Lecture #18

Archiascomycetes: Early Diverging Ascomycetes

Open with impact of molecular data and polyphyly of yeasts

Archiascomycetes: Early Diverging Ascomycetes

• Diverse group including saprobic and parasitic fungi that have been grouped primarily on the basis of rDNA sequence analysis (Sugiyama and Nishida, 1994; Nishida and Sugiyama, 1993).
• Saitoella, and Schizosaccharomyces are asexual, saprobic soil-living yeast
• Taphrinaand Protomyces are dimorphic with a saprobic yeast stage and a parasitic mycelial stage on plant hosts
• Pneumocystis carinii, is the infective agent of a pneumonia that strikes individuals with supressed immune systems.

Members of the group lack ascogenous hyphae and ascocarps, and the asci have sometimes been homologized with sporangia. In some members of the group the asci are said to form within thick-walled ascogenous cells that have been called cysts or chlamydospores. Whether this is, in fact, merely a thick walled ascus or not is unclear.

Free cell formation

Studies of ascosporogenesis indicate that the process may occur either by formation of an EMS similar to the cylindrical type found in filamentous ascomycetes (in Pneumocystis) or by development of the EMS in association with individual nuclei in Taphrina and Schizosaccharomyces

Ascospore Release

• Ascospore release may be by forcible discharge of the ascus in some species of Taphrina and Protomyces, but may be variable even within these species depending on environmental conditions
• Ascospores are released passively upon rupture of the ascus wall in Schizosaccharomyces and Pneumocystis

Ploidy level of nuclei

• Taphrinahas a haploid yeast stage and a mycelium that is dikaryotic or multinucleate with paired nuclei
• Protomyceshas haploid and diploid yeast stages as well as a diploid mycelium
• Somatic cells of Schizosaccharomyces and Pneumocystis are haploid

Septal Pores

There is some question about the occurrence of simple septal pore and Woronin bodies in the mycelial archiascomycetes.

Classification

• Individual members of the archiascomycetes have been classified as protozoa, ascomycetes, basidiomycetes, oomycetes, and zygomycetes
• Taphrinahas been the object of special attention in phylogenetic speculation. It has been proposed as a primitive ascomycete closely related to rust fungi, as a basidiomycete, as an organism only distantly related to ascomycetes or basidiomycetes, as a sister group to the Saccharomycetales and as a member of the Saccharomycetales
• Protomyces,Saitoella, and Pneumocystis also have been suggested as being related to certain basidiomycetes at some time in their history.
• Pneumocystis, was considered to be a protozoan for many years, although evidence of ascosporogenesis has been available for some time

Archiascomycetes

• Taphrinales
  • Taphrinaceae Taphrina
  • Protomycetaceae Protomyces + 4 genera
• Schizosaccharomycetales
  • Schizosaccharomycetaceae Schizosaccharomyces
  • Pneumocystis

Taphrinales

Taphrina and members of the Protomycetaceae have mycelial stages that are parasitic on angiosperms, causing galls, thickenings, blisters, lesions and witches' brooms.

Protomycetaceae has a restricted host range and none of the species infects important crop plants; primarily weedy species in the Umbelliferae and Asteraceae

Taphrina occur on a wide variety of hosts, including some economically important species; best known of these is Taphrina deformans, the cause of peach and almond leaf curl diseases that produces obvious symptoms on the leaves and may lead to almost complete defoliation of the host tree;T. coerulescens on oak

• Dimorphic with a saprobic haploid uninucleate yeast phase that probably undergoes plasmogamy to produce the infective mycelial phase in the presence of the host
• Mycelial compartments are delimited by septa with simple pores, that appear similar to those of filamentous ascomycetes
• Mycelium is reported as dikaryotic or multinucleate with paired nuclei (Taphrina) or diploid (Protomycetaceae) from its initiation.
• Within the plant terminal or intercalary cells of the mycelium become differentiated as ascogenous cells and undergo development, eventually to produce ascospores.
• In Taphrina karyogamy occurs in the ascogenous cell, a process that takes place immediately after plasmogamy in the Protomycetaceae.

Classification

Taphrinales includes two families, the monotypic family Taphrinaceae with almost 100 species and the Protomycetaceae with five genera and twenty species

Taphrinaceae

• Mycelium of Taphrina is composed of septate hyphae that may be intercellular or subcuticular and sometimes grow within the walls of the epidermal cells of the host
• Asci are formed in a layer on the host surface from subcuticular mycelium that bursts through the cuticle
• Ascospores may bud within the ascus and the cells continue to bud after ascospore release to comprise the saprobic yeast stage
• Although the asci are forcibly discharged they rarely have been obtained in air samples, except in the immediate area of infected plants
• An the spring when growth resumes for the potential host, the yeast cells initiate the mycelial phase that infects the young tissues

Life Cycle

• Soon after they are formed, the ascospores produce small, round or ovoid spores by budding. This process may begin before the ascospores are released from the asci and continue after the spores have been released to the substrate. These spores, like the ascospores, are uninucleate and haploid. On the surface of the host, the spores continue to bud, then at some point they initiate the mycelial stage, perhaps by fusing in pairs after which they form the infective mycelium.

• Mycelium has often been said to be dikaryotic; however, evidence from serial sections and electron microscopy that showed multinucleate hyphal compartments with the nuclei arranged in pairs.

• Mycelium grows and branches, spreading between the cells and penetrating into the tissues of the host. Hyphal strands eventually become massed in the subcuticular region, and some of the hyphal cells enlarge to form ascogenous cells. Karyogamy occurs, apparently between the paired nuclei present within each ascogenous cell, and the cell begins to elongate. While elongation is proceeding, the diploid nucleus may divide mitotically, and the daughter nuclei move toward opposite ends of the cell. A septum develops between these two nuclei, separating the ascogenous cell into a basal stalk cell and an upper ascus cell. Meiosis and a subsequent mitotic division result in the formation of eight nuclei.

• Ascosporogenesis in Taphrina deformans takes place by the invagination of the ascus plasma membrane to form an EMS near the spindle pole body of each nucleus and eventually ascospores are delimited

• As the ascogenous cells enlarge and elongate, they exert pressure on the host cuticle from below, and eventually break through to form a compact surface layer of asci -- "the hymenium" -- on the epidermis of the host. The asci usually discharge the ascospores into the air.

• The ascus of Taphrina is unitunicate and without a differentiated apical apparatus for ascospore release. The tip of the ascus simply bursts at its thinnest point, responding to a buildup of pressure within the ascus. After the ascospores are released, they bud to form numerous spores, a process that may have begun in the ascus, and the life cycle is ready to begin anew.

Schizosaccharomycetales

Schizosaccharomyces, the fission yeasts, have been isolated from slime fluxes, honey, fruit, and fruit products, habitats that also are common to a number of the Endomycetales.

Characteristics

• Several characters distinguished these species from the yeasts of the Saccharomycetales including their method of cell division and cell wall polysaccharides.
• After nuclear division the cell elongates and a wall is produced between the daughter nuclei. The new wall material apparently originates from the area between the cell wall and plasma membrane abutting the original cell wall. It develops as an annulus with inward growth until the new daughter cells are completly partitioned. Parts of the devoloping wall thicken during their formation, and eventually separate in a middle translucent layer leaving scars at the ends of the new cells.
• Although low levels of chitin may be present in fission yeasts compared to the true yeasts, they also differ in other wall polysaccharides, particularly mannan structure and the presence of several unique major structural polysaccharides

Classification. Although the four species of the Schizosaccharomyces were traditionally included with the yeasts of the Saccharomycetales the genus long has been set apart from the group by its method of cell division and wall carbohydrates. Kurtzman (1993) used rDNA sequence analysis as the basis for placement in a separate order, and Sugiyama and Nishida (1994) included the genus in the archiascomycetes.

Pneumocystis

• An extracellular parasite of a number of mammals, including human beings. It is best known as the causal agent of a virulent pneumonia in immunocomprised humans, primarily those infected with HIV. The fungus binds to epithelial cells of the aveoli, and in addition to a lung infection, it may become disseminated to other organs and throughout the bone marrow.
• Pneumocystis cariniiwas first described as a protozoan from lungs of rats in France (Delanoe and Delanoe, 1912), but was not reported in humans until forty years later (van der Meer and Brug, 1942). Although it was known to occur in individuals who were severly malnurished or taking immunosuppressant drugs after transplant surgery or in cancer treatment, it was only after the spread of HIV that this organism became widely known (Masur,1989).
• Orginally described as a protozoan from rats; however, its taxonomic position as a protozoan was never clear. An ultrastructural study by Vavra and Kucera (1970) demonstrated an EMS in spore formation, an important character to distinguish them as ascospores. DNA sequences have made it clear that Pneumocystis carinii is a fungus (Edman et al., 1988); but, there has been some debate about what kind of fungus it actually is. Although the best evidence available indicates that Pneumocystis is a member of the archiascomycetes, the matter is not completely settled

Saccharomycetales (=Endomycetales): The Ascomycetous Yeasts

• Unicellular fungi that reproduce by budding
• The term yeast has no taxonomic standing and simply denotes a growth form
• Some zygomycetes, ascomycetes, and basidiomycetes may be dimorphic and shift from mycelial growth to yeast growth under certain environmental conditions
• Other fungi, both ascomycetes and basidiomycetes, exist primarily as yeasts most of their lives. It is important to note that although the unicellular stage often is emphasized as a taxonomic character, a number of the yeasts in the Saccharomycetales also produce mycelium.

Saccharomycetales

• Characterized, not by the yeast phase, but by the absence of ascogenous hyphae and ascocarps
• Asci of these fungi are thin-walled and may release their ascospores by deliquescing or eventually breaking; only a few have a specialized region for ascospore release
• Plasmogamy occurs by fusion of cells that function as gametangia.
• There is no dikaryotic phase in the life cycle, as karyogamy occurs immediately after plasmogamy; a diploid stage may persist in some species with cells multiplying by budding
• The zygote, derived from the copulation of two cells or parthenogenetically, either becomes transformed directly into an ascus or produces an erect, septate ascophore from which the asci develop
• Ascospore formation is similar to that of other ascomycetes; however, the EMS usually originates in association with the SPB of each individual nucleus
• Asexual reproduction is by budding, production of conidia, or mycelial fragmentation.
• Cell wall polysaccharides, primarily mannans and B-1,3 and B-1,6 glucans as well as reduced amounts of chitin, localized primarily in bud scars
• The group also has been characterised by DNA sequence analysis, a fact that has facilitated the placement of some yeast species without known sexual stages in Saccharomycetales. Molecular data also have been important in separating them from some of the archiascomycetes.

Members of Saccharomycetales are important in the baking, brewing, distilling, and related industries including fuel ethanol production, as supplements to food, in the manufacture of riboflavin and citric acid, as plant pathogens and mycoparasites, in the contamination and spoilage of foods, as human pathogens, as important components of ecosystems, and as model organisms for scientific study.

Food Yeasts

• It is their usefulness in conversion of a varity of raw materials low in protein to nutritious products that has led to efforts to develop their use as food.
• Substrates for conversion into single cell protein (SCP), as it has come to be known, include waste products from other processes such as cheese-making, potato processing, brewery wastes, and paper production.

Fermentations and Alcoholic Beverages. Yeasts almost certainly have been used by humans since the dawn of civilization. Their use is first documented about 6,000 B.C. Fermentations were welcome discoveries because they have the effect of preservation by lowering the pH and, in some cases, producing alcohol, conditions in which few microorganisms prosper; yeasts are noted particularly for their ability to ferment carbohydrates; hence, the name saccharomycetes (Gr. saccharon = sugar + myketes = fungi) is applied to them.

Human Pathogens. Many of us harbor a yeast as part of our normal intestinal and urogenital mycota that may turn on us one day causing severe infection and even death. Candida albicans is seldom isolated outside the bodies of animals, and is known from 58 species including wild and domesticated mammals and birds.

• Candida albicans, the most important disease agent of the genus, is not known to reproduce sexually and appears to be permanately diploid. This yeast may produce pseudomycelium and mycelium as well as yeast cells both in culture and in tissue

Somatic Structures

• Somatic structures and life cycles of members of Saccharomycetales are more variable than is generally recognized
• Except for the zygote, some species are unicellular throughout their life cycles
• Others may have a life cycle with both haploid and diploid phases; parthenogenic or asexual forms may remain haploid or diploid
• Some yeasts produce a number of buds in succession that remain joined for a period of time forming a pseudomycelium
• A septate mycelium may be produced by many yeasts and is sometimes abundant, particularly on low nutrient agars

• Several types of septa have been described in the hyphae of yeasts. Although the simple septal pore typical of the filamentous ascomycetes is known in some species, a septum with a single central or many micropores (perforate septum) is more common; the pores form after septum formation and the plasma membrane is therefore not continuous through the pores

Asexual Reproduction

• Budding and formation of arthrospores and conidia are the usual methods of asexual reproduction in Saccharomycetales.
• In Saccharomyces budding is initiated at the time of duplication of the spindle pole body
• An area of enzymatic wall softening within a newly developed chitinous ring allows a portion of the cell contents surrounded by newly synthesized wall to push out through the constricting ring.
• At the time nuclear division is almost complete, an abscission plate begins to form. This plate consists of a thin layer of chitin secreted at the invaginated plasma membrane of the separating cells.
• Separation of the bud from the mother cell then takes place and a bud scar is evident on the mother cell opposite the birth scar of the daughter cell. The bud scar resembles a crater on the surface of the mother cell with a raised circular rim with a punctation in the middle. By counting the bud scars on a cell, one can determine the number of buds that have been produced from it. Sometimes a succession of buds may be formed before each bud separates from the parent cell and the chain of buds is the pseudomycelium.
• The position of bud formation on a cell has been used as a character at higher taxonomic levels

(IMAGES)

Sexual Reproduction

Saccharomyces cerevisiae

• After plasmogamy is completed to produce the zygote cell, karyogamy typically occurs in this cell which usually is the young ascus
• In Cephaloascus and a few other genera an ascophore is produced
• There is no dikaryotic phase in yeasts
• After ascus development ascosporogenesis occurs; usually four ascospores are produced per ascus, but some species may have eight or even hundreds from postmeiotic mitoses.

Meiosis and Ascosporogenesis

• Both divisions of meiosis occur within the original intact nuclear envelope.
• SPBs are replicated at the beginning of the first division and migrate to lie on opposite sides of the nuclear envelope as the nucleus elongates.
• At the completion of the first division, the nuclear envelope is still intact.
• SPB replication then occurs for the second time, and the SPBs separate in pairs so that members of a pair are on opposite sides of the nucleus.
• The two spindles now are oriented perpendicular to the spindle of the first division.
• The nuclear envelope invaginates at the end of the second division, and the four nuclei at last separate
• EMS form in association with each nucleus.
• Ascosporogenesis in ascomycetous yeasts differs only in detail from that of other ascomycetes in the absence of a common vesicle enclosing all the nuclei before ascospore delimitation

Classification

Saccharomycetaceae

1. Have a predominantly unicellular thallus that may produce pseudomycelium,
2. Reproduce asexually primarily by multilateral budding, and
3. Produce ascospores in a free ascus that originates either from a zygote or parthogenetically from a single somatic cell.

The best known member of the family is Saccharomyces cerevisiae.

Life cycle of S. cerevisiae, an example of a species that may have both haploid and diploid budding stages

• Heterothallic, with two of the ascospores in each ascus carrying one mating type (a) and two, the other mating type (a)
• After cell fusion, the zygotes begin to bud and several generations of diploid cells ensue before ascospore formation occurs
• One to four globose, smooth-walled ascospores are formed
• Pseudomycelium may be formed on occassion

Nadsoniaceae and Saccharomycodaceae.

• Commonly known as apiculate (lemon-shaped) yeasts
• Species may have bipolar-budding
• Pseudomycelium is formed in some species, but never true mycelium.
• Saccharomycodes ludwigii; life cycle features a long diploid somatic stage and a very short haploid phase.

Eremotheciaceae and Metschnikowiaceae.

• Most important group of plant pathogens in Saccharomycetales, and cause damage to beans, tomatoes, nuts, and especially cotton. They probably are dispersed to flowers by hemipterous insects, and they enter the flower through the stigma (Batra, 1973).