Identification of macro-fungi of the Hymenomycetes with special reference to isozyme patterns and basidiospore morphology : a dissertation submitted in part fulfilment of the requirements for the Bachelor of Horticultural Science (Hons.), Lincoln College, Canterbury, New Zealand

Abstract

Several identification techniques were used with the following: Auricularia (2spp.), Peniophora (1sp.), Chondrostereum (1sp.), Agaricus (5 spp.), Amanita (2 spp.), Agrocybe (2 spp.), Panaeolus (1 sp.), Marasmius (1 sp.), Laccaria (2 spp.), Paxillus (1 sp.) and Russula (2 spp.). All species were not studied with every technique. Initial identifications were based on morphological macro-features. Light microscopy was used to study features of Agaricus (2 spp.), Russula (1 spp.), and Laccaria (1 spp.) to determine their importance in identification. Scanning electron microscopy was used to determine the size and surface structure of basidiospores of Auricularia (2 spp.), Agaricus (5spp.), Amanita (2 spp.), Agrocybe (2 spp.), Panaeolus (1 sp.) Marasmius (1 sp.), Laccaria (2 spp.), Paxillus (1 sp.), and Russula (2 spp.). Surface structure was found to be more useful in the delineation of genera and species than spore dimensions. Several methods for preparing spores for viewing in the scanning electron microscope (SEM) were evaluated . Critical point drying of hymenial fragments produced good results from fresh or rehydrated material and was found to be better than other methods. Fixation was not of value in preparing spores. Chemical tests were used on Agaricus (3 spp.), Amanita (1 sp.),Agrocybe (1 spp.), Laccaria (2 spp.), and Russula (2 spp.). No single chemical test separated all species studied, but some, such as Melzers reagent and potassium hydroxide proved to be useful in the separation of some species. Amino acid profiles were developed for Agrocybe parasitica and Agaricus vaporarius and comparison of levels with those published for other species revealed differences, but not considered important in identification. lsozyme polyacrylamide gel electrophoresis (PAGE) of the pileus, hymenium and stipe of fruiting bodies of Auricularia (1 sp.) Agaricus (5 spp.),Amanita (2 spp.),Marasmius (1 sp.), Paxillus (1 sp.), and Agrocybe (1 sp.), and mycelia of Agrocybe (2 spp.), Auricularia ( 1 sp.), Peniophora (1 sp.), Chondrostereum (1sp.), and Agaricus (1sp.) was carried out using stains for catechol oxidase, succinate dehydrogenase and alcohol dehydrogenase. Extracts of the pileus, hymenium and stipe from most species produced different banding patterns when stained for each of the 3 enzymes studied. When stained for the activity of each enzyme, different overall banding patterns were recorded for the fruiting body of each species as well as for the mycelial extracts studied. Of the three enzymes, catechol oxidase was of greatest value as extracts from all species not only reacted more quickly and positively but extractions were easier to perform. lsozyme PAGE is of value for the identification of members of the Hymenomycetes and potential exists for incorporation of isozyme banding patterns into data bases used for identification. However, further samples from different locations must be studied to determine the intraspecific variation, and thus confirm the value of isozyme PAGE in mushroom species identification. Low molecular weight (LMW) RNA PAGE enabled the seperation of 2 species of Agaricus. LMW RNA profiles could provide a means of identifying subspecies with less variation in banding pattern than might be observed using isozyme PAGE. Again further study of samples from different locations is necessary to determine the true value of LMW RNA for fungal identification.