Enhanced production of laccase in Trametes vesicolor by the addition of ethanol

In a medium containing 40 g ethanol l^–1, laccase production by Trametes versicolor was 2.6 unit per ml of the supernatant, which was over 20 times higher than that without ethanol. Laccase activity with ethanol was quite comparable to that with the well-known inducers such as veratryl alcohol, xylidine and guaiacol. With other white-rot fungi, Coriolus hirsutus and Grifola frondosa, ethanol had a similar stimulatory effect on laccase production.     

Fungal spore concentrations in the air at Tiruchirapalli (India) 1987–1988

Summary Using an aeroscope, airborne fungal spores were sampled for two years, 1987–1988 at Tiruchirapalli, Tamil Nadu, India. The aerospora components, their seasonal and annual variations in incidence in the air are discussed and a spore calendar for Tiruchirapalli is presented.     

Betulachrysoquinone hemiketal: A p-benzoquinone hemiketal macrocyclic compound produced by Phanerochaete chrysosporium

Betulachrysoquinone hemiketal was isolated from pre-extracted wood of Betula lutea Michx. inoculated with Phanerochaete chrysosporium Burds. Acid-catalysed hydrolysis of betulachrysoquinone hemiketal produced betulachrysoquinone which was shown to be 2-hydroxy-6-(13′-hydroxytetradecanyl)-p-benzoquinone.     

An improved method for light- and electron microscopial studies of nematode/fungal interactions

A method is presented that enables studies to be made of single nematode-fungal interactions under conditions where fungal growth at the expense of external nutrients is prevented. The nematophagous fungus Arthrobotrys ologospora was used as a model organism in these studies. The method is based on removal of the traps from the vegetative mycelium, immediately after a nematode was captured and transfer of the trap with the captured nematode into a droplet of sterile distilled water placed in a moisture chamber. In the absence of external nutrients, such isolated traps of A. oligospora were fully effective in penetrating and subsequently digesting the captured nematode. Solely vegetative mycelium was formed at the expense of the digested nematode; this developed from the trap that originally had captured the nematode. One advantage of the present method is that studies on various stages of the nematode-fungal interaction can now be performed under conditions that exclude major influences of external nutrients which otherwise could be communicated to the trap cells by way of the vegetative mycelium.     

Efficient integrative transformation of the phytopathogenic fungus Alternaria alternata mediated by the repetitive rDNA sequences

An attempt was made to transform Alternaria alternata protoplasts using a plasmid vector, pDH25, bearing the Escherichia coli hygromycin B (Hy) phosphotransferase gene (hph) under the control of the Aspergillus nidulans trpC promoter. Transformants arose on a selective medium containing 100 μg Hy/ml. There were two types of transformants, forming large and small colonies on the selective medium. Transformation with one μg of the vector produced an average of 4.5 large colonies and 600 small ones. In large-colony transformants, the vector often integrated into the recipient chromosome in the form of highly rearranged tandem arrays. To increase transformation efficiency, fragments of the highly repetitive ribosomal RNA gene cluster (rDNA) of A. alternata were used to construct four new vectors for homologous recombination system. Use of these vectors gave higher transformation efficiency than the original plasmid. The best vector, pDH25r1a, gave rise to large-colony transformants at a frequency 20 times higher than pDH25. Transformation events in A. alternata with pDH25r1a occured by homologous recombination as a single crossover between the plasmid-borne rDNA segment and its homologue in the chromosome, often giving rise to tandemly repeated vector DNA.     

Extracellular enzyme localization during interspecific fungal interactions

Abstract Confronted colonies of Phlebia radiata, P. rufa, Coriolus versicolor, Stereum hirsutum, Phanerochaete velutina and Hypholoma fasciculare showed spatially and temporally heterogeneous patterns of loccase-α-naphthol and peroxidase activities. These activities were coincident in axenic cultures. but were not always so during interaction. Confrontation between species resulted in induction of phenoloxidase activities, even within coenocytic colony regions of Phlebia species which were normally void of such activities in axenic culture. These events resulted in restriction of C. versicolor growth during interaction with P. rufa.     

High-level expression of a tobacco chitinase gene in Nicotiana sylvestris. Susceptibility of transgenic plants to Cercospora nicotianae infection

Endochitinases (E.C. 3.2.14, chitinase) are believed to be important in the biochemical defense of plants against chitin-containing fungal pathogens. We introduced a gene for class I (basic) tobacco chitinase regulated by Cauliflower Mosaic Virus 35S-RNA expression signals into Nicotiana sylvestris. The gene was expressed to give mature, enzymatically active chitinase targeted to the intracellular compartment of leaves. Most transformants accumulated extremely high levels of chitinase-up to 120-fold that of non-transformed plants in comparable tissues. Unexpectedly, some transformants exhibited chitinase levels lower than in non-transformed plants suggesting that the transgene inhibited expression of the homologous host gene. Progeny tests indicate this effect is not permanent. High levels of chitinase in transformants did not substantially increase resistance to the chitin-containing fungus Cercospora nicotiana, which causes Frog Eye disease. Therefore class I chitinase does not appear to be the limiting factor in the defense reaction to this pathogen.     

Purification and infectivity ofEntomophaga grylli (Fresenius) Batko Pathotype 2 againstMelanoplus differentialis (Thomas) [Ort.: Locustidae]

The life stages ofEntomophaga grylli (Fresenius) Batko Pathotype 2 were purified and separated by centrifugation in Percoll^R density-gradient medium. The ranges of buoyant densities for germinated resting spores, germ conidia, and resting spores respectively were: 1.040–1.050, 1.055–1.085, and 1.080–1.120 g/ml. Cuticular invasion by germinated germ conidia was the means by whichMelanoplus grasshoppers became infected. Scanning electron micrographs revealed germination of germ conidia on the visible host integument at 100% RH, but not at 90% RH. Significantly higher mortality (P<0.05) was obtained after 3 weeks with grasshoppers incubated in constant light than in constant dark for 24 h following treatment. The disease was not transmitted by ingestion of any life stage. Contribution N^o 85-153-J, Department of Entomology. Agricultural Experiment Station, Kansas State University, Manhattan, KS 66506.     

Studies on the biosynthesis and metabolism of the phytoalexin lubimin and related compounds in Datura stramonium L.

Arachidonic acid, cellulase, CuSO₄, a sonicate of Phytophthora infestans mycelium and a spore suspension of Penicillium chrysogenum all elicited the formation of the sesquiterpenoid phytoalexins lubimin, 3-hydroxylubimin and rishitin in fruit cavities of Datura stramonium. 3-Hydroxylubimin was the predominant phytoalexin formed after treatment of the fruits with arachidonic acid, cellulase and the P. infestans preparation. Copper sulphate was a potent elicitor of lubimin but not 3-hydroxylubimin. The fungus P. chrysogenum metabolized lubimin and 3-hydroxylubimin to 15-dihydrolubimin and 3-hydroxy-15-dihydrolubimin respectively, both in fruit cavities inoculated with spores of this fungus and in pure culture. The 15-dihydrolubimin formed in the fruits by the fungus was further metabolized (by the fruits) to both isolubimin and 3-hydroxy-15-dihydrolubimin. The precursor-product relationships between all of the subject compounds was investigated by feeding experiments with ³H-labelled compounds. 2-Dehydro-[15-³H₁]lubimin was rapidly and efficiently incorporated into lubimin and may be the direct precursor of lubimin in planta. 3-Hydroxy[2-³H₁]lubimin was incorporated into the nor-eudesmane rishitin but 10-epi-3-hydroxy[2-³H₁]lubimin was not. An updated scheme for the biosynthesis and metabolism of lubimin and related compounds in infected tissues of solanaceous plants is presented.We thank Mr Vic Swetez for the provision of plant material, Mrs Margaret Huffee for technical assistance, Dr David Ewing for help with obtaining NMR spectra, and the Agricultural and Food Research Council for financial support.