Introduction

Fungi included in the Homobasidiomycetes possess holobasidia, in contrast to the heterobasidia (phragmobasidia) of the Heterobasidiomycetes, rusts and smuts (see Chapters 21–23). The traditional classification of the Homobasidiomycetes, founded by the nineteenth century Swedish mycologist Elias Fries, was based on a number of different arrangements of the hymenium on the hymenophore. The most common types, shown in Fig. 19.1, are (A) agaricoid, i.e. gill-bearing (lamellate); (B) poroid, i.e. bearing pores instead of gills; (C) hydnoid, i.e. with a toothed or spiny hymenium; (D) clavate, with a club-shaped or coralloid fruit body, the outside of which is covered by the hymenium; (E) resupinate, i.e. with a flattened (corticioid) hymenium appressed to the underside of solid surfaces; and (F) epigeous or (G) hypogeous gasteroid or secotioid (non-ballistosporic) hymenophores (see Chapter 20). It has long been suspected that the different hymenial arrangements have evolved separately in unrelated fungal groups, i.e. that they represent examples of convergent evolution. They can be interpreted as different ways of maximizing the hymenial area for a given amount of fungal tissue (Pöder, 1983; Pöder & Kirchmair, 1995). Examples of similar hymenophore arrangements in unrelated fungi are seen in the tubular hymenia characteristic of Boletus (Fig. 19.21) and Trametes (Fig. 19.26) or the toothed hymenia found in the homobasidiomycete Hydnum (Fig. 19.1c) and the heterobasidiomycete Pseudohydnum (Fig. 21.9b).

Introduction

Fungi living predominantly or exclusively as yeasts are encountered in three classes of Basidiomycota, namely the Heterobasidiomycetes (Chapter 21), Urediniomycetes (Chapter 22) and Ustilaginomycetes (Chapter 23). We shall discuss basidiomycete yeasts together in the present chapter because these organisms, although taxonomically diverse, are unified by many biological features.

Ecology

Little is known about the ecology of basidiomycete yeasts. They occur in marine and freshwater habitats, the soil and the plant rhizosphere, and especially on above-ground plant surfaces such as tree bark, leaves, flowers and fruits. A certain degree of specificity of yeast species relative to plant species or organs of a given plant host has been observed (Phaff, 1990). Basidiomycete yeasts may be found in all climatic zones from the arctic to the tropics. They generally exist as saprotrophic phylloplane organisms. When nutrients become available, there may be a steep increase in the population density of these yeasts. Many yeasts isolated from soil have their origin in vegetation which becomes incorporated into humus after leaf fall. Basidiomycete yeasts are not noted as plant pathogens, but some species can infect animals. Cryptococcus neoformans is one of the most serious fungal pathogens of humans (pp. 661–665). Members of another group (Malassezia spp.) live commensally on the skin of humans and other mammals, causing superficial dermatomycoses under suitable conditions (p. 671).

Introduction

The order Pezizales contains the operculate discomycetes which are the most readily recognized cup fungi. The order is large, containing some 15 families, about 160 genera and over 1100 species (Kirk et al., 2001). Most are terrestrial and saprotrophic on soil, burnt ground, decaying wood, compost or dung, but some form sheathing mycorrhiza (ectomycorrhiza) with trees (Maia et al., 1996). A somewhat exceptional case is Rhizina undulata, which causes root rot of conifers in plantation situations, usually starting from areas affected by recent fires (Callan, 1993). Whilst most species of Pezizales produce epigeous fruit bodies above ground level and have active ascus discharge mechanisms with wind-dispersed ascospores, the truffles (e.g. Tuber and Terfezia) form subterranean (hypogeous) ascomata. The dispersal of truffles relies on the ripe ascomata being eaten by rodents and other mammals attracted by their strong odour. The ascospores survive digestion and defaecation. There are also aquatic Pezizales, growing on wood in streams or other wet places. An overview of the Pezizales may be found in Pfister and Kimbrough (2001). Keys to genera are given by Korf (1972) and Dissing et al. (2000).

The ascocarp is generally an apothecium (p. 245) which can range in diameter from less than one millimetre to several centimetres. It is often cup-shaped or disc-like, fleshy, sometimes stalked, and frequently brightly coloured. The asci are, in most cases, cylindrical with a well-defined lid called operculum (see pp. 239–241) which is the characteristic feature of the Pezizales.

What are fungi?

About 80 000 to 120 000 species of fungi have been described to date, although the total number of species is estimated at around 1.5 million (Hawksworth, 2001; Kirk et al., 2001). This would render fungi one of the least-explored biodiversity resources of our planet. It is notoriously difficult to delimit fungi as a group against other eukaryotes, and debates over the inclusion or exclusion of certain groups have been going on for well over a century. In recent years, the main arguments have been between taxonomists striving towards a phylogenetic definition based especially on the similarity of relevant DNA sequences, and others who take a biological approach to the subject and regard fungi as organisms sharing all or many key ecological or physiological characteristics – the ‘union of fungi’ (Barr, 1992). Being interested mainly in the way fungi function in nature and in the laboratory, we take the latter approach and include several groups in this book which are now known to have arisen independently of the monophyletic ‘true fungi’ (Eumycota) and have been placed outside them in recent classification schemes (see Fig. 1.25). The most important of these ‘pseudofungi’ are the Oomycota (see Chapter 1973).

Introduction

The phylum Oomycota, alternatively called Peronosporomycetes (Dick, 2001a), currently comprises some 800–1000 species (Kirk et al., 2001). The Oomycota as a whole have been resolved as a monophyletic group within the kingdom Straminipila in recent phylogenetic studies (e.g. Riethmüller et al., 1999; Hudspeth et al., 2000; see Fig. 4.2), although considerable rearrangements are still being performed at the level of orders and families. A scholarly treatment of the Oomycota has been published by Dick (2001a) and will remain the reference work for many years to come. Because of the outstanding significance of Oomycota, especially in plant pathology, we give an extended treatment of this group.

The vegetative hypha

Although some members of the Oomycota grow as sac-like or branched thalli, most of them produce hyphae forming a mycelium. Oomycota are now known to be the result of convergent evolution with the true fungi (Eumycota), and their hyphae differ in certain details. However, the overall functional similarities are so great that they provide a persuasive argument for the fundamental importance of the hypha in the lifestyle of fungi (Barr, 1992; Carlile, 1995; Bartnicki-Garcia, 1996). Much physiological work has been carried out on hyphae of Oomycota (see Chapter 1), and the results have a direct bearing on our understanding of the biology of the Eumycota. Like them, the hyphae of Oomycota display apical growth and enzyme secretion, ramify throughout the substratum by branching to form a mycelium, and can show morphogenetic plasticity by differentiation into specialized structures such as appressoria or haustoria.

Introduction

Gasteromycetes are an unnatural assemblage of basidiomycetes sharing the common negative character that the basidiospores are not discharged violently from their basidia. Instead of the ballistosporic basidiospores of other basidiomycetes which are asymmetric in side view (see Fig. 18.5), those of the gasteromycetes are usually symmetrically poised on their sterigmata or are sessile. Dring (1973) has termed such basidiospores statismospores. Commonly the basidia open into cavities within a fruit body, and the basidiospores are released into these cavities as the tissue between them breaks down or dries out. A recognizable fertile layer (hymenium) may be present or absent (see Reijnders, 2000). The internal production of basidiospores has given the gasteromycetes their name (Gr. gaster = stomach). The gasteromycete fruit body is termed the gasterocarp, and the spore mass enclosed by the gasterocarp wall (peridium) is the gleba. Sometimes, as in Lycoperdon or Geastrum, the gasterocarp opens by a pore through which the spores escape, but in forms with subterranean (hypogeous) fruit bodies there is no special opening, and it is possible that the spores are dispersed by rodents and other burrowing animals (Colgan & Claridge, 2002). In Phallus and its allies, the basidiospores are exhibited in a sticky mass attractive to insects, whilst in Cyathus and Sphaerobolus the spores are enclosed in separate glebal masses or peridioles which are dispersed as units. In spite of these variations in gasterocarp morphology, the life cycles of most gasteromycetes follow the general Homobasidiomycete pattern outlined in Fig. 18.4.

Introduction

The Basidiomycota (colloquially basidiomycetes) are a large group of fungi with over 30 000 species. They include many familiar mushrooms and toadstools, bracket fungi, puffballs, earth balls, earth stars, stinkhorns, false truffles, jelly fungi and some less familiar forms. Also classified here are the rust and smut fungi, which are pathogens of higher plants and may cause serious crop diseases. Most basidiomycetes are terrestrial with wind-dispersed spores, but some grow in freshwater or marine habitats. Many are saprotrophic and are involved in litter and wood decay, but there are also pathogens of trees such as the honey fungus, Armillaria, which attacks numerous tree species, and Heterobasidion annosum, which can seriously damage conifer plantations. Common woodland mushrooms such as species of Amanita, Boletus and their allies grow in a mutually symbiotic relationship with the roots of trees, forming ectotrophic (sheathing) mycorrhiza. Species of Rhizoctonia, representing mycelial forms of basidiomycetes, behave as pathogens towards a wide range of plants but are mycorrhizal associates of orchids. As saprotrophs, basidiomycetes play a vital role in recycling nutrients but they also cause severe damage as agents of timber decay, e.g. dry rot of house timbers by Serpula lacrymans. The fruit bodies (basidiocarps) of many mushrooms are edible, and some are grown commercially for food, notably Agaricus bisporus (= A. brunnescens, the white button mushroom), Pleurotus spp. (oyster mushrooms) and Lentinula edodes (shii-take). It is also well known that the basidiocarps of certain mushrooms are poisonous to eat, e.g. Amanita phalloides (the death cap).

Introduction

The phylum Ascomycota (colloquially called ascomycetes) is by far the largest group of fungi, estimated to include more than 32 000 described species in 3400 genera (Kirk et al., 2001). It is assumed that the majority of ascomycetes has yet to be discovered, and the total number of species may well be higher by a factor of 10–20 or even more (see Hawksworth, 2001). The name is derived from the Greek words askos (a leather bottle, bag or bladder) and mykes (a fungus), so ascomycetes are sac fungi. The characteristic feature of the group is that the sexually produced spores, the ascospores (see p. 25), are contained within a sac, the ascus. In most ascomycetes the ascus contains eight ascospores and is turgid, ejecting its spores by a squirt mechanism.

There is a very wide range of lifestyles. Some ascomycetes are saprotrophs, others necrotrophic or biotrophic parasites of plants and animals, including humans. Examples of biotrophic parasites are the Erysiphales, the cause of many powdery mildew diseases of plants (Chapter 13), the Taphrinales (p. 251) causing a range of plant diseases associated with growth abnormalities, and the Laboulbeniales, relatively harmless ectoparasites of beetles and some other insects (Blackwell, 1994; Weir & Blackwell, 2001). Many ascomycetes grow as endophytes in symptomless associations with plants. Some are mutualistic symbionts, for example the lichens (Chapter 13) which make up about 40% of the described species of ascomycetes.

Introduction

The class Plectomycetes originally contained all ascomycetes which produce their asci within a cleistothecium, i.e. a ‘closed case’. DNA sequence comparisons have revealed that this character was a fairly good one because, with the major exception of the powdery mildews (Erysiphales; see Chapter 13) and few scattered examples in the Pyrenomycetes (Chapter 12) and Helotiales (Chapter 15), most cleistothecium-forming fungi and the anamorphs associated with them have been found to be monophyletic (Berbee & Taylor, 1992a; Geiser & LoBuglio, 2001). As they stand now, the Plectomycetes can be defined by the following set of characters (Alexopoulos et al., 1996; Geiser & LoBuglio, 2001).

(1) A cleistothecium or gymnothecium is usually present; a cleistothecium proper has a thick and continuous (pseudoparenchymatous) wall, whereas in the gymnothecium the wall consists of an open cage-like construction of hyphae, the reticuloperidium (Greif & Currah, 2003). Naked asci are produced only in rare cases.

(2) Ascogenous hyphae are usually not conspicuous.

(3) Asci are scattered throughout the cleistothecium, not produced by a fertile layer (hymenium).

(4) Asci are mostly globose and thin-walled, and the ascospores are released passively after disintegration of the ascus wall, not by active discharge.

(5) Ascospores are small, unicellular and usually spherical or ovoid.

(6) Conidia are commonly produced from phialides (in Eurotiales) or as arthroconidia, which are typically formed as chains of conidia alternating with sterile cells. An arthroconidium becomes released when the neighbouring cells disintegrate. This rhexolytic secession is typical of the microconidia of Onygenales and Ascosphaerales.

Introduction

The characteristic feature of this group is that the ascus is bitunicate and fissitunicate; it has two separable walls (see p. 240). The outer wall (ectotunica or ectoascus) does not stretch readily, but ruptures laterally or at its apex to allow the stretching of the thinner inner layer, the endotunica or endoascus (Figs. 17a–c). Asci are generally non-amyloid. The fruit body with asci is regarded as an ascostroma, and each cavity in which asci develop is termed a locule. In contrast to the Hymenoascomycetes, in which ascocarps develop following plasmogamy and the pairing up of two genetically dissimilar nuclei (ascohymenial development), in the Loculoascomycetes the ascoma is already present before the compatible nuclei are brought together (Barr & Huhndorf, 2001). The development of asci in pre-formed locules is called ascolocular. The ascostroma has therefore been defined as an aggregation of vegetative hyphae not resulting from a sexual stimulus (Wehmeyer, 1926). However, Holm (1959) has questioned the accuracy of this definition, since examples are known where the ascocarps do develop following the pairing of nuclei (Shoemaker, 1955). Within the developing ascocarp, one or more locules are formed by the downgrowth of pseudoparaphyses (see below) and the development of asci. One or more ostioles then develop by the breakdown (lysis) of a pre-formed mass of tissue (lysigenous development). Where a single locule develops, a structure resembling a perithecium results and, although this term is commonly used for such loculoascomycete fruit bodies, they should strictly be called pseudothecia (see p. 245).

Major advances, especially in DNA-based technology, have catalysed a sheer explosion of mycological knowledge since the second edition of Introduction to Fungi was published some 25 years ago. As judged by numbers of publications, the field of molecular phylogeny, i.e. the computer-aided comparison of homologous DNA or protein sequences, must be at the epicentre of these developments. As a result, information is now available to facilitate the establishment of taxonomic relationships between organisms or groups of organisms on a firmer basis than that previously assumed from morphological resemblance. This has in turn led to revised systems of classification and provided evidence on which to base opinions on the possible evolutionary origin of fungal groups. We have attempted to reflect some of these advances in this edition. In general we have followed the outline system of classification set out in The Mycota Volume VII (Springer-Verlag) and the Dictionary of the Fungi (ninth edition, CABI Publishing). However, the main emphasis of our book remains that of presenting the fungi in a sensible biological context which can be understood by students, and therefore some fungi have been treated along with taxonomically separate groups if these share fundamental biological principles. Examples include Microbotryum, which is treated together with smut fungi rather than the rusts to which it belongs taxonomically, or Haptoglossa, which we discuss alongside Plasmodiophora rather than with the Oomycota.

Introduction

In contrast to the Pezizales (see preceding chapter) which produce apothecia with asci discharging their spores through a detachable lid at their apex, the asci of inoperculate discomycetes liberate their spores either through a valve or a slit. In the inoperculate as well as operculate discomycetes, the asci may contain two or more layers, i.e. they are often described as bitunicate. However, these layers do not separate during ascus discharge, i.e. they are non-fissitunicate. Fine structural details of the asci of Helotiales have been described by Verkley (1993, 1994, 1996). Two large ecological groups of inoperculate discomycetes can be distinguished: the lichenized and non-lichenized species. This feature correlates approximately with the taxonomy at the level of orders, and here we shall discuss the Helotiales (sometimes alternatively called Leotiales) which contain mostly non-lichenized fungi. The Lecanorales and other orders with mainly or exclusively lichen-forming fungi are described in Chapter 16.

Those relatively few phylogenetic studies that have so far been performed on the Helotiales lack the necessary power of resolution to delimit natural groups. Thus, it is not clear at present whether this order is monophyletic or not, and several different classification schemes are in use (Gernandt et al., 2001; Kirk et al., 2001; Pfister & Kimbrough, 2001). The families currently associated with the Helotiales are listed in Table 15.1. Thus circumscribed, the order Helotiales contains some 2300 species. The Orbiliaceae, formerly included here (Pfister, 1997), are now considered to be more closely related to the Pezizales.

INTRODUCTION

Phagocytosis, an essential component of the innate immune response, is a complex process whereby extracellular particles of diameter ≥ 0.5μm are internalized into a specialized membrane-bound vacuolar compartment. Following particle engulfment the resulting vacuoles, better known as phagosomes, undergo a maturation sequence involving fusion and fission events with components of the endocytic pathway. The resulting hybrid compartments, termed phagolysosomes, acquire the molecular machinery necessary to destroy the ingested pathogens.

The process of internalization can be envisaged as involving five distinct steps, beginning with particle recognition, receptor signaling, membrane remodeling and actin polymerization allowing for pseudopod extension, and climaxing with particle uptake into the cytosol. Although the mechanism of internalization is largely conserved among phagocytic cell types, the purpose of phagocytosis is diverse. Unicellular organisms such as amoebae engulf bacteria to obtain nutrients. In mammalian cells, neutrophils and macrophages employ phagocytosis to prevent the spread of infectious agents, but the same process is also employed for the clearance of apoptotic bodies.

Phagocytosis is a receptor-mediated event initiated by ligand recognition and particle binding. Multiple receptors recognize and prompt the elimination of the varied army of pathogenic organisms; a different set of receptors is needed to identify and clear apoptotic bodies. Indeed, the former elicit an immune and inflammatory response, whereas the latter do not.

The objective of this review is to summarize succinctly the current knowledge of the events that lead to phagocytosis.

INTRODUCTION

The epithelial layer of mucosal surfaces in the gastrointestinal tract is a busy surface for communication between the host and both commensal and pathogenic bacteria. In the case of invasive pathogens, the epithelium represents the major barrier that has to be overcome. Important virulence determinants include those mediating adhesion to and invasion of the epithelium. Different strategies are used to induce the uptake of invasive bacteria (Figure 8.1). For example, Yersinia spp. and Listeria monocytogenes express adhesins that bind tightly to host cell-surface proteins. This binding interaction initiates invasion by triggering signaling pathways normally involved in the regulation of cell adhesion. This invasion mechanism has been termed “zipper” because the host cell membrane is wrapped tightly around the bacteria (Mengaud et al., 1996). For more details on the zipper mechanism; see Alonso and Garcia-del Portillo (2004). More recently, a variant of this mechanism, termed the “tandem β-zipper,” has been identified. As an example of this entry mechanism, pathogenic bacteria like Staphylococcus aureus express surface proteins that bind to human fibronectin. This fibronectin coat facilitates binding to host cell-surface receptors and mediates invasion (Figure 8.1b) (Schwarz-Linek et al., 2004). A third strategy, termed “trigger mechanism,” is employed by Salmonella and Shigella spp. This involves specialized protein-transport systems called type III secretion systems (TTSS) to inject virulence factors (effector proteins) directly into the cytosol of the host cells. The translocated effector proteins trigger host signaling cascades that mediate a variety of responses, including pathogen uptake (Hueck, 1998).

INTRODUCTION

The airway epithelium represents a primary site for the introduction and deposition of potentially pathogenic microorganisms into the body through inspired air. The ciliated epithelium lining the airways possesses several mechanisms to prevent colonization by inhaled bacteria and, despite repeated exposures to a wide variety of organisms, the lower respiratory tract usually remains sterile. The airway is defined anatomically as the upper respiratory tract, which includes the nasal sinuses and the nasopharynx, and the lower respiratory tract, which begins at the larynx and continues to the trachea, before dividing into the smaller airways until they reach the alveoli. The luminal surface of the airways is lined by a layer of epithelial cells. In the conducting airways, these cells are pseudostratified columnar epithelial cells, which become simple cuboidal epithelium as the branches extend to the alveoli (Diamond et al., 2000). The respiratory epithelium is an essential barrier that features tight intercellular apical junctions between the cells, a superficial liquid layer or film that contains mucous-gland and goblet-cell secretions, immunoglobulins, and lysozyme, components that are propelled and cleared by cilia.

INNATE HOST DEFENSES AGAINST BACTERIAL LUNG PATHOGENS

In the upper airways, the nose functions as a filter by trapping large particulate matter (>10 μm) in nasal hair or on the surface of the turbinates and septum. Smaller particles, including bacteria ranging in size from 2 μm to 10 μm, are inhaled and deposited in the lower conducting airways.

Bacteria must overcome multiple obstacles in order to achieve successful pathogenesis. In many cases, this requires bypassing the first line of host defense: the barrier provided by epithelial surfaces of the integument and the gastrointestinal, respiratory, and urinary tracts. To overcome these barriers, pathogenic organisms frequently initiate mechanisms that exploit essential cellular processes of the epithelium. These cellular processes are therefore critical to our understanding of bacterial pathogenesis. Their description is the goal of this chapter.

GASTROINTESTINAL TRACT

The gastrointestinal epithelium forms a critical interface between the internal milieu and the lumen. The latter should be considered the external environment, since the gut is essentially a tube running through the body that communicates with the external environment at each end. Thus, like the skin, the barrier formed by the gastrointestinal epithelium is critical in preventing noxious luminal contents from accessing the internal tissues. In contrast to the skin, the gastrointestinal tract must also support passive paracellular and active transcellular transport of nutrients, electrolytes, and water. The barrier formed by the gastrointestinal epithelium must therefore be highly regulated and selectively permeable. Consistent with this, barrier permeability and epithelial transport function vary at individual sites within the gastrointestinal tract according to regional differences in the specific nutrients and ions transported.

The oral cavity and esophagus are lined by stratified squamous epithelium (Figure 1.1), much like the skin.

INTRODUCTION

Consumption followed by digestion has developed from a nutrition mechanism in unicellular eukaryotes into a highy regulated and indispensable mechanism of host defense against infection in mammals. Phagocytosis of pathogenic microorganisms by phagocytes, or “eating cells,” is a major host defense mechanism of the innate immune system. The process of phagocytosis was first described at the beginning of the twentieth century by Elie Metchnikoff, who observed ingestion of small particles by cells from starfish larvae. Phagocytosis is generally defined as the internalization of particles with a diameter of at least 0.5μm, such as bacteria, viruses, parasites, large immune complexes, or apoptotic cells and cell debris. Ingestion of smaller particles, such as small immune complexes or other macromolecules, occurs through a fundamentally distinct mechanism, called endocytosis. Phagocytosis and endocytosis are distinguishable by the importance of actin polymerization, which directs membrane motility during phagocytosis, but not endocytosis. Another distinction can be made by the presence of clathrin coats around vacuoles formed during some forms of endocytosis, but not phagocytosis (Greenberg 1986). Recently, one more distinction has been added by showing that endocytosis by IgG Fc receptors (FcγR), but not phagocytosis, requires ubiquitylation (Booth et al. 2002). Thus, the specific molecular pathways that direct the process of ingestion depend on the size of the particle. When the target particle is too large to be ingested, a process designated “frustrated phagocytosis” may occur, involving activation of pathways partially similar, but not identical, to those activated during phagocytosis.