L.2 –Biotechnology Mycology D.Ebtihal Muiz Life cycles, spore formation and germination The times at which a fungus reproduces sexually and asexually, the nuclear condition and morphology of these stages are known as the life cycle of that fungus. The life cycles of a fungus reflects the adaptations of that organism to its environment and/or niche, and is of great interest from an evolutionary and biological point of view. A complete understanding of an organisms life cycle is necessary if we want to control or use a particular species. As you might expect for the fungi, life cycles are not as consistent as those found for flowering plants. The life cycle of an organism can be described as the series of events from zygote to zygote. Superimposed on the life cycle is often an asexual reproductive phase - a manner of increasing numbers of individuals without going through sexual reproduction (fusion, karyogamy and meiosis). Asexual reproduction depends on mitotic division of the nucleus, requires less expenditure of energy and reduces the role of chance. On the negative side, it provides less genetic variation and relies on mutation and the parasexual cycle as a source of variation. The structures formed during asexual reproduction include: zoospores (Chytridiomycota), sporangiospores (Zygomycota) and conidia (Ascomycetes, Basidiomycetes). The generic term for asexual spores is mitospore. Sexual reproduction requires the fusing of two cells and/or nuclei, sometimes through the production of specialized structures such as gametangia. Thus the process involves chance (since the two mates must find each other) and is energetically more expensive than asexual reproduction On the positive side it provides new combinations of alleles. Structures formed during meiosis include: resting sporangia giving rise to zoospores (Chytridiomycota); zygospores (Zygomycota); asci and ascospores (Ascomycota); basidia and basidiospores (Basidiomycetes). One thing is quite clear in the fungi, their life cycles are quite different from those of higher plants and animals where the organism is diploid and meiosis occurs in cells which are totally dependent on the diploid organisms to produce haploid cells which are short-lived since they must fuse to form a zygote or die. Haploid-diploid - these phases alternate regularly, an unusual condition in fungi restricted almost entirely to a few species of aquatic Chyridiomycota. Diploid - the haploid phase is restricted to the gametes or gametangial phase. Most of the Oomycota may conform to this pattern 1 and it is more similar to your life cycle and that of flowering plants. This supports the alignment of this group with the Stramenopila. Spore Germination Spores are generally considered dormant structures. Dormancy is any rest period or reversible interruption of the phenotypic development of the organism. The spore stage is a quiescent phase situated in the life cycle of a fungus between two phases of active growth. There are two basic types of dormancy: constitutional (memnospores) and exogenous (xenospores). Constitutional dormancy is due to an innate property of the species. For example a spore might have a physical or chemical barrier in the wall that inhibits the uptake of water or nutrients or there may be a metabolic block that needs to be overcome or an inhibitor that must be leached out before germination will occur. With exogenous dormancy, theoretically the spore is capable of germinating immediately after it is produced but it remains dormant due to unfavorable environmental conditions such as water availability, temperature, pH, etc. Factors required for germination. I. Water. Water is essential for spore germination and the amount required varies among taxa. 80% humidity - Aspergillus, Penicillium 90-95% - Alternaria, colletotrichum, Ustilago, Cladosporium More that 95% - Venturia, Magnesia Liquid water - many species including Endoconidia, peronosporales, Sclerotinia conidia, rust spores, etc. 2. Temperature. The temperature required for germination varies with the species and often reflects geographical distribution, habitat and life cycle. 3. Nutrition. Many spores carry with them internally all the nutrients needed but some require a special exogenous factor, e.g., vitamins, inorganic ions, particular carbon source. 4. pH. Spores of most fungal species will germinate at a pH between 4.56.5. 5. Oxygen. Small quantities of oxygen are needed for spore germination and low quantities of oxygen are not thought to limit germination. Changes during spore germination. 1. Water content increases and the spore swells due to the absorption of water. The spore comes out of the quiescent metabolic state as enzymes go into solution. 2 2. Vacuolation increases. 3. Endoplasmic reticulum and other cell organelles increase. 4. Respiration increases and the mitochondria enlarge and form more cristae. 5. Vesicles thought to be involved in wall synthesis also appear. 6. In spores with lipid bodies, the lipid bodies disappear as the lipids are used up as an energy source. Spores in general: by wet weight spores generally contain 25% protein and 20% fat. and they have a low water content relative to vegetative mycelium. Cell walls of spores are generally not fibrillar, but they are multi—layered and often contain melanin and have Ornamentations. Spores contain all normal mycelial organelles. Respirators reserves include lipids, glycogen, phospholipids and polysaccharides that can include sugar alcohols Like Trehalose), Respiration rates in spores are only 1-4% those of vegetative mycelium, but obviously the more reserves a spore has, the longer it will survive. Dormancy: Dormancy occurs when spores do not immediately germinate after formation. Dormancy is a break in the life cycle. There are two types, endogenous (constitutive) and exogenous (induced). Endogenous dormancy is clue to sonic internal quality of the spore, a barrier to water or nutrient entry, a metabolic block, or an inhibitor. Self inhibition prevents spores from germinating in dense suspensions. It can be by excessive sensitivity to oxygen or carbon dioxide levels, nutrient competition, or most usually due to the presence of inhibitors. These molecules are often active in the 1-10 nanomolar range. These inhibitors have to be leached away before germination takes place. There can also lie physical barriers to germination. In one of the athlete foot fungi, Microsporium gypseum, there is a protein layer around the spore which prevents the uptake of water. This layer is removed by the action of a fungal acid phosphatase enzyme. This enzyme is inhibited by high levels of phosphate, and until phosphate levels in the environment drop the fungus spore does not germinate. 3 Endogenous (induced) dormancy occurs because of some external condition, and whilst these conditions prevail the spore will not geminate. As soon as the limiting factor is removed the spore germinates. Table I . A summary of the characteristics of fungal spores with endogenous and exogenous dormancy: Endogenous dormancy: Exogenous dormancy: Displaced from point of origin Remain at porn of origin Definite launch mechanisms Released by autolysis Small and thin walled spores Large and thick walled Short survival time Survive for a long time Germinate readily tinder suitable Germinate after a specific conditions stimulus or removal of an inhibitor Optimal environmental signals trigger the end of dormancy and the onset of germination. Chemical stimuli can trigger germination. This is frequently seen in pathogens where host compounds can act as germination stimulants. Germination begins with imbibition, the uptake of water, which can cause a 3 to 20 fold increase in size. Spherical growth also accounts for some of the swelling. Eventually polarized growth starts, with the emergence of a germ tube from the spore. The spore wall may he ruptured and a new cell wall covered germ tube emerges, or the spore wall may be softened and the germ tube then emerges. 4 General View of fungi What is a fungus? 1. Heterotroic organism devoid of chlorophyll 2. Has cell walls 3. Non motile (exception is zoospores) 4. Reproduces by spores 5. Primary storage product is glycogen 6. Usually filamentous, eukaryotic and multicellular Hypha (pl. hyphae) 1. Filaments constituting the body (thallus) of a fungus. 2. Hyphae elongate by apical grow th. 3. Hyphac arc referred to, collectively, as mycelium (pl. mycelia). 4. Hyphae have cross walls called septa (sing. septum). 5. Hyphac with septa are known as septate. 6. Hyphae without septa arc known as aseptate or coenocytic. 7. Fungal cell wall consists of B linked glucans and chitin. Hyphae organized into structures 1. Stroma (pl. Stromata)— compact, somatic structures that is cushion -like on or in which fruiting bodies are formea. 2. Sclerotium (pl, sclerotia) — hard testing body resistant to unfavorable conditions. 3. Rhizomrph —strands to thick shoestring—like Structures that exhibit a high degree of internal structure where hyphae may lose their individuality and have a thick rind and growing tip. Hyphal behavior may indicate type of organism 1.Nectroph (Perthotroph) — uses enzymes or toxins to kill host cells in advance of their hypltae and then hyphae grow between dead or dying cells. 2. Biotroph (obligate parasite) — obtains nutrients only from living cells, They have specialized hyphal branches (haustorium —pl. haustoria) that penetrate the host cell wall and invaginate the host cell membrane 3. Hemibiotroph — requires living host cells but soon act like nectrophs (Colitotrichium, lindemuthianumn). 5 Other hyphal structures 1. Appressorium (pl. appressoria) — specialized infection structures that form at the tips of hyphae. 2. Appressoria adhere to the host surface and form penetration pegs. 3. Penetration pegs can penetrate hard artificial surfaces such as gold foil and nonbiodegradable Mylar membranes. 4. Protoappressoria — slight hyphal tip swellings that act like an appressoria. 5. Reproduction in Fungi 6. Fungi have both asexual (anamorphic) and sexual (teleomorphic) reproduction. 7. Some fungi are holocarpic (entire fungus converts into one or more reproductive structures). 8. Most fungi are eucarpic (reproductive organs arise from only a portion of the fungus body — thallus — pl. thalli). Asexual Reproduction 1. Fragmentation of somatic hyphae or throcpores (thallic conidia) 2. Fission of somatic cells into daughter cells 3. Budding 4. Mitotic spores Asexual spores I. Chlamydospores arc thick walled thaIlic conidia that function as a resting spore 2. Asexual spores are either sporangiospores or conidia. 3. Sporangiospores vs. Conidia 4. Sporangiospores are borne in a sac called a sporangium. The cytoplasm within the sac is cleaved into sporangiospoies. The sporangium is borne on a specialized hypha known as a sporangiophore. 5. A conidium is borne on the tip or side of a specialized hypha known as a conidiophore. Sexual reproduction in Fungi 1. Plasmogamy — (p!) fusion of protoplasts (bring nuclei close together). N+N 2. Karyogamy (k!) — fusion of nuclei. 2N 3. Meiosis (mi!)— reduction of chromosome number to haploid state. N 6 4. In some fungi, k! follows p! quickly; in others, K! is separated from p! by time and space. Sex Spores in Fungi 1. Most plant pathogenic fungi exist as N or N+N and are only 2N in the zygote. 2. In the Oomycota, individuals are 2N for most of their life cycle. 3. Some fungi have hyphae with different genotypes of nuclei within the same hyphal cells. This is known as heterokaryosis. Such individuals are referred to as being heterokaryotic. 4. Gametes in Fungi 5. Sexual organs — gametangia 6. Isogamete I isogametangia — male and female are indistinguishable 7. heterogametes I heterogametangia — male and female are morphologically different 8. Male — antheridium; femaIe — oogonium 9. Classification How Fungi Obtain Nutrients All fungi obtain food by secreting digestive enzymes and then absorbing the organic molecules produced (external digestion). — extensive hyphae network provides enormous surface area for absorption — many fungi able to break down cellulose in wood Ecology of Fungi • Fungi and bacteria are the principal decomposers in the biosphere. — mineral cycling Fungi are virtually the only organisms capable of breaking down lignin. • Fungi often act as disease—causing organisms for both plants arid animals. —agricultural damage — human health • Mutualistic associations — lichens — fungi and green algae — mycorrhizae — fungi and plant roots 7