Why and who put yeast in Ascomycota

Microscopy course 'Structure and function of plants',
Part mushrooms
(Jan Nechwatal / Ralf Vögele)

 

 

Preliminary meeting I

Importance of mushrooms for the global metabolism

 

•           Mushrooms cover a wide spectrum Hydrolases that can break down macromolecules.

•           The fragments are absorbed into the fungal cells and mostly aerobic reduced.
Examples: Breakdown of starch, cellulose and lignin.

•           Fungi are important organisms in the Decomposition of foliage andWood and therefore crucial for
the return of organic carbon to the material cycle
(=> Mineralization).

 

Interactions with plants

 

1. Braiding

 

Symbioses of mushrooms (Mycobiont) and cyanobacteria or green algae (Photobiont). They represent a unit Thalluswhich is mainly formed by the fungus. Lichen often colonize extreme habitats in which the individual partner cannot survive alone (mountains, arctic). The photobiont receives protection from drought and is supplied with minerals, the fungus uses the photosynthetic partner as a C, possibly N sources.

 

2. Mycorrhiza

 

symbiosis between plants and mushrooms in the root area, with low specificity. The plant experiences a better supply of water and minerals, especially phosphate, the mushroom receives carbohydrates. There are different types of mycorrhizae, with the VA mycorrhiza and the Ectomycorrhiza the most important are.

- Ectomycorrhiza

Dominated in moderate latitudes, especially on trees, mostly Basidiomycetes, more rarely Ascomycetes. Coating of the secondary roots with dense mushroom mycelium => coat. Infected roots stay short. The hyphae penetrate between the rhizodermis and the outer cortex cells => Hartig network. Minerals first get into the fungus before they are absorbed by the roots.

- VA mycorrhiza (vesicular-arbuscular type)

Dominated in the tropics,Zygomycetes The fungus invades with you Appressorium through the rhizodermis into the cortical layer. There the hyphae continue to grow and form in the inner cortex cells Arbuscle and Vesicle out. Arbuscules are enclosed by the vegetable plasma membrane of the cortical cells => exchange of substances

 

 

3. Phytopathogenic fungi

                                                                                                                     Diets:

necrotrophic - The parasite draws its energy from cells that have been killed (e.g. Cochliobolus heterostrophus).

optional / obligatory biotrophic - The parasite draws energy from living host cells (e.g. Cladosporium, Rust fungi, powdery mildew).

 

                                                                                                                     Penetration Strategies:

Directly through epidermal cells with the help of hydrolytic enzymes or pressure

Penetration through stomata. Penetration through wounds

 

Interactions with animals

                                                                                                                      

                                                                                                                     Insect pathogenic fungi

mostly specific

are used for biological control

 

                                                                                                                     Symbioses between fungi and insects

Leaf cutter ants

some Termites

 

                                                                                                                     Symbioses between mushrooms and ruminants

in the rumen there are chytridiomycetes, which split cellulose and gain energy through fermentation

Effects on humans

 

Mushrooms as pathogens
  • opportunistic mycoses (e.g. Candida albicans)
  • systemic mycoses, dangerous in immune problems such as AIDS (e.g. Cryptococcus neoformans)
  • Allergies
Mushrooms as destroyers of goods
  • Food spoilage due to mycotoxins (e.g. aflatoxin)
  • Damage to building material => wood degradation
Mushrooms as a food source
  • "Hat mushrooms" (Basidiomycetes), truffles (Ascomycetes), morels (Ascomycetes)
  • "Quorn": meat substitute, Fusarium (Ascomycet)
Biotechnological use
  • Fermentation: beer, wine, bread, soy sauce, etc.
  • Making cheese
  • Production of antibiotics: penecillin, cephalosporin, taxol
  • Production of technical enzymes
  • Organic acids production
  • Fungi as antagonists of pests

 

                   

Some technical terms in mycology

 

Hyphaefilamentous rows of cells from fungi
MyceliumThe entirety of the hyphae of a fungus
YeastsFungi that live as single cells and divide by budding
GametangiumSex organs that contain the sex cells (gametes) or nuclei that simply function as gametes. The gametangia are at Phycomyces (Zygomycete) are identical (Isogametangium), at Pythium (Oomyzet) on the other hand differently (Oogonium and Antheridium)
SeptaPartitions within a hypha that divide the hypha into mononuclear (haploid or diploid) or binuclear (dikaryotic) sections (cells). With differently designed Pores
Spursmostly in large quantities in Sporangia formed, specialized cells that serve to spread and after germination can grow into a new mycelium
HymeniumHyphae of asci or basidia
Conidiaasexually formed, uncultivated spores (spread by the wind); usually arise on special hyphae (Conidiophores)
Mitosporesspores formed after mitosis (e.g. conidia)
Meiosporeshaploid spores formed after maturation, e.g. Ascospores of the Ascomycetes and Basidiospores the Basidiomycetes
Oosporessexual, in Oogonia after fertilization through Antheridia resulting diploid spores of the Oomycetes
HeterothalliaSexual processes only take place between genetically different (+) and (-) mycelia or their sex cells (self-incompatibility)
PlasmogamyFusion of the plasma of male and female sex cells
KaryogamyFusion of sexually different, haploid nuclei
AscusTubular cell in which, after meiosis, usually eight haploids Ascospores arise (àAscomycetes)
BasidieCell from which, after meiosis, usually four haploids Basidiospores exogenous (in sterigms) arise (àBasidiomycetes)

 

 

 

Systematics of mushrooms

 

Empire: Protozoa

Tribe: Dictyosteliomycota (cellular slime mold)

Dictyostelium discoideum *

Tribe: Acrasiomycota (cellular slime molds)

Tribe: Myxomycota (real plasmodial slime molds)

Physarum polycephalum

Tribe: Plasmodiophoromycota

Plasmodiophora brassicae

Polymyxa graminis

Rich: Chromista

Tribe: Oomycota

Order: Peronosporales

Pythium sp.*

Phytophthora infestans

Kingdom: Fungi (Real Mushrooms)

Tribe: Chytridiomycota

Tribe: Zygomycota (yoke mushrooms)

Order: Mucorales

Phycomyces blakesleeanus *

Rhizopus stolonifer

Order: Endogonales (VA mycorrhia fungi)

Glomus mossae

 

Tribe: Ascomycota (ascomycota)

Saccharomyces cerevisiae* (Baker's yeast)

Fusarium graminearum *

Penicillium roquefortii *

Aspergillus nidulans *

Sordaria fimicola *

Blumeria graminis (Powdery mildew)

Claviceps purpurea (`Ergot´)

 

Tribe: Basidiomycota

Order: Agaricales

Agaricus bisporus *

Amanita muscaria

 

*) These species will be processed as part of the course.

 

literature:

 

Alexopoulos, C.J., Mims, C.W., Blackwell, M. (1996): Introductory Mycology. Wiley & Sons, New York.

Carlile, M.J. & Watkinson, S.C. (1995): The Fungi. Academic Press, London.

Strasburger, E. (1991): Textbook of Botany. Fischer Verlag, Stuttgart.

Esser, K. (1985): Cryptogams (2 volumes), Springer Verlag, Berlin.

 

   

 

Dictyostelium discoideum

 

Dictyostelium is a representative of the cellular slime molds (Dictyosteliomycota), which are assigned to the protists, but are not closely related to the real fungi. A large number of individual amoebas feed on the bacterial lawn. After consuming the food, the amoeba aggregate into one Pseudoplasmodiumthat moves as a functional unit and then to one Sorocarp (= Fruiting body: consisting of foot disc, stem [with later dying cells] and head [with spores]).

 

 

Development cycle of Dictyostelium discoideum (from Alexopoulos; Fig. 27-2)

 

 

Tasks:

The microscopy is carried out directly on the agar plate at the lowest magnification (3.2 x objective) or by binocular. The aggregation stage, pseudoplasmodium ('slugs') and Sorokarp (with footplate, stem and spore head) as well as transition stages of different stages of development should be observed and drawn.

 

 

Pythium sp.

 

Mushrooms of the genus Pythium often cause so-called fall-over diseases in young seedlings of various plants and are difficult to control. They are facultative parasites that can also feed saprophytically.Pythium belongs to the Oomycetes (just like the causative agent of potato rot, Phytophthora infestans, as well as the obligate biotrophic downy mildew fungi) and forms a diploid, unseptated mycelium.

The reproduction takes place asexually through sporangia, sexually through oospores. Depending on the temperature conditions, sporangia and oospores germinate either with a germ tube (at more than 18 ° C) or with the release of zoospores (at 10-18 ° C).

 

 

Development cycle from Pythium sp. (from Agrios: Phytopathology, Figure 11-16)

Tasks:

A dissecting needle is used to cut an approx. 5x5 mm piece down to the bottom of the plate from the overgrown nutrient medium. This is then placed on a slide so that the bottom of the piece is facing up. Finally, it is covered with a drop of water and a cover slip. Early stages with oogonia and antheridia (on the outer edge of the Petri dish) can be observed, as well as later stages with oospores, which are characterized by strongly thickened cell walls (in the middle of the Petri dish). The structures of both stages as well as the surrounding hyphae are to be drawn (40x objective).

 

 

Phycomyces blakesleeanus

 

The Zygomycota (yoke mushrooms) belong to the "real" mushrooms along with the Asco and Basidiomycota, while other groups such as the Oomycota are treated with the mushrooms ("sensu lato") for traditional reasons. Are zygomycetes Haplonts, contain Chitin as the main component of the cell wall and usually have a unseptated mycelium (Coenocytium). Mito- and meiospores are generally not flagellated. With the mitospores (vegetatively formed) there is a progression from endogenously formed sporangiospores to exogenously formed conidiospores. The formation of the zygote occurs through gametangiogamy (fusion of the reproductive organs).

 

Our course example, Phycomyces blakesleeanus, is a saprophyte found naturally on cow dung, bread and fatty substrates.P. blakesleeanus Depending on the environmental conditions, forms two different types of mitospores, the micro- and macrospores, which arise in micro- or macrosporangia. In the light (like our course plates) only the macrosporangia (spherical containers) are formed, which sit at the end of a few cm long, unbranched sporangia carriers (macrosporangiophores). A macrosporangium contains up to 100,000 spores. It reacts to gravity, light and touch stimuli. The latter can, if necessary, be observed on the lid of the Petri dish.

 

There P. blakesleeanus is heterothallic, it requires the simultaneous presence of two colonies that differ by the type of mating (- or +). In the light at temperatures below 25OC the sexual cycle is initiated. When the two mycelia come into contact, the hyphae tips thicken (formation of the zygophores), which wrap around each other and lift off the substrate. At the tip of the zygophores, the gametangia constrict, which look the same in both mating types (à isogametangiogamy) and are each carried by the suspensor. As the gametangia push apart during further growth, loops are formed. The two gametangia merge to form the zygosporangium (plasmogamy) in which pairs of - and + nuclei (karyogamy) and subsequent meiosis soon occur. During the maturation of the haploid zygospore, thick, black cell walls are formed with decorations similar to deer antlers on the outside, which make it easier for animals to spread.

 

 

 
 
 

(Pictures from http://www.es.embnet.org/~genus/phycomyces.html)

 

Life cycle of Phycomyces spp.

 

(from http://www.es.embnet.org/~genus/phycomyces.html)

 

 

Tasks:

The agar dishes are viewed with the microscope in the area of ​​the zygote line at the lowest magnification. Different stages of union with traghyphae (suspensors), gametangia and finished zygospores can be observed. Asexual macrosporangia are formed on upright traghyphae a few cm long. The different stages of development and structures should be drawn.

 

 

General characteristics of the Ascomycota

 

The Ascomycota (ascomycota) comprise a large number of species (> 300,000), belong to the "higher" mushrooms in addition to the Basidomycota and are characterized by the following characteristics:

 

·                    usually terrestrial way of life

·                    septate mycelium, with simple pores

·                    Cell walls mainly made of chitin and ß-glucans

·                    All spore forms are immobile

·                    haploid phase dominates in the life cycle

·                    short dikaryophase, maintained by "hook formation"

·                    Meiosis products in asci, these mostly in fruit bodies

·                    Mitospores: mostly exogenously formed conidiospores

 

 

 

Development of the Ascomycetes. Asci mostly arise in the fruiting bodies

 

 

Saccharomyces cerevisiae

 

Saccharomyces cerevisiae, The baker's or brewer's yeast (Ascomyzet) is of immense importance in the food industry. The term 'yeast' describes a type of vegetative growth in higher fungi, namely the sprouting of single cells. As we recently learned, neighboring yeast colonies can use ammonia gas to communicate with one another and prevent them from growing together; in this way they prevent excessive competition for food and can use the existing substrate “fairly”. With some fungi, a transition between both types of growth is possible (dimorphism), depending on environmental conditions or mutations. In the case of yeast which is pathogenic to humans Candida albicans E.g. the filamentous stage (see figure below) is decisive for the penetration of the host tissue and thus also for the pathogenicity.

S. cerevisiae Laboratory strains only grow as yeast cells, but naturally occurring wild types can also do so under suitable conditions Pseudohyphae (Middle thing between yeast single cells and real hyphae). In contrast to the yeast cells, the pseudohyphae are able to penetrate into a solid substrate, e.g. agar. This should be demonstrated in the course.

The wild type strain used in the course is diploid (like most naturally occurring Saccharomyces yeasts). This strain grows yeast-like on complete medium (YPD) on (nitrogen-limited) SLAHD medium with pseudohyphae (after approx. 4-5 days of incubation at 28-30 ° C).

 

 

Yeast cells (above) and pseudohyphae (below) from Candida albicans

 

Tasks:

Dividing yeast cells from a fresh liquid culture are examined under a microscope. For microscopy of pseudohyphae, the minimal medium plate without the lid is viewed with the 10x objective. A flat piece of agar a few mm wide with a few colonies is then removed with a scalpel, placed on a slide with a little water and examined under the microscope (40x objective).

 

 

Fusarium graminearum

 

In the food industry, biotechnology with mushrooms and their products plays a major role. By fermentation of the grain pathogen Fusarium graminearum (Strain A3 / 5) the mycoprotein with the trade name Quorn is obtained.

F. graminearum is the secondary crop form of the Ascomycetes Giberella zeae. So it is the genus Fusarium around vegetatively reproducing mushrooms (= Fungi Imperfecti). The structures of this type of reproduction are conidia or spores, the morphology of which helps to distinguish these fungi.

 

 

 

Tasks:

A dissection needle is used to cut an approx. 5x5 mm piece down to the bottom of the plate from the overgrown nutrient medium. This is then placed on a slide.

 

 

Aspergillus (Emericella) nidulans

 

The genus Aspergillus includes many species, which are mostly saprophytic, more rarely phytopathogenic or animal or human pathogenic.Some species are of great importance in the production of enzymes (e.g. amylases), chemicals (e.g. citric acid) or food (e.g. soy sauce). Most of the species in the group are traditionally classified among the Deuteromycetes ("Fungi Imperfecti"), which have no sexual stages (teleomorphs) or have not (yet) been found. However, there are some ways in the group like A. nidulans, which form sexually formed fruiting bodies (celistothecia: closed ascocarp), which makes it possible to classify them within the ascomycetes. These then also have the generic name "Emericella".

The species treated in the course can reproduce sexually or vegetatively, depending on the environmental conditions. The mitospores are exogenous conidiospores ("head mold") formed on club-shaped carriers. The following structures are formed from the substrate mycelium: foot cells, conidiophores, vesicles, metulae, phialides, conidia.

 

 

 

 

 

Conidiophores from Aspergillus sp.

Tasks:

At the second On the day of the course, material is removed from the colonies, placed on a slide with a drop of Abel's solution and carefully (!) Covered with a cover slip. Draw the conidiophores with the different cell types (metulae, phialides and conidia). In addition, the mushrooms are observed directly on the plates under the microscope in order to see the long chains of conidia (3.2x objective). You drive in from the edge of the colony to follow the development of the fungi.

 

 

Penicillium spp.

 

The genus Penicillium includes many species, most of which are saprophytic, more rarely plant or human pathogens. Penicillium- Species play an important role in the production of antibiotics (e.g. penicillin), food processing (e.g. camembert, salami) and the production of chemicals (e.g. oxalic acid). How Aspergillus becomes Penicillium classified with the Deuteromycetes. Teleomorphs belonging to the Ascomycetes are known in some species.

The species covered in the course is isolated from blue cheese and has been identified from not to the species level; possible here would be, for example. P. camembertii. The mitospores are conidiospores ("brush mold") formed exogenously on hyphae. The following structures are formed from the substrate mycelium: foot cell, conidiophores, ramus (branch, not present in all species), metula, phialids, conidia.

 

Conidia carriers of Penicillium sp.

 

Tasks:

On the first day of the course, starting from a blue cheese, Penicillium-Cultures can be set up on agar plates. To do this, a small amount of the cheese from the blue-green spore area (the size of a pinhead) is placed on a corn agar plate. On the second day of the course, material is removed from the colonies, placed on a slide with a drop of Abel's solution and carefully (!) Covered with a cover slip. Draw the conidiophores with the different cell types (metulae, phialides and conidia). In addition, the mushrooms are observed directly on the plates under the microscope in order to see the long chains of conidia (3.2x objective). You drive in from the edge of the colony to follow the development of the fungi.

 

 

Sordaria fimicola

 

S. fimicola is a dung dweller and related to Neurospora crassa, one of the best studied filamentous fungi. Sexual processes in ascomycetes usually take place in the fruiting bodies in which asci are formed, where nuclear fusion takes place and immediately afterwards the maturation process, from which eight ascospores generally arise after a further mitosis.

 

Sordaria fimicola:
Fruiting bodies and asci with ripe (pigmented) and immature (unpigmented) ascospores

tasks:

With tweezers, mycelium parts with fruiting bodies (perithecia) are removed from the borderline of the two fungal trunks and transferred to a slide with a drop of water. A cover slip is placed and the fruiting bodies are squeezed by applying pressure with the index finger so that the asci become visible. A fruiting body and several asci with different arrangements of dark and light ascospores are drawn (40x objective).

 

 

General characteristics of the Basidiomycota

 

The Basidiomycota (stand mushrooms) belong to the higher fungi next to the Ascomycetes and count about a third of all known fungi. They have the following characteristics:

 

  •   usually terrestrial way of life
  • septate mycelium, with complicated pores (e.g. dolipores)
  • Cell walls mainly made of chitin and ß-glucans
  • immobile spurs
  • The dikaryotic phase dominates the life cycle and is maintained by a complex pattern of division (buckle formation)
  • Meiosis products (basidiospores) are formed exogenously on basidia, these mostly in fruiting bodies (e.g. cap mushrooms!)

 

The fruiting bodies of the basidiomycetes consist of a two-nucleus hyphae. In the case of the lamellar fungi, basidia are formed on the edge of the exposed hymenium, in which the nuclear fusion and immediately after the meiosis take place Agaricusbisporus only 2) Basidiospores are shot down a few mm by an osmotic explosion mechanism and can thus be spread via the air.

 

 

 

Agaricus bisporus (Cultivated mushrooms)

 

Besides the fly agaric, the culture champion is probably the best-known representative of the basidiomycetes, but compared to other representatives it has a special feature. Agaricus bisporus forms per base only two, not four, basidiospores, as the name suggests.

 

Section through lamellae of a basidiomycete (http://www.biodiversity.ea.gov.au/biodiversity/abrs/publications/fungi/kingdoms.html#perithecium)

Basidia and basidiospores of some basidiomycetes (right A. bisporus)

(http://home.tiscalinet.ch/mki/images/Basidie.jpg, http://www.bfafh.de/inst4/44/cp_basidie2.htm, http://www.rz.uni-karlsruhe.de /~botanik/anf-prakt/ba-kut-3.html)