portion nearest the point of inoculation) at first, and then gradually spread to the younger portion. Immediately after the tip of hyphae reached the other end of the growth tube, perithecia were induced in the youngest portion of the hyphae irrespective of the photoconditions; then formation proceeded toward the older portion. This induction was not age-dependent, because in growth tubes with different lengths, perithecia always became visible ca. 24 hr after the tip of hyphae reached the other end of growth tube. The photoinhibitory effect was no longer observed thereafter, but photopromotive effect was still evident.     

Methods of investigating populations of the take-all fungus (Ophiobolus graminis) in soil

Attempts to isolate Ophiobolus graminis directly from infested soils failed, so host-infection techniques were used to study soil-borne populations of the fungus. Extracting organic debris from soils and grading it by wet sieving through standard meshes concentrated the fungus. Fractions were tested for infectivity either as layers in pots of sand or by packing into short lengths of polyvinyl chloride tubing, through which wheat seedlings were grown. Coarse debris (retained by 420 μ aperture sieves) was most infectious and usually caused lesions within 3 weeks; whole soil and especially fine debris (< 420 μ) caused fewer and less severe infections, which often became apparent only after 3 weeks. Slight infectivity of the sediment fractions was attributed to imperfect separation of debris. Soil sampled in crop or stubble rows caused more infections than soil from between rows. Usually seedling infection was made manifest by root lesions and runner hyphae, but these symptoms were not always plain or conclusive. Many seedling roots that rotted when kept moist and in the light produced perithecia within 6 weeks. Although perithecia formed on some roots where neither lesions nor hyphae were found, they did not form on all diseased roots. More needs to be known about the factors influencing perithecial formation before it can be used as a reliable confirmatory test.     

板栗褐缘叶枯病协同致病菌Ophiognomonia castaneae的生活史

采用室外定点观察,子实体诱导及rDNA ITS、MS204、tef1-α 3种分子标记进行系统发育分析等方法,对板栗褐缘叶枯病Phomopsis castaneae-mollissimae的协同致病菌板栗蛇孢日规壳Ophiognomonia castaneae的生活史进行了研究。结果表明,每年7月下旬至8月初叶片发病初期很少分离到O. castaneae,随着病斑扩大该菌的分离频率逐渐增大,至发病后期其分离频率可高达78.5%,甚至可超过致病菌P. castaneae-mollissimae,10月下旬板栗落叶背面的病斑上开始形成O. castaneae的分生孢子盘,11月下旬开始形成O. castaneae的子囊壳原基,次年5、6月越冬落叶背面的病斑上长出子囊壳;带病斑的叶片经室内外诱导,0-25℃范围均可产生成熟子囊壳;湿度是决定子囊壳能否形成的关键因素,强光照不利于子囊壳的产生;分离物的菌丝体在PDA培养基上培养,易产生分生孢子;将分离物分为两种交配型,相互交配后6个月所有处理均未长出该菌的有性型子实体。室外定点观察及rDNA ITS、MS204、tef1-α 3种分子标记表明分离物和病斑上的子囊孢子及其萌发菌丝为O. castaneae的不同生长发育阶段。     

Perithecium morphogenesis in Sordaria macrospora

The perithecium of the self-fertile ascomycete Sordaria macrospora provides an excellent model in which to analyse fungal multicellular development. This study provides a detailed analysis of perithecium morphogenesis in the wild type and eight developmental mutants of S. macrospora, using a range of correlative microscopical techniques. Fundamentally, perithecia and other complex multicellular structures produced by fungi arise by hyphal aggregation and adhesion, and these processes are followed by specialization and septation of hyphal compartments within the aggregates. Perithecial morphogenesis can be divided into the ascogonial, protoperithecial, and perithecial stages of development. At least 13 specialized, morphologically distinct cell-types are involved in perithecium morphogenesis, and these fall into three basic classes: hyphae, conglutinate cells and spores. Conglutinate cells arise from hyphal adhesion and certain perithecial hyphae develop from conglutinate cells. Various hypha-conglutinate cell transitions play important roles during the development of the perithecial wall and neck.     

Experiments using sterilized apple-leaf discs to study the mode of action of urea in suppressing perithecia of Venturia inaequalis (Cke.) Wint.

Laboratory experiments using sterilized apple-leaf discs showed that treatment of leaves with urea during the early stages of perithecial initiation induced a high nitrogen content of the leaves, which prevented further development of perithecia although mycelial growth was unaffected. Treatments applied at a later stage of fungal development inhibited both perithecial and mycelial growth. Some of the bacteria isolated from urea-treated leaves in the field restricted perithecial development, particularly when applied in the first month after inoculation with suspensions of conidia from sexually compatible strains of the fungus. One isolate, a Pseudomonas sp., was shown to be important in the decomposition of apple leaves.     

Studies on the development of perithecium of Ceratocystis stenoceras

The development of the perithecium of Ceratocystis stenoceras was observed by a light microscope and by a scanning electron microscope.The fungus has developed dark brown perithecia on wheat agar medium in three days of incubation. Perithecial primordia appeared as tightly knotted coils. At the center of it an oval ascogonium was observed. The ascogonium was developed from a lateral wall of a hypha, and the hyphae covering the ascogonium branched at the basal part where the ascogonium was attached. These hyphae branched repeatedly in the developmental growth to cover the ascogonium, and it was finally covered tightly. The plasmogamy of this fungus is much probably performed by the gametangial contact. As the stage proceeded, the ascogonium elongated, the terminal and the basal portions of it swelled and cleavage of the ascogonium resulted. Each of the cleaved ascogonia germinated continuously and stretched out the ascogenous hyphae. About that time the cells consisting of perithecia were vacuolated from the center and successively dissolved, so that a space was formed in the center of the body. Ascogenous hyphae continued to develop downwards, and their end were fixed to the inner wall of the body.The upper portion of the hyphae converged to the center of the body and the ascogenous hyphae became the supporting tissue for ascus formation.Hook formation was observed prior to the ascus formation. After completion of karyogamy by hook formation, the fissure appeared on the ascus and the end portion was released. The released portion included eight ascospores. The ascus had a smooth surface and no special structure was seen on the top. As the asci were matured, they evanesced by themselves and concurrently ascospores came out. Finally the body was massively filled with ascospores.     

MORPHOLOGY OF CHAETOMIUM ERRATICUM

Development of perithecia from single, uninucleate ascospores disclosed a homothallic condition for Chaetomium erraticum. This species was found to produce sessile ascogonial coil initials from uninucleate vegetative cells that become enveloped by hyphae formed at the base of the ascogonium. The ascogonium consists of several cells that are uninucleate or binucleate. A perithecium forms from numerous divisions and enlargement of the surrounding uninucleate cells. Differentiation of the perithecial cells results in the formation of a carbonaceous wall, perithecial hairs, and an ostiole lined with periphyses. A convex hymenial cluster of ascogenous cells forms in the lower half of the centrum from which typical croziers develop. Asci push up into the pseudoparenchyma cells of the centrum. The growth of the ascogenous system is in part responsible for increase in perithecial size. The breakdown of the pseudoparenchyma cells around the developing asci results in the formation of a central cavity in which ascospores are released when the asci deliquesce. No paraphyses are present. The type of development and features of the centrum of C. erraticum and other species of Chaetomium indicate a distinct Xylaria-type centrum.     

A new Ophiostoma species isolated from the ambrosia beetle Xyleborus dryographus (Coleoptera: Scolytidae)

An undescribed ophiostomatoid fungus was isolated from the scolytid beetle Xyleborus dryographus in south-west Germany. The fungus is described as Ophiostoma verrucosum sp. nov. with a Hyalorhinocladiella anamorph. The teleomorph is characterized by thick-sheathed ascospores, numerous divergent ostiolar hyphae and an ornamented perithecial base.     

A Glomerella species phylogenetically related to Colletotrichum acutatum on Norway maple in Massachusetts

A fungus isolated from Norway maple (Acer platanoides) in the Boston, Massachusetts, area was determined to be a species of Glomerella, the teleomorph of Colletotrin chum acutatum. Pure cultures of the fungus were obtained from discharged ascospores from perithecia in leaf tissue. This fungus was determined to be homothallic based on the observation of perithecial development in cultures of single-spore isolates grown on minimal salts media and with sterile toothpicks. A morphological and molecular analysis was conducted to determine the taxonomic position of this fungus. Parsimony analyses of a combined nucleotide dataset of the ITS and LSU rDNA region, and of the D1-D2 LSU rDNA region, indicated that this species has phylogenetic affinities with Colletotrichum acutatum, C. acutatum f. sp. pineum, C. lupini, C. phormii and G. miyabeana. These results are significant because C. acutatum has not been reported on Acer platanoides. In addition the consistent presence of perithecia on leaf tissue and in culture is unusual for Colletotrichum, suggesting that the teleomorphic state is important in the life cycle of this fungus.     

Perithecial formation in Gelasinospora reticulispora III. Inhibitory effects of near-UV and blue light during the inductive dark period

Growing hyphae of Gelasinospora reticulispora required a continuousdark period prior to photoinduction of perithecia. The inductivedark period was interrupted by brief exposure of the hyphaeto white light so that the formation of perithecia no longertook place. Photosensitivity of the hyphae in terms of the light-breakeffect gradually changed during the inductive dark period. Sensitivityreached its maximum at the 18th hr of the dark period when anirradiation of 1?10⁵ ergs cm^–2 of near-UV light or 4?10⁴ergs cm^–2 of blue-light was sufficient for the light-break.Red and far-red light had no effect at all. The light-breakeffect was limited to the irradiated portion of the hyphae anddid not affect any unirradiated portions. Inhibitory effecton perithecial formation of continuous white light could betotally replaced for several days with intermittent irradiationof near-UV or blue light if given for 5 min every 4 hr. (Received December 18, 1973; )     

Theissenia rogersii sp. nov. and phylogenetic position of Theissenia

Theissenia rogersii deviates from known Theissenia species primarily in having large ascospores with a thick wall layer and a unique configuration of two stromatal tissue types, one carbonaceous and the other fibrous. The carbonaceous tissue forms palisades on and beneath the perithecial layer as well as encasing individual perithecia, whereas the fibrous tissue fills the spaces between columns of the palisades as well as between encased perithecia. Phylogenetic analyses based on DNA sequences of beta-tubulin and alpha-actin genes placed Theissenia in the subfamily Hypoxyloideae among the genera that are characterized by having bipartite stromata (i.e. with the stromata differentiated into an outer dehiscing layer and an inner perithecium-bearing layer).     

Conidioma development in Ophiodothella vaccinii

The perithecial ascomycete Ophiodothella vaccinii causes a leafspot disease of sparkleberry (Vaccinium arboreum), in which an anamorph is produced early in the life cycle of the fungus. The anamorph forms shiny, black, pulvinate conidiomata that contain a single central pore. After initial infection, fungal hyphae permeate the interior tissues of the leaf, creating lesions. Conidiomata are initiated by the formation of a small layer of intertwined, thicker-walled hyphae beneath the epidermis of the lesion. Near the center of this hyphal layer a subglobose collection of thick-walled hyphae is formed. This hyphal collection grows upward, becoming conical and pressing against the epidermis. Elongation of a columnar apex of the hyphal collection ruptures the epidermis, creating a pore. Subsequent expansion and development of conidiophores and conidia push the epidermis upward, lifting it away from the column, opening the pore and allowing conidia to emerge. The conidioma is regarded as a modified acervulus.     

Relations entre production et localisation de mycosporine et morphogenèses reproductrices chez le Pyrénomycète Gnomonia leptostyla

Relations between production and localisation of mycosporin and reproductive morphogenesis in the Pyrenomycete Gnomonia leptostyla.
The production of mycosporin (P310) has been analysed in Gnomonia leptostyla (FT.) Ces. et de Not. during mycelial growth and reproductive morphogenesis (macroconidiogenesis, microconidiogenesis and differentiation of perithecia). Conidiogenesis is induced in illuminated cultures while darkness promotes perithecial development. At 20°C, the cultures produce either macroconidia or perithecia with abortive sporophyte. Microconidia differentiation and perithecia maturation require low temperature (10°C). Mycosporin is, at all times, present in the thallus. However, the concentration of mycosporin in highest in the conidiogenous thallus, intermediate in the perithecial thallus. and lowest in the vegetative mycelium. In the conidiogenous thallus, macroconidia and microconidia are both sites of mycosporin accumulation. On the contrary, in the perithecial thallus, mycosporin levels are not higher in perithecia than in mycelia, even during their maturation period. The quantitative variations of mycosporin during the thallus development and its accumulation inside conidia suggest translocation from sites of synthesis towards reproductive cells.     

Anatomy and mycotrophy of the achlorophyllous Afrothismia winkleri

Afrothismia winkleri develops fleshy rhizomes, densely covered with small root tubercles, narrowing to filiform roots with age. The exclusively intracellular mycorrhizal fungus has distinct morphologies in different tissues of the plant. In the filiform root the hyphae grow straight and vesicles are borne on short hyphal stalks. The straight hyphae are present in the epidermis of the root tubercles, but change to loosely coiled and swollen hyphae in the rhizome tissue. No penetration from epidermis to root cortex was found. From the rhizome, a separating cell layer permits only one or rarely two hyphal penetrations into the cortex of each root tubercle. The hyphae proceed apically within the root hypodermis in a spiral row of distinctively coiled hyphae, branches of which colonize the inner root cortex. In the inner root cortex the hyphal coils degenerate to amorphous clumps. In older roots the cortex itself also deteriorates, but epidermis, hypodermis, endodermis and central cylinder persist. The mycorrhizal pattern in A. winkleri is interpreted as an elaborate exploitation system whereby the fungus provides carbon and nutrients to the plant and, simultaneously but spatially distinct, its hyphae are used to translocate and store the matter within the plant. Several features indicate that the endophyte is an arbuscular mycorrhizal fungus.     

Effect of Trichoderma harzianum on perithecial production of Gibberella zeae on wheat straw

Conidial suspensions and cell-free filtrates of Trichoderma harzianum isolates were evaluated for their effectiveness in reducing perithecial and ascospore production of Gibberella zeae on wheat straw. Isolate T-22, which is registered in the US as a biological control agent (Plant Shield™), was included in the study as a positive control. When co-inoculated with G. zeae all 11 isolates of T. harzianum significantly reduced perithecial numbers on wheat straw. Five T. harzianum isolates, including T-22, reduced perithecial formation by 70% or greater. All isolates of G. zeae, varied in their ability to produce perithecia. Isolate 192132 produced the greatest number of perithecia and was used to further evaluate the effect of application time of the T. harzianum isolates. Perithecial reduction was highest (96-99%) when T. harzianum conidial suspension or cell-free filtrate was applied to straw 24 h prior to inoculation with G. zeae. Control was less effective when T. harzianum was applied at the same time (co-inoculated) or 24 h after G. zeae. Treatments which reduced perithecial numbers also reduced ascospore numbers; however, the average numbers of ascospores per perithecia were not significantly lowered. Field trials showed significant reduction of perithecia on residues treated with T. harzianum prior to placement on the soil surface. Both T. harzianum and G. zeae were re-isolated from residues sampled in July and August after 30 and 60 days of exposure to the environment.     

Barbatosphaeria gen. et comb. nov., a new genus for Calosphaeria barbirostris

The new genus Barbatosphaeria is described for a perithecial ascomycete known as Calosphaeria barbirostris occurring on decayed wood of deciduous trees under the periderm. The fungus produces nonstromatic perithecia with hyaline, 1-septate ascospores formed in unitunicate, nonamyloid asci. Anamorphs produced in vitro belong to Sporothrix and Ramichloridium with holoblastic-denticulate conidiogenesis; conidiophores of the two types were formed in succession during the development of the colony. Phylogenetic analyses of nuLSU rDNA sequences indicate that this fungus is distinct from morphologically similar Lentomitella, tentatively placed in the Trichosphaeriales. It groups with freshwater Aquaticola and Cataractispora and terrestrial Cryptadelphia in maximum parsimony analysis; the same grouping but without Cryptadelphia was inferred from Bayesian analysis. Cultivation, morphology and phylogenetic studies of the nuLSU rDNA support the erection of a new genus for C. barbirostris.     

Field and laboratory studies of the effect of urea on ascospore production of Venturia inaequalis (Cke.) Wint

Applications of urea after harvest but before leaf-fall restricted perithecial production by Venturia inaequalis. Immersion of detached leaves in urea appeared to be the most effective method of preventing perithecial formation, although spraying attached leaves was equally effective when leaf abscission occurred within a week of treatment. A high nitrogen content within the leaf was one of the major factors contributing to suppression. Urea-treated leaves decomposed rapidly, thus destroying the overwintering substrate for the fungus. When apple plants (clone M. 111) were sprayed in autumn with 5 % urea, followed by a second (pre-bud-burst) application at 2 %, ascospore production in the spring was suppressed. The second treatment appeared to prevent the release of ascospores from mature perithecia.     

Nutrient-dependent inhibition of perithecial development due to sequential crosses on the same mycelium of Neurospora tetrasperma

Summary An explanation of perithecial inhibition in the second of two sequential crosses at different locations on the same mycelium of Neurospora tetrasperma was sought by (1) assaying media that had supported inhibited and uninhibited portions of the mycelium which contained no developing perithecia, (2) determining the effect of these media on perithecial development, (3) adding nutrients to inhibited portions of the mycelium, and (4) assaying carbon sources in media that had supported portions of the mycelium which contained developing perithecia, and portions, both inhibited and uninhibited, which contained no developing perithecia. Different kinds and volumes of media and various intervals of time between sequential crosses were used to aid in determining limits of perithecial inhibition. Perithecial inhibition was observed to be independent of volatile metabolites and pH, independent of non-volatile metabolites, reversible by addition of nutrients, dependent upon nutrient volume, and correlated with the concentration of the carbon source in the medium. It is proposed that second crosses are inhibited because of a previous lowering of the concentration of nutrients in the medium in second-cross locations, owing to prior demand upon those nutrients by the developing perithecia in first-cross locations. The possibility of an activation signal between first- and second-cross locations is discussed. No inhibitory substance in inhibited locations was detected.Supported in part by a National Science Foundation Traineeship.