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Background: Ergot, caused by the fungal pathogen Claviceps purpurea, infects the female flowers of a range of cereal crops, including wheat. To understand the interaction between C. purpurea and hexaploid wheat we undertook an extensive examination of the reprogramming of the wheat transcriptome in response to C. purpurea infection through floral tissues (i.e. the stigma, transmitting and base ovule tissues of the ovary) and over time., Results: C. purpurea hyphae were observed to have grown into and down the stigma at 24 h (H) after inoculation. By 48H hyphae had grown through the transmitting tissue into the base, while by 72H hyphae had surrounded the ovule. By 5 days (D) the ovule had been replaced by fungal tissue. Differential gene expression was first observed at 1H in the stigma tissue. Many of the wheat genes differentially transcribed in response to C. purpurea infection were associated with plant hormones and included the ethylene (ET), auxin, cytokinin, gibberellic acid (GA), salicylic acid and jasmonic acid (JA) biosynthetic and signaling pathways. Hormone-associated genes were first detected in the stigma and base tissues at 24H, but not in the transmitting tissue. Genes associated with GA and JA pathways were seen in the stigma at 24H, while JA and ET-associated genes were identified in the base at 24H. In addition, several defence-related genes were differential expressed in response to C. purpurea infection, including antifungal proteins, endocytosis/exocytosis-related proteins, NBS-LRR class proteins, genes involved in programmed cell death, receptor protein kinases and transcription factors. Of particular interest was the identification of differential expression of wheat genes in the base tissue well before the appearance of fungal hyphae, suggesting that a mobile signal, either pathogen or plant-derived, is delivered to the base prior to colonisation., Conclusions: Multiple host hormone biosynthesis and signalling pathways were significantly perturbed from an early stage in the wheat - C. purpurea interaction. Differential gene expression at the base of the ovary, ahead of arrival of the pathogen, indicated the potential presence of a long-distance signal modifying host gene expression.

Pathogen-derived cytokinins (CKs) have been recognized as important virulence factor in several host-pathogen interactions and it was demonstrated multiple times that phytopathogenic fungi form CKs via the tRNA degradation pathway. In contrast to previous studies, the focus of this study is on the second step of CK formation and CK degradation to improve our understanding of the biosynthesis in fungi on the one hand, and to understand CK contribution to the infection process of Claviceps purpurea on the other hand. The ergot fungus Claviceps purpurea is a biotrophic phytopathogen with a broad host range including economically important crops causing harvest intoxication upon infection. Its infection process is restricted to unfertilized ovaries without causing macroscopic defense symptoms. Thus, sophisticated host manipulation strategies are implicated. The cytokinin (CK) plant hormones are known to regulate diverse plant cell processes, and several plant pathogens alter CK levels during infection. C. purpurea synthesizes CKs via two mechanisms, and fungus-derived CKs influence the host-pathogen interaction but not fungus itself. CK deficiency in fungi with impact on virulence has only been achieved to date by deletion of a tRNA-ipt gene that is also involved in a process of translation regulation. To obtain a better understanding of CK biosynthesis and CKs' contribution to the plant-fungus interaction, we applied multiple approaches to generate strains with altered or depleted CK content. The first approach is based on deletion of the two CK phosphoribohydrolase (LOG)-encoding genes, which are believed to be essential for the release of active CKs. Single and double deletion strains were able to produce all types of CKs. Apparently, log gene products are dispensable for the formation of CKs and so alternative activation pathways must be present. The CK biosynthesis pathway remains unaffected in the second approach, because it is based on heterologous overexpression of CK-degrading enzymes from maize (ZmCKX1). Zmckx1 overexpressing C. purpurea strains shows strong CKX activity and drastically reduced CK levels. The strains are impaired in virulence, which reinforces the assumption that fungal-derived CKs are crucial for full virulence. Taken together, this study comprises the first analysis of a log depletion mutant that proved the presence of alternative cytokinin activation pathways in fungi and showed that heterologous CKX expression is a suitable approach for CK level reduction.

Claviceps purpurea is an ovarian parasite infecting grasses (Poaceae) including cereals and forage plants. This fungus produces toxic alkaloids and consumption of contaminated grains can cause ergotism in humans and other mammals. Recent molecular genetics studies have indicated that it included three cryptic species (G1, G2, G3). In this study, reproductive isolation amongst these groups and among material from Phragmites and Molinia was tested using gene flow statistics for five polymorphic loci, and to support these data, phylogenetic affiliations based on gene trees and a multigene phylogeny were used. The four recognized species are characterized based on morphology and host spectrum and formal taxonomic names are proposed. Claviceps purpurea sensu stricto (G1 group) represents a typical rye ergot, but infects various other grasses. Typical hosts of Claviceps humidiphila (new name for G2 species), like Phalaris arundinacea, belong to grasses preferring humid locations. Claviceps spartinae (G3) is specific to chloridoid grasses from salt barches. The material from Phragmites and Molinia can be authenticated with the species Claviceps microcephala for which the new name Claviceps arundinis is proposed here. The divergence time between species was estimated and the tools for species identification are discussed., (Copyright © 2014 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.)

It is well recognized, that outcomes of mutualistic plant-microorganism interactions are often context dependent and can range from mutualistic to antagonistic depending on conditions. Instead, seemingly pathogenic associations are generally considered only harmful to plants. The ergot fungus (Claviceps purpurea) is a common seed pathogen of grasses and cereals. Ergot sclerotia contain alkaloids which can cause severe toxicity in mammals when ingested, and thus the fungal infection might provide protection for the host plant against mammalian herbivores. Theoretically, the net effect of ergot infection would positively affect host seed set if the cost is not too high and the defensive effect is strong enough. According to our empirical data, this situation is plausible. First, we found no statistically significant seed loss in wild red fescue (Festuca rubra) inflorescences due to ergot infection, but the seed succession decreased along increasing number of sclerotia. Second, in a food choice experiment, sheep showed avoidance against forage containing ergot. Third, the frequency of ergot-infected inflorescences was higher in sheep pastures than surrounding ungrazed areas, indicating a protective effect against mammalian grazing. We conclude that, although ergot can primarily be categorized as a plant pathogen, ergot infection may sometimes represent indirect beneficial effects for the host plant. Ergot may thus serve as a conditional defensive mutualist for its host grass, and the pathogenic interaction may range from antagonistic to mutualistic depending on the situation.

Sorghum ergot (caused by Claviceps africana) is a disease that affects sorghum seed development and yield. The interaction between pollen tube growth and hyphal development determines whether ovaries will be fertilized or colonized. Thus their respective deposition times on the stigma are critical. The effect of the time interval between pollination and inoculation on stigma receptivity and seed production was measured under field conditions in the male-sterile line A9 at Montecillo, State of México (2240m altitude). Pollination and inoculation treatments, from simultaneous application to 2 and 4h difference, were imposed when all stigmas on the panicle had emerged. Control panicles were either only pollinated or only inoculated. Eighteen hours later, pollen grains that adhered to, and germinated within the stigma, pollen tubes in the style and ovary, and fertilized pistils were counted. Pistils showing some disease expression (germinated spores, mycelium growth, or tissue necrosis) at 18, 48, and 72h were recorded. The number of diseased florets was registered at the dough growth stage, while number of seeds, grain yield and 100-seeds weight was measured at the physiological maturity. The pathogen applied in a water suspension of macro and secondary conidia caused a decrease in stigma receptivity; the greatest decrease (40-60%) occurred when the pollen and the inoculum were deposited almost simultaneously, regardless of which was deposited first. The route of the pollen tube was also the route for fungal infection. On average, treatments first inoculated had 60% more diseased florets and 36% less grain yield, 30% fewer seeds and seed size decreased 8%, than those first pollinated., (Copyright © 2010 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.)

The ergot diseases of grasses, caused by members of the genus Claviceps, have had a severe impact on human history and agriculture, causing devastating epidemics. However, ergot alkaloids, the toxic components of Claviceps sclerotia, have been used intensively (and misused) as pharmaceutical drugs, and efficient biotechnological processes have been developed for their in vitro production. Molecular genetics has provided detailed insight into the genetic basis of ergot alkaloid biosynthesis and opened up perspectives for the design of new alkaloids and the improvement of production strains; it has also revealed the refined infection strategy of this biotrophic pathogen, opening up the way for better control. Nevertheless, Claviceps remains an important pathogen worldwide, and a source for potential new drugs for central nervous system diseases.

Sorghum ergot, caused predominantly by Claviceps africana Frederickson, Mantle, de Milliano, is a significant threat to the sorghum industry worldwide. The objectives of this study were firstly, to identify molecular markers linked to ergot resistance and to two pollen traits, pollen quantity (PQ) and pollen viability (PV), and secondly, to assess the relationship between the two pollen traits and ergot resistance in sorghum. A genetic linkage map of sorghum RIL population R931945-2-2 x IS 8525 (resistance source) was constructed using 303 markers including 36 SSR, 117 AFLP , 148 DArT and two morphological trait loci. Composite interval mapping identified nine, five, and four QTL linked to molecular markers for percentage ergot infection (PCERGOT), PQ and PV, respectively, at a LOD >2.0. Co-location/linkage of QTL were identified on four chromosomes while other QTL for the three traits mapped independently, indicating that both pollen and non pollen-based mechanisms of ergot resistance were operating in this sorghum population. Of the nine QTL identified for PCERGOT, five were identified using the overall data set while four were specific to the group data sets defined by temperature and humidity. QTL identified on SBI-02 and SBI-06 were further validated in additional populations. This is the first report of QTL associated with ergot resistance in sorghum. The markers reported herein could be used for marker-assisted selection for this important disease of sorghum.

Ergoline alkaloids occur in taxonomically unrelated taxa, such as fungi, belonging to the phylum Ascomycetes and higher plants of the family Convolvulaceae. The disjointed occurrence can be explained by the observation that plant-associated epibiotic clavicipitalean fungi capable of synthesizing ergoline alkaloids colonize the adaxial leaf surface of certain Convolvulaceae plant species. The fungi are seed transmitted. Their capacity to synthesize ergoline alkaloids depends on the presence of an intact differentiated host plant (e.g. Ipomoea asarifolia or Turbina corymbosa [Convolvulaceae]). Here, we present independent proof that these fungi are equipped with genetic material responsible for ergoline alkaloid biosynthesis. The gene (dmaW) for the determinant step in ergoline alkaloid biosynthesis was shown to be part of a cluster involved in ergoline alkaloid formation. The dmaW gene was overexpressed in Saccharomyces cerevisiae, the encoded DmaW protein purified to homogeneity, and characterized. Neither the gene nor the biosynthetic capacity, however, was detectable in the intact I. asarifolia or the taxonomically related T. corymbosa host plants. Both plants, however, contained the ergoline alkaloids almost exclusively, whereas alkaloids are not detectable in the associated epibiotic fungi. This indicates that a transport system may exist translocating the alkaloids from the epibiotic fungus into the plant. The association between the fungus and the plant very likely is a symbiotum in which ergoline alkaloids play an essential role.

Phenoloxidase (PO) is an important component of the insect immune system and is frequently used to measure an individual's immune defence ability. However, evidence documenting positive correlations between the immune assay and resistance against pathogens is scarce and contradictory. We used replicate lines of yellow dung flies Scathophaga stercoraria (L.) with different PO levels to investigate whether PO levels affect resistance against parasitic mites and entomopathogenic fungi. Prevalence of flies exposed to pathogens was the same in all selection regimes, although pathogens clearly negatively affected fitness. PO measurements alone therefore do not necessarily predict overall resistance against pathogens. Furthermore, under starvation lines selected for high PO levels did not survive longer than those selected for low PO levels, irrespective of exposure to pathogens. This suggests that even if elevated immune levels increase an individual's ability to combat pathogens, the benefits may not outweigh the costs of increased investment in immunity.

Grass-associated fungi (grass symbionts) in the family Clavicipitaceae (Ascomycota, Hypocreales) are species whose host range is restricted to the plant family Poaceae and rarely Cyperaceae. The best-characterized species include Claviceps purpurea (ergot of rye) and Neotyphodium coenophialum (endophyte of tall fescue). They have been the focus of considerable research due to their importance in agricultural and grassland ecosystems and the diversity of their bioactive secondary metabolites. Here we show through multigene phylogenetic analyses and ancestral character state reconstruction that the grass symbionts in Clavicipitaceae are a derived group that originated from an animal pathogen through a dynamic process of interkingdom host jumping. The closest relatives of the grass symbionts include the genera Hypocrella, a pathogen of scale insects and white flies, and Metarhizium, a generalist arthropod pathogen. These data do not support the monophyly of Clavicipitaceae, but place it as part of a larger clade that includes Hypocreaceae, a family that contains mainly parasites of other fungi. A minimum of 5-8 independent and unidirectional interkingdom host jumps has occurred among clavicipitaceous fungi, including 3-5 to fungi, 1-2 to animals, and 1 to plants. These findings provide a new evolutionary context for studying the biology of the grass symbionts, their role in plant ecology, and the evolution of host affiliation in fungal symbioses.

Epichloë species are fungal symbionts (endophytes) of grasses, six European and four North American biological species in genus Epichloë have been described in previous researches. In this study we describe a new Epichloë species, Epichloë yangzii Li et Wang, found in natural symbioses with Roegneria kamoji native to China. We investigated the host specificity, morphology, interfertility tests and molecular phylogenetic evidences of this new species. The results indicated that E. yangzii is host specific and seedborne. Most morphological characteristics of this new species are typical in the genus. However differences are evident in several features including size of perithecia, asci and ascospores. In mating tests E. yangzii was not interfertile with E. elymi isolates from related hosts in genera Elymus. Phylogenetic relationships based on sequences of beta-tubulin gene (tub2) introns and translation elongation factor 1-alpha gene (tef1) introns showed that members of the new species grouped into exclusive clades with high bootstrap value.

Sorghum ergot in India is caused by Claviceps africana and C. sorghi. The distributions of these two species in India is not known. Eighty-nine sorghum ergot isolates were cultured from young sphacelia obtained from male sterile sorghum plants artificially inoculated using inoculum collected in the field. Based on cultural characteristics, the isolates were separated into two groups which differed distinctly in the morphology of their sphacelia, conidia, and sclerotia. Marked differences also were observed in rates of secondary conidial production and disease spread between the groups. In combination with molecular evidence, our results confirm that the isolates placed in Group I represent C. africana and Group II isolates represent C. sorghi. C. africana was found to be widely distributed in all sorghum growing areas of India. The species first described as occuring in India, C. sorghi, appears to be restricted to a few locations in the states of Maharashtra, Andhra Pradesh, and Karnataka.

Claviceps purpurea, a biotrophic pathogen of cereals, has developed a unique pathogenic strategy including an extended period of unbranched directed growth in the host's style and ovarian tissue to tap the vascular system. Since the small GTPase Cdc42 has been shown to be involved in cytoskeleton organization and polarity in other fungi, we investigated the role of Cdc42 in the development and pathogenicity of C. purpurea. Expression of heterologous dominant-active (DA) and dominant-negative (DN) alleles of Colletotrichum trifolii in a wild strain of C. purpurea had significant impact on vegetative differentiation: whereas DA Ctcdc42 resulted in loss of conidiation and in aberrant cell shape, expression of DN Ctcdc42 stimulated branching and conidiation. Deletion of the endogenous Cpcdc42 gene was not lethal but led to a phenotype comparable to that of DN Ctcdc42 transformants. DeltaCpcdc42 mutants were nonpathogenic; i.e., they induced no disease symptoms. Cytological analysis (light microscopy and electron microscopy) revealed that the mutants can penetrate and invade the stylar tissue. However, invasive growth was arrested in an early stage, presumably induced by plant defense reactions (necrosis or increased production of reactive oxygen species), which were never observed in wild-type infection. The data show a significant impact of Cpcdc42 on vegetative differentiation and pathogenicity in C. purpurea.

A study was conducted in College Station, TX, to determine the viability of Claviceps africana spores in the digestive tract of adult corn earworm moths, Helicoverpa zea (Boddie). Both sexes were exposed to ergot-infected sorghum panicles for 30 min, and spores were recovered from excreta of the moths at 24-, 48-, and 72-h intervals after feeding. Recovered spores were quantified, and viability was determined by the germination rate of macroconidia. Nearly a 100-fold greater concentration of spores was recovered from female excreta at the three time intervals compared with male excreta. Concentration of spores in female and male excreta was greatest at 24 h, with a significant reduction at the later time intervals. Spore germination rates for both sexes were greater at 24 h, with survival being significantly reduced at the 72-h interval. Spores in female excreta survived longer than those from male excreta. Spore survival over time was significantly reduced in male excreta. Spore concentration and survival were greater from female excreta, which is key, because egg-laying activities on sorghum panicles intensify during flowering, and this source of ergot spores could contribute to the spread of the disease. This study demonstrates that corn earworm moths can internally carry viable ergot spores for several days and can act as primary dispersal agents for the fungus. This is important because contaminated moths migrating from areas in Mexico and southern Texas where ergot is endemic could transmit and spread the disease to other sorghum growing regions of the United States.

Types of asexual sporulation in 17 Claviceps species and the closely related Corallocytostroma ornicopreoides were revised in relation to the phylogeny of clavicipitaceous fungi. We observed: (1) enteroblastic conidiation from branched phialidic conidiophores typical of the genus (anamorph Sphacelia) in all species including Corallocytostroma; (2) widespread and often sequential formation of terminal holoblastic secondary conidia on tapering hyphae arising from sphacelial macroconidia: and (3) in addition to sphacelial conidiation, sympodial holoblastic conidiation of the Ephelis-type in cultures of C. zizaniae and in both the culture and sphacelial tissue of C. citrina. Secondary conidiation was not found in C. purpurea, C. citrina and C. sorghicola. During sphacelial fructification, most species produced macroconidia and microconidia. Only macroconidia formed in planta underwent secondary conidiation whereas microconidia did not germinate at all. In C. phalaridis, the formation of holoblastic 2-3 celled appendaged conidia was observed, similar to that of Aciculosporium and Neoclaviceps. In dendrograms based on ITS1-5.8S rDNA-ITS2 sequences, genera and species with appendaged conidia grouped on a highly supported clade with ancestral Corallocytostroma. The clade was placed inside a group of tropical species of Claviceps, without any relationship to Balansiae.

Sixteen isolates of Claviceps spp. were analyzed for the production of polysaccharides, oligosaccharides, and sucrose metabolism under conditions of submerged fermentation. Physiological markers calculated by the Verhulst-Pearl law were used for hierarchical cluster analysis. Low correlation was found between physiologically based dendrogram and phylogenetic analysis constructed from an alignment of rDNA sequences. To confirm the intraspecific uniformity of physiological markers three isolates of C. africana from different hosts and locations were included. The influence of genotype, physiological variability, environmental location and habitat on metabolite production is discussed.

The infectivity of 4 isolates of Metarhizium anisopliae to puparia of Ceratitis capitata treated as late third-instar larvae in unsterilized soil was investigated in the laboratory under controlled temperature and moisture. At 20-30 degrees C, mortality in puparia was highest at water potential of -0.1 and -0.01 mega Pascal (MPa) and lowest at water potential of -0.0055 and -0.0035 MPa in all the isolates. In wetter soil however, isolates ICIPE 20 and 60 caused significantly higher mortality than ICIPE 18 and 69. The survival of conidia in drier soil (-0.1 MPa) was not adversely affected at all temperatures. However, in wet soil (-0.0035 MPa) there was drastic reduction in colony counts in ICIPE 18 and 69 at 25 and 30 degrees C but conidial density in ICIPE 20 and 60 remained at the initial level at 14 days after inoculation at all temperatures. When ICIPE 20 was evaluated against three other fruit fly species (Ceratitis cosyra, Ceratitis rosa, and Ceratitis fasciventris), significant reduction in adult emergence and higher pupal mortality occurred in C. cosyra and C. fasciventris than in C. rosa at a combination of 15 and 20 degrees C and -0.1 and -0.0035 MPa. However, at higher temperature and the same moisture level, the isolates were equally pathogenic across the 3 species. It is probable that in addition to pathogen cycling and multiplication from dead infected insects in the soil, a balance between microbial degradation and replenishment of inoculum of virulent isolates occur through fluctuations in, and intricate interactions between temperature and moisture levels. This study is indicative of the potential of using isolate ICIPE 20 for soil inoculation against pupariating third-instar larva of fruit flies, thus providing a novel alternative to chemical soil application.

We introduce our paper by briefly outlining how we came to the conclusion that neurons in the brain and cells in other tissues can be killed by high intracellular concentrations of calcium ions (Ca(2+)). The life cycle of Claviceps purpurea is then described before theoretical information is drawn upon which indicates that ergot alkaloids can inactivate alpha adrenergic receptors for noradrenaline released from sympathetic nerve terminals. We deduce that inhibition of these receptors by ergot alkaloids leads to an increase in intracellular Ca(2+) concentration which can cause the death of the insulin producing cells of the pancreas., (Copyright 2003 Elsevier Science Ltd.)

The effects of Cordyceps sinensis (CS) and its extracted fractions on steroidogenesis in MA-10 cells were determined. Different concentrations of CS and 3 fractions of CS (F1, a water-soluble polysaccharide; F2, a water-soluble protein; and F3, a poorly water-soluble polysaccharide and protein) were added to MA-10 mouse Leydig tumor cells with or without human chorionic gonadotropin (hCG), and the production of steroid and the expression of steroidogenic acute regulatory protein (StAR) were examined. The results showed that CS alone (2-10 mg/mL) stimulated MA-10 cell progesterone production in a dose-dependent relationship. Fractions F1 and F3 (2-10 mg/mL) also had significant (P < .05) stimulatory effects on MA-10 cell steroidogenesis with a dose-dependent relationship. However, fraction F2 did not have an effect on MA-10 cells. CS and F3, but not F1, significantly induced more steroid production in hCG-stimulated MA-10 cells (P < .05). As a temporal relationship, F1 and F3 (2 mg/mL) maximally stimulated progesterone production between 1 and 3 hours after stimulation in MA-10 cells. In addition, CS and F3 significantly enhanced MA-10 cell StAR protein expression, which indicates that CS and F3 may use a cyclic adenosine monophosphate signal transduction pathway to activate MA-10 Leydig cell steroidogenesis in a manner to that of luteinizing hormone.

Reproductive and preweaning data on 233 Angus (A), Brahman (B), and reciprocal-cross cows (AB, BA) and 455 two- and three-breed cross calves managed on common bermudagrass or endophyte-infected tall fescue were used to evaluate the interaction of forage type with individual and maternal heterosis and maternal and grandmaternal breed effects. Cows were born from 1988 to 1991 and calves from 15 Polled Hereford sires were born from 1991 to 1994. Heterosis for calving rate was similar and important on both forages (P < .01), but maternal effects were small on each forage. Maternal heterosis for birth weight differed between common bermudagrass and tall fescue (P < .10) and grandmaternal effects were evident on bermudagrass (P < .05) but not tall fescue. Forage effects were generally substantial for 205-d weight, weaning hip height, and weaning weight:height ratio (P < .01), and maternal heterosis for these traits was larger on tall fescue than on common bermudagrass (P < .01). Grandmaternal effects were in favor of Angus for 205-d weight, hip height, and weight:height ratio on common bermudagrass (P < .05) but not on tall fescue. Heterosis for 205-d weight per cow exposed was substantial on both forages (P < .01) and was numerically larger on tall fescue than on bermudagrass, but maternal effects were not significant. These results suggest more advantage for Brahman-cross cows over purebreds on endophyte-infected tall fescue than a similar comparison on common bermudagrass. They also suggest an advantage for Angus in grandmaternal effects on bermudagrass but not tall fescue.

A saprotrophic strain of Claviceps purpurea VNIIA 312A, an organism producing peptide +ergot alkaloids with prolactin inhibiting activity was shown to die under lyophilization conditions. To provide long-term storage of strain 312A, L-drying or drying under vacuum from liquid state was used with success. Three protective media were tested. Favourable results were obtained by using 25 per cent maltose solution as a protective medium. Preservation of the culture viability was accompanied by maintenance of the culture capacity for active formation of the biomass and production of +ergot alkaloids.

A homokaryotic, clavine alkaloid-producing strain of ergot, Claviceps sp. strain SD 58, was used in an attempt to demonstrate parasexuality. Genetically marked auxotrophic strains were produced by mutation with N-methyl-N'-nitro-N-nitrosoguanidine. Protoplast fusion of pairs of unlike doubly auxotrophic strains and isolation of stable prototrophic fusion products were carried out. By growth of the fusion products on complete medium, selective pressure for prototrophy was removed and auxotrophic segregants were allowed to form. Analysis of these and recovery of segregants with nonleaky, non-parent-type combinations of auxotrophic characteristics has provided strong evidence that a parasexual cycle can function in Claviceps sp. strain SD 58. Preliminary work suggests that the genetics of ergot might be studied by mitotic analysis and that protoplast fusion and selection procedures might be useful for the enhancement of favorable characteristics in Claviceps strains.

The mutant strain Claviceps purpurea 244 forming hyphae composed mainly from sclerotium-like cells was found to sporulate both in liquid and solid media, particularly in the form of terminal chlamydospores (4.0 x 6.5 micrometer). Chlamydospores produced during submerged cultivation germinated, new chlamydospores being formed directly from germ tubes, or, occasionally, conidia (the so-called microcycle) or new vegetative mycelium were formed. The ultrastructures of the chlamydospores and vegetative cells was identical. The cytoplasm was filled with ribosomes and contained lipid inclusions and vacuoles with membrane invaginations. Strain 244 cultivated under submerged conditions produced 150 microgram/ml clavins, with elymoclavin predominating (82%). The parent strain Claviceps pururea 129 only produced chlamydospores on the vegetative mycelium, whereas no microcycle was detected; under submerged conditions it produced mainly agroclavin (85%) at a concentration of 4 mg/ml.

A non-mutant saprophytic strain C. paspali which forms conidia both on a solid medium and during submerged fermentation is described. Conidiation proceeded in parallel with culture growth and production of alkaloids. The effect of composition of culture media on the intensity of conidiation is described.

Several strains of Claviceps purpurea, a phytopathogenic Ascomycete, contain mitochondrial (mt) plasmids in high molar excess relative to mtDNA. Comparative analysis of plasmids of four strains of different geographic origin revealed that all the plasmids are structurally related (size; linearity; restriction map; probably 5'-linked terminal protein; terminal inverted repeats, TIRs); two of them are even identical, indicating a possible mobility of these genetic entities. In strain K it was shown that plasmid titres are comparably high in axenic cultures and in parasitic structures (sclerotia). Detailed analysis of plasmid pClK1 proved the existence of a perfect TIR of 327 bp; the plasmid's structure and details of its nucleotide sequence indicate a replication modus comparable to that of adenoviruses. pClK1 is almost completely transcribed resulting in two major transcripts of 3.5 and 3.15 kb, respectively. In plasmid-free strains (cured by ethidium bromide treatment) these mRNAs are not detectable; nevertheless they show no significant difference in phenotype. As judged from their structural properties they could be derived from viral ancestors. In this context the plasmids' close relationship to mt plasmids of higher plants may be of special interest.