No description available.

In order to investigate whether epigenetics plays a role in <i>P. infestans, </i>the expression patterns of genes involved in transcriptional and post-transcriptional gene silencing were investigated and several were found to be up-regulated in both pre-infection stages and <i>in planta. </i>A histone deacetylase (Pi-HDA1) was selected for functional characterization using RNA interference (RNAi). Silencing <i>Pi-HDA1 </i>produced large aberrant zoospores indicating that it may be required for development. In addition silencing of <i>Pi-HDA1 </i>resulted in reactivation of INF1 production in <i>inf1</i>-transgenic silenced strains, therefore suggesting that <i>Pi-HDA1 </i>may be directly involved in the silencing of <i>inf1 </i>in <i>inf1-</i>transgenic silenced strains. In addition an RNA helicase, <i>Pi-RNH1</i> was also found to be specifically up-regulated in zoospores. <i>Pi-RNH1</i>-silenced strains produced large aberrant zoospores that had undergone partial cleavage and often had multiple flagella on their surface. The <i>Pi-RNH1</i>-silenced zoospores were also sensitive to osmotic pressure and they often ruptured upon release from the sporangia. These findings indicate that <i>Pi-RNH1</i> has a major function in zoospore development. In addition the mechanism of internuclear gene silencing was further investigated. In <i>P. infestans </i>internuclear gene silencing is a process where silencing is transmitted from nucleus to nucleus in heterokaryotic strains. Previously it was demonstrated that internuclear gene silencing is a transcriptional silencing process. In several eukaryotes transcriptional silencing has been shown to involve methylation of cytosines. Methylation sensitive sequencing was performed and it was concluded that DNA methylation is not involved in transcriptional silencing in <i>P.</i> <i>infestans. In solution </i>hybridization assays provided preliminary evidence that small RNAs may be the diffusible factor responsible for the spread of silencing.

Chitin synthetase has been characterized from stipes of the toadstool Coprinns cinereus. This enzyme makes chitin, a major component of the walls of many fungi. It is probably associated with the plasmalemma and can be solubilized from a microsomal preparation by treatment with 10 mg/ml digitonin. This treatment results in a large increase in stability of the enzyme. The enzyme requires only magnesium ions and UDP-N-acetyl-glucosamine for activity and it is assumed that any primer required is firmly bound to the enzyme molecules. Glucose, N-acetylglucosamine and diacetylchitobiose activate chitin synthetase, while adenine and uridine nucleotides are inhibitory. The enzyme product UDP and the antibiotic polyoxin D are competitive inhibitors. Kinetic studies of the variation of activity with UDP-N-acetylglucosamine concentration suggest that this substrate is an allosteric activator of chitin synthetase, and the properties of the enzyme are related to various standard models of allosteric enzymes and to the responses of effectors present in vivo. The solubilized chitin synthetase preparation also contains proteolytic and nucleoside phosphatase activities. The properties of the uridine diphosphatase are examined for its effect on chitin synthetase through the enzyme product UDP. The enzyme is partially purified by ammonium sulphate precipitation, ion exchange chromatography and molecular exclusion chromatography. In these preparations the enzyme exists as aggregates of molecular weights of several millions which break down in the presence of salt. When eluted with 0.2 M-NaCl the enzyme anpears as a single peak with a molecular weight of about 150,000.

<I>Bacillus brevis</I> Nagano, gramicidin S producing strain and <I>B. brevis</I> E1 an antibiotic negative mutant were tested for their efficacy to control disease in two growing seasons. <I>B. brevis</I> Nagano had been extensively tested in Scotland, UK and it was found to be very effective again at <I>B. cinera</I> in Chinese cabbage and tomato crops. Also the strain E1 which produces an unidentified biosurfactant which is believed to reduce leaf wetness duration was tested for the first time <I>in planta</I>. Along with these well characterized strains three native <I>Bacillus</I> strains were tested in the greenhouse. One <I>B. subtilis</I> and two<I> B. pumilus</I> strains producing antifungal compounds were tested <I>in vitro, in vivo</I> and <I>in planta</I>.<I> B. brevis</I> E1 proved to be the most effective treatment. Its effectiveness was assigned to the performance of the biosurfactant under conditions of temperatures ranging between 16-24°C when percentage relative humidity was marginally below 90%. <I>B. brevis</I> Nagano WT which also produces biosurfactant was moderately effective in both years. The reduction of antibiotic activity due to binding of gramicidin S on ethanol soluble components of the leaf surface is considered as a possible cause for its reduced activity. Both strains managed to delay epidemics and reduce the rate of disease in an overall study of disease progress. The consistent level of disease reduction (despite the variability of the epidemics over the two years), indicated that <I>B. brevis</I> E1 is a potential BCA. Disease reduction was affected mostly by environmental factors as well as the survival of the antagonists but clear conclusions on the duration of this effect were not drawn. The number of viable colony forming units of <I>B. brevis</I> on the phylloplane decreased following an inverse exponential pattern in most of the cases. An increase was occasionally monitored which was difficult to be attributed to any abiotic or biotic factor. Biological control agents did not seem to have any detrimental effect on naturally occurring fungi and bacteria, but they were inadequate to control <I>B. cinerea</I> quiescent infection of flowers.

Porphyra umbilicalis is a monostromatic thallus consisting of a single layer of cells with thick cell walls, embedded in a mucilaginous intercellular matrix and sandwiched between the tough outer 'cuticles'. It has been reported in the past that the cell wall consists of β-1,3 xylan, the 'cuticle', a β-1,4 mannan and the intercellular matrix, a sulphated D-L-galactan. These studies had been made on materials extracted by rather drastic chemical treatments. An attempt to separate these regions in a pure and native form using biochemically mild techniques was met with success. Chemical, physical and biochemical studies show these to be complex structures whose formation and function are biologically controlled. All three extracellular regions are found to contain proteins in addition to polysaccharides which form their major constituent. The protein component contains all the common amino acids with relatively higher amounts of serine and threonine. Cysteine was found to be high in the 'cuticle' and in the cell wall. Hydroxyproline was absent from all three regions studied. Mannose and glucose were found to form the major constituents of the polysaccharide component of the 'cuticle', xylose and glucose with small amounts of uronic acid in that of the cell wall and galactose, 6-0-methyl galactose, 3,6-anhydrogalactose and sulphate half ester in that of the intercellular matrix. The presence of an enzyme causing methylation of galactose units of the intercellular matrix is reported. This enzyme appears to be playing an important role in bringing about fine structure modifications of the matrix polysaccharide, thereby preserving the structure and conformation for its biological function. Also an enzyme activity, degrading the matrix polysaccharide, is demonstrated which suggests that the production and functioning of this extracellular region is biologically controlled.

1 Methods have been devised for growing mated and unmated surface cultures of Mucor mucedo to allow analysis of cell constituents during growth and reproduction. 2 Several strain differences were revealed. The minus strain has a 20% faster linear growth rate than the plus strain. The plus strain accumulated glycogen to a greater extent than minus. Measurements of the protein, ergosterol and β-carotene contents of the minus strain always exceeded those for the plus strain. 3 When two compatible mating types meet 'head on', the levels of protein and glycogen increases as the sexual hyphae, the zygophores, are induced, but these levels drop in the later stages of zygospore formation. 4 Very large accumulations of the isoprenoids ergosterol and β-carotene result when the two strains are mated or when they are treated with externally applied trisporic acid. 5 Methods were devised to study the growth enzyme, chitin synthase, from cell-free preparations and 'in situ' preparations of Mucor mucedo during growth and differentiation. 6 Chitin synthase was most active at the hyphal apices and decreased with age and differentiation. 7 Chitin synthase from Mucor mucedo has a pH optima of pH 5.5-6.0, temperature optima of 15-20°C, requires the Mg2+ ion and N-acetylglucosamine for activation and is inhibited by Polyoxin D and colloidal chitin. 8 A membrane-bound chitinase enzyme was extracted from surface agar grown cultures of Mucor mucedo. 9 The 3,4-dinitrophenyl-tetra-N-acetyl-β-D-chitotetraoside colorimetric test enabled a detailed study of chitinase. 10 Chitinase from Mucor mucedo has a pH 5.5 optima, a temperature optima of 55°C, does not require any metal cation for activation and is inhibited by colloidal chitin.

The overall aim of this research was to provide information to assist in the conservation and management of rare stipitate hydnoid fungi that form ectomycorrhizal associations with Scots pine.  Research based on sporocarp production indicates that stipitate hydnoid fungi have declined in Europe and many species are thought to be threatened by extinction.  The development of a management strategy for these fungi relies on a better understanding of their ecology, including information on their below-ground distribution and persistence. An ITS sequence database was established for 12 species.  Species-specific primers were developed within the ITS1 and ITS2 regions and the specificity of the primer pairs was tested by conducting PCR amplifications for each primer/DNA combination as well as BLAST searches. The primary were used in conjunction with real-time PCR to compare the below-ground distribution of 2 species with their sporocarp distribution. There was no direct relationship between the below-ground mycelium and the sporocarp distribution.  A spatial analysis suggests that the mycelium of both species form aggregates. The ability of four species to persist below-ground at locations where sporocarps were previously, but are no longer, recorded was investigated.  DNA and RNA detections demonstrated that the target species were present and metabolically active. The below-ground structure of the co-existing stipitate hydnoid species was studied around two typical fruiting sites. Compared to the below-ground distribution of stipitate hydnoid fungi, the fruiting areas were small and limited to sites with a shallow or lacking organic horizon.  There were no indications for below-ground interactions between stipitate hydnoid species. Inoculum of 2 species was introduced to new native woodlands.  The persistence of both species was monitored by DNA and RNA detection and showed that it is possible to introduce stipitate hydnoid fungi.  However, the success was not predictable.

No description available.

No description available.

No description available.