To become familiar with mycological culture techniques.
To visualize and identify the structural components of fungi.
The fungi are a group of eukaryotic protists that lack chlorophyll. We have seen the velvety blue and green growth on rotting oranges and lemons as well as on stale cheeses, the whitish –gray furry outgrowth on bread and jam and the mushrooms in the fields. These are the bodies of various fungi. Thus fungi have a diversity of morphological appearances, depending on the species. Fungi comprise the molds and yeasts. Molds are filamentous and multi cellular whereas yeasts are usually unicellular.
Figure 1: (a) Penicillium growing on apple Figure 1: (b) Rhizopus growing on bread
The fungi possess rigid cell walls containing chitin, mannan and other polysaccharides. The cytoplasmic membrane contains sterols. They possess true nuclei with nuclear membrane and paired chromosomes. They divide asexually, sexually or by both processes. They reproduce by means of spores. Fungi that reproduce by asexual spores and sexual pores are called perfect fungi and the fungi that reproduce
only by asexual spores are called imperfect fungi (deuteromycetes)
Most fungi are saprophytes, securing their nutrients from dead organic materials. Like many bacteria, fungi release hydrolytic exoenzymes that digest external substrates. They then absorb the soluble products. They are Most fungi are saprophytes, securing their nutrients from dead organic materials. Like many bacteria, fungi release hydrolytic exoenzymes that digest external substrates. They then absorb the soluble products. They are chemoorganoheterotrophs and use organic compounds as a source of carbon, electrons, and energy.
Glycogen is the primary storage polysaccharide in fungi. Most fungi use carbohydrates (preferably glucose or maltose) and nitrogenous compounds to synthesize their own amino acids and proteins. Fungi usually are aerobic. Some yeast, however, are facultatively anaerobic and can obtain energy by fermentation, such as in the production of ethyl alcohol from glucose. Obligately anaerobic fungi are found in the rumen of cattle.
Fungi grow best in dark, moist habitats, but they are found wherever organic material is available. They are primarily terrestrial organisms, although a few are freshwater or marine. Many are pathogenic and infect plants and animals. Fungi also form beneficial relationships with other organisms. For example, the association between the roots of vascular plants and fungi called mycorrhizae. Fungi also are found in the upper portions of many plants. These endophytic fungi affect plant reproduction and palatability to herbivores. Lichens are associations of fungi and either algae or Cyanobacteria.
Figure 2: (a) Mycorrhizal root tip (b) Foliose lichen
Fungi are important to humans in both beneficial and harmful ways. With bacteria and a few other groups of heterotrophic organisms, fungi act as decomposers, which is of great significance. They degrade complex organic materials in the environment to simple organic compounds and inorganic molecules. In this way carbon, nitrogen, phosphorus, and other critical constituents of dead organisms are released and made available for living organisms.
As parasites, fungi cause diseases in plants, humans and other animals. Although fungal diseases are less commonly encountered than bacterial or virus disease in humans and other animals, they are of great importance in causing diseases of plants.
Figure 3: Grey mold disease on tomato
Fungi, especially the yeasts, are essential to many industrial processes involving fermentation. Examples include the making of bread, wine, and beer. Fungi also play a major role in the preparation of some cheeses and soy sauce, in the commercial production of many organic acids (citric, gallic) and certain drugs (ergometrine, cortisone); and in the manufacture of many antibiotics (penicillin, griseofulvin) and the immunosuppressive drug cyclosporine.
In addition, fungi are important research tools in the study of fundamental biological processes. Cytologists, geneticists, biochemists, biophysicists, and microbiologists regularly use fungi in their research. Based on this research the yeast Saccharomyces cerevisiae is the best understood eucaryotic cell. The branch of science dealing with the study of fungi is known as Mycology.
The body or vegetative structure of a fungus is called a thallus. It varies in complexity and size, ranging from the single-cell microscopic yeasts to multicellular molds.
A yeast is a unicellular fungus that has a single nucleus and reproduces either asexually by budding and transverse division or sexually through spore formation. Each bud that separates can grow into a new yeast, and some group together to form colonies. Generally yeast cells are larger than bacteria, vary considerably in size, ranging from 1 to 5µm in width and from 5 to 30µm or more in length. They are commonly spherical to egg shaped. They have no flagella but do possess most of the other eucaryotic organelles.
A mold or filamentous fungi consists of long, branched, threadlike filaments of cells called hyphae that form a mycelium, a tangled mass or tissue like aggregation. Vegetative hyphae grow on the surface of culture media. They form aerial hyphae, called reproductive hyphae, that bear asexual reproductive spores or conidia. The hyphae that grow below the surface of culture media are called rhizoidal hyphae. The hyphal strand of some molds may be separated by a cross-wall called a septum. Hyphae that contain septa are called septate hyphae. Molds with hyphae that lack septa are called coenocytic hyphae.
Figure 5: (a) Aspergillus niger with conidiophores (b) Conidiophores of Penicillium sp.
Many fungi, especially those that cause diseases in humans and animals, are dimorphic -that is, they have two forms. Dimorphic fungi can change from the yeast (Y) form in the animal to the mold or mycelial form (M) in the external environment in response to changes in various environmental factors (nutrients, CO2 tension, oxidation-reduction potentials, temperature). This shift is called the YM shift. In plant-associated fungi the opposite type of dimorphism exists: the mycelial form occurs in the plant and the yeast form in the external environment
The isolation, culture, and microscopic examination of molds require the use of suitable selective media and special microscopic slide techniques. The process of transferring hyphae to a slide breaks up the structure of hyphae and sporangiophores so that identification becomes very difficult. In this method, sporangiophores, the hyphae, and the spores remain intact when stained.
When molds are collected from the environment, the Sabouraud’s agar is most frequently used. It is a simple medium constituting 4% glucose, 1% peptone, and 2% agar-agar. The pH of the medium is then adjusted to 5.6 to inhibit the growth of other bacteria.
For some molds the pH of Sabouraud’s agar is too low while the glucose content of the agar is too high. A better medium for the growth of these organisms is one contributed by C. W. Emmons known as Emmons' medium that contains 2% glucose, with 1% neopeptone, and an adjusted pH of 6.8 to7.0. To inhibit bacterial growth, 40 mg of chloramphenicol is added to one litre of the medium. In addition to these two media, cornmeal agar, potato dextrose agar, Czapek solution agar, are some of the available media for culturing molds.
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