The Fungal Kingdom

In 1996 the genome of Saccharomyces cerevisiae was published and marked the beginning of a new era in fungal biology ( 1 ). Since then, rapid advancements in both sequencing technologies and computational biology have resulted in the sequencing of genomes for more than 800 species (e.g., http://genome.jgi.doe.gov/fungi/). These genomes represent a windfall of data that are informing evolutionary studies of fungi and the search for biological solutions to alternative fuels, bioremediation, carbon sequestration, and sustainable agriculture and forestry ( 2 ). Indeed, the marriage between genomics and phylogenetics occurred early, both in the use of phylogenetic techniques to study genome evolution and in the use of genome-scale data to infer evolutionary relationships. In this article we will review the impact of genomic-scale phylogenies, along with standard molecular phylogenies, on our understanding of the evolution of the fungal tree of life and the classification that communicates it.

The fungal lineage is one of the three large eukaryotic lineages that dominate terrestrial ecosystems. They share a common ancestor with animals in the eukaryotic supergroup Opisthokonta and have a deeper common ancestry with plants, yet several phenotypes such as morphological, physiological, or nutritional traits make them unique among all living organisms. This article provides an overview of some of the most important fungal traits, how they evolve, and what major genes and gene families contribute to their development. The traits highlighted here represent just a sample of the characteristics that have evolved in fungi, including polarized multicellular growth, fruiting body development, dimorphism, secondary metabolism, wood decay, and mycorrhizae. However, a great deal of other important traits also underlie the evolution of the taxonomically and phenotypically hyperdiverse fungal kingdom, which could fill up a volume on its own. After reviewing the evolution of these six well-studied traits in fungi, we discuss how the recurrent evolution of phenotypic similarity, that is, convergent evolution in the broad sense, has shaped their phylogenetic distribution in extant species.

It is a certainty that the tree of life can be broken up into large units of biodiversity that span such great evolutionary distances that one could call them kingdoms. However, if these units are to have any meaning, they must represent radiations of diverse groups underpinned by unique shared derived adaptations that drove the evolutionary success of each group. Without such evolutionary characters, these classifications are merely abstract concepts or arbitrarily demarcated lineages. Even the five kingdoms (commonly called plants, animals, fungi, protists, and prokaryotes) that gained popular acceptance from Whittaker ( 1 ) were abstractly defined and nonmonophyletic. These kingdoms were defined based on cellular organization (prokaryote or eukaryote), multicellularity (protist or “higher” eukaryote), and trophic level (producer [plants], consumer [animals], or decomposer [fungi]). Somewhat more phylogenetically sound definitions ( 2 ) have replaced these classifications, but the names, defining characters, and number of kingdoms are still not settled (e.g., 3 , 4 ).

In 1825, Elias Magnus Fries (1794–1878) predicted that the fungi would prove to be the largest group in the vegetable kingdom, analogous to the insects in the animal kingdom. Notwithstanding that fungi are not actually part of the plant kingdom, how right he has proved to be as the bicentenary of his prediction approaches. By the 1960s a few mycologists were speculating that there might be as many fungal as plant species, but almost no attempts to calculate estimates from the available data were made. As concern over the conservation of biodiversity in general grew in the subsequent decades, culminating in the signing of the Convention on Biological Diversity in 1992, more precise figures on species numbers of all kinds of organisms were required. A series of estimates of the number of fungi settled on figures ranging from 500,000 to almost 10 million species, with 1.5 to perhaps 5 million receiving most support among mycologists. A recent study even predicts up to a trillion species of microorganisms globally ( 1 ); how many of these are supposed to be fungi is not specified, but if this estimate holds true and only 1% of these were fungi, the global estimate of fungal diversity would be a thousand times higher than the current highest estimate of 10 million species.

Microsporidia were initially described about 150 years ago with the identification of Nosema bombycis as the organism responsible for the disease pébrine in silkworms ( 1 ). Microsporidia are ubiquitous in the environment and infect almost all invertebrates and vertebrates, as well as some protists ( 2 ). Spores from microsporidia are commonly found in surface water ( 3 ). These organisms are eukaryotes that have a nucleus with a nuclear envelope, an intracytoplasmic membrane system, chromosome separation on mitotic spindles, vesicular Golgi, and a mitochondrial remnant organelle lacking a genome termed a mitosome ( 4 ). For insects, fish, laboratory rodents, and rabbits microsporidia are important pathogens, and they have been investigated as biological control agents for destructive species of insects ( 2 ). Several species of microsporidia have caused significant agricultural economic losses including Nosema apis and Nosema ceranae in honeybees ( 5 ), Loma salmonae in salmonid fish ( 6 ), and Thelohania species in shrimp ( 7 ). Franzen ( 8 ) published an excellent review of the history of research on these pathogens, and a recent textbook by Weiss and Becnel ( 2 ) provides a comprehensive examination of what is known about these organisms. In 1977 Sprague elevated the class or order Microsporidia to the phylum Microspora ( 9 ), and a decade later Sprague and Becnel ( 10 ) suggested that the term Microsporidia should instead be used for the phylum name. These organisms were previously considered “primitive” protozoa ( 11 ), but molecular phylogenetic analysis has resulted in the insight that these organisms are not primitive but instead are degenerate, and that Microsporidia are related to the Fungi, either as a basal branch of the Fungi or as a sister group ( 12 – 16 ).

There are ∼64,000 known species within the Ascomycota, making it the largest phylum of Fungi. Major subphyla include the Taphrinomycotina (e.g., Schizosaccharomyces pombe), the Saccharomycotina (including Candida and Saccharomyces clades), and the Pezizomycotina (the largest subphylum, which includes the Eurotiomycetes, Dothideomycetes, Sordariomycetes, and Leotiomycetes) (see Fig. 1 ). Most Saccharomycotina grow as budding yeast or are dimorphic (can grow as yeast or filaments), whereas most Pezizomycotina are predominantly filamentous, although some are also dimorphic.

In the phylum Basidiomycota, a wide variety of lifestyles are represented. These range from well-known and conspicuous wood-decaying mushrooms, plant growth-promoting and mutualistic mycorrhizae, and crop-destroying smut and rust fungi, to yeast-like human pathogens. Lifestyle differences have consequences for the mating and breeding systems of these fungi (see “Glossary,” below, for definitions of specialist terms used in this article), which are reflected in the genetic evolution of mating-type determination. For over a century fungi have been recognized as having diverse breeding systems, from homothallism (i.e., universal compatibility among gametes, including among clonemates) to heterothallism (i.e., mating among haploid gametes carrying different mating-type alleles). The study of breeding systems, for example, led to the discovery of the astounding variability in mating-type alleles among mushrooms, with thousands of different mating types in some species ( 1 ), and to the realization that in many fungal pathogens the process of sexual reproduction is closely linked to infection and pathogenicity ( 2 ) ( Fig. 1 ). The importance of basidiomycete fungi and their great research tractability, from ecology to genomics, have brought major insights into the diversification of genetic mechanisms used to achieve sexual reproduction.

The Mucoromycota is a newly formalized phylum of fungi that are one of what are sometimes considered the basal lineages in the fungi ( 1 ). These species have undergone a different evolutionary trajectory than the Ascomycetes and Basidiomycetes. Generally, the species are difficult to develop into experimental models, but despite this our understanding of mating and sex in the fungi overall has been punctuated with major discoveries being made in this lineage.

Sexual reproduction is a ubiquitous feature of the eukaryotic kingdom with the many benefits of sex in generating genetic diversity as substrates for evolutionary selection being well known. When two different partners come together, there is the generation of genetic variation in the offspring, through the processes of crossover and recombination during meiosis, enabling response of future generations to environmental selection pressures ( 1 – 4 ). Sexual reproduction also allows the repair of random epigenetic or conventional genetic damage by recombination with homologous chromosomes and can mask lethal mutations ( 4 , 5 ). In addition, sexual recombination alleviates clonal interference and prevents deleterious mutations hitchhiking to fixation ( 6 ). Indeed, there are so many benefits to sexual reproduction that exceptions that are purely asexual have been termed “evolutionary scandals” ( 7 ). As a result, supposed ancient asexual species such as the bdelloid rotifers (an exclusively female class of over 460 rotifer species thought to date back several million years) and darwinulid ostracods (a family of around 30 crustacean species thought to have been exclusively female and asexual for over 200 million years, but for which very rare living males have recently been described) have gained notoriety ( 8 – 10 ). It therefore comes as a great surprise that, until recently, approximately 20% of all fungal species were considered to reproduce only by asexual means, with no recognized sexual cycle, based on knowledge of described fungal species ( 11 , 12 ). Indeed, in some phylogenetic groupings such as the Ascomycotina, up to 40% of taxa surveyed were deemed to be asexual ( 13 ). This is despite the fact that sexual reproduction in fungi can have additional benefits such as the production of fruit bodies and sexual spores that are resistant to adverse environmental conditions, thereby promoting survival of sexual offspring; it can provide a transient diploid arena for selection of genes; and sex can favorably impact genome evolution ( 14 – 17 ). Asexual species are also proposed to be short-lived evolutionary “dead ends” subject to rapid extinction ( 12 , 18 ). Fungal species lacking a known sexual cycle have been referred to as “imperfect” or “mitosporic” fungi and have been grouped into the “Fungi Imperfecti” or “Deuteromycota,” although phylogenetic analysis has shown that these are artificial groupings not based on taxonomic relationship ( 19 ). The terms “asexual” and “imperfect” are used synonymously in this review.

Cell-cell fusion is an essential biological process that occurs in organisms throughout the tree of life. It is involved in both sexual and asexual developmental processes in most species and has been shown to occur in multicellular and in unicellular organisms. Somatic cell fusion events are widespread in eukaryotic organisms, including animals, where they are important for muscle differentiation, placental development, and formation of multinucleate giant cells in the immune system ( 1 – 4 ).

Filamentous fungi are a large and ancient clade of microorganisms that occupy a broad range of ecological niches ( 1 , 2 ). Fungi are recyclers, being major decomposers of plant debris ( 3 ); they form mycorrhizal symbiosis with 93% of all flowering plant families ( 4 ), and they serve in the industrial production of proteins ( 5 ). However, fungi pose a threat to public health, the ecosystem, and our food security ( 6 , 7 ). The success of filamentous fungi is largely due to their elongate hypha, a chain of cells separated from each other by septa ( 8 ). Hyphae grow rapidly by polarized exocytosis at the apex ( 9 – 11 ), which allows the fungus to extend over long distances and invade many substrates, including soils and host tissues. Hyphal tip growth is initiated by establishment of a growth site and the subsequent maintenance of the growth axis, with transport of growth supplies, including membranes and proteins, delivered by motors along the cytoskeleton to the hyphal apex ( 12 ). Among the enzymes delivered are cell wall synthases that are exocytosed for local synthesis of the extracellular cell wall ( 13 ). Exocytosis is opposed by endocytic uptake of soluble and membrane-bound material into the cell ( 14 ). The first intracellular compartment in the endocytic pathway is the early endosomes (EEs), which emerge to perform essential additional functions as spatial organizers of the hyphal cell ( 15 ). Individual compartments within septated hyphae can communicate with each other via septal pores, which allow passage of cytoplasm or organelles ( 16 ) to help differentiation within the mycelium ( 17 ). This article introduces the reader to more detailed aspects of hyphal growth in fungi.

Fungal cell walls are dynamic structures that are essential for cell viability, morphogenesis, and pathogenesis. The wall is much more than the outer layer of the fungus; it is also a dynamic organelle whose composition greatly influences the ecology of the fungus and whose composition is highly regulated in response to environmental conditions and imposed stresses. A measure of the importance of the cell wall can be appreciated by the fact that approximately one-fifth of the yeast genome is devoted to the biosynthesis of the cell wall ( 1 , 2 ). Of these approximately 1,200 Saccharomyces cerevisiae genes ( 2 ), some are concerned with the assembly of the basic components, others provide substrates for wall materials, and many regulate the assembly process and couple this to environmental challenges. They include genes that encode carbohydrate active enzymes (which can be found in the CAZy database [http://www.cazy.org]) ( 3 ) and include multigene families of chitin and glucan synthases as well as remodeling enzymes such as the glycohydrolases (glucanases, chitinases) and transglycosidases. Many of the building blocks of the cell wall are conserved in different fungal species ( 4 ), while other components of the wall are species-specific, and there is perhaps no part of the cell that exhibits more phenotypic diversity and plasticity than the cell wall.

Decomposer fungi, by their very nature, continually deplete the organic resources in which they grow and feed. They therefore rely on continual successful spread to new resources. In terrestrial ecosystems resources are distributed heterogeneously in space and time ( 1 , 2 ). They are often discrete, ranging in size from small fragments, e.g., bud scales, to large tree trunks, though discrete leaves en masse can form a continuous layer on the forest floor. The processes of arrival and spread are thus crucial to the success of saprotrophic fungi. Following arrival at a resource, their competitive ability determines whether they are successful in colonization and how long they retain that territory. Colonization and competition are the main focus of this paper and are discussed separately below, largely drawing on wood decay fungi for illustrative examples.

The relative degree to which organisms move is a process operating at multiple temporal and physical scales ( 1 ). In recent years dispersal has received a great deal of attention in fields ranging from mathematics and physics to ecology and molecular biology, but only a patchy framework exists to explain dispersal over very large distances. Modeling patterns of long-distance dispersal (LDD) among macroorganisms, ranging from vertebrates and flying insects to seed plants, appears tractable, but documenting the geographic distributions and dispersal dynamics of microscopic propagules and microbes presents multiple theoretical and methodological challenges ( 2 – 4 ). The majority of empirical research directly measuring the dispersal of microbes or microscopic propagules is restricted to relatively short distances, and tracking dispersal at greater spatial scales involves mathematical or genetic models, e.g., in studies of moss ( 5 – 9 ), ferns ( 10 – 13 ), bacteria ( 14 – 19 ), and fungi ( 19 – 23 ). However, fitting dispersal data (e.g., from the tracking of spore movement) to mathematical functions often over- or underestimates LDD and imprecisely describes the trajectory of spore movement across large distances ( 24 – 28 ). Inferences based on population genetics data capture rare instances of successful LDD but incompletely describe underlying demographic processes and typically cannot speak to mechanisms of LDD ( 1 ). Besides the limitations of mathematical and genetic methods, important details about the natural history of species are often ignored or remain unknown, leaving many questions unanswered, including, e.g., how ephemeral propagules remain viable while exposed to harsh environments over extended periods of time.

Growth as an interconnected mycelial network is characteristic of filamentous fungi and has been subject to scientific investigation since the seminal works of Buller at the start of the 20th century ( 1 – 3 ). We have increasingly detailed understanding of the fundamental cellular processes needed to form a network, such as hyphal tip growth ( 4 ), septation ( 5 , 6 ), hyphal orientation ( 7 ), branching ( 8 ), and fusion ( 9 – 13 ) ( Fig. 1A and C ). In contrast, we know far less about the molecular events at the next physical scale that leads to hyphal aggregation and hyphal differentiation, and how these impact physiological processes such as long-distance resource distribution and biomass recycling. For example, while direct uptake and intrahyphal nutrient diffusion may be sufficient to sustain short-range local growth when resources are abundant ( 14 ), long-distance translocation is required to deliver nutrients at a sufficient rate to growing tips, particularly in fungi that form large networks on the forest floor that are too large to distribute nutrients through diffusion alone. We know little about the quantitative contribution of different potential transport pathways, such as cytoplasmic streaming, vesicle transport, growth-induced mass flow, or evaporative mass flow, to net fluxes and overall nutrient dynamics, and how they might vary between species and developmental stage ( 15 – 17 ). Nevertheless, the behavior of the growing mycelial network emerges from the interaction of many such processes and requires an integrated view to understand the overall impact on fungal behavior ( 18 – 20 ). Our understanding is further constrained by inferences drawn from a limited number of genetically tractable model filamentous species grown under laboratory conditions (abundant, evenly dispersed, low-molecular-weight resources, high relative humidity, constant light and temperature) compared with real-world conditions (patchy, recalcitrant, ephemeral resources, fluctuating temperature, light and relative humidity).

The significance of fungi in natural environments is extensive and profound. Their most obvious roles are as decomposers of organic materials and as animal and plant pathogens and symbionts. It is therefore obvious that they are of major importance in the global carbon cycle through such activities and as important determinants of plant growth and productivity. However, their importance in terms of nutrient and element cycling greatly extends beyond these core activities, and they are involved in the biogeochemical cycling of many other elements and substances, as well as many other related processes of environmental significance. The growing discipline of geomicrobiology addresses the roles of microorganisms in geological and geochemical processes ( 1 , 2 ), and geomycology can be considered to be a part of this topic that focuses on the fungi ( 3 , 4 ). The often clear demarcation between mycological and bacteriological research has ensured that the geoactive properties and significance of fungi have been unappreciated in wider geomicrobiological contexts. The range of prokaryotic metabolic diversity found in archaea and bacteria, including their abilities to use a variety of different terminal electron acceptors in respiration and effect redox transformations of many metal species ( 5 , 6 ), has also contributed to a narrow overall view of the significance of eukaryotic organisms in important biosphere processes. A recent collection of geomicrobiology review articles managed to completely exclude fungi (as well as algae), even to the extent of defining “microbes” as being only bacteria and archaea ( 7 ). Nevertheless, appreciation of fungi as agents of geochemical change is growing, and their significance is being discovered even in locations not usually regarded as prime fungal habitats, e.g., rocks, acid mine drainage, deep aquatic sediments, hydrothermal vents, and the igneous oceanic crust ( 8 – 11 ). Their significance as bioweathering agents of rocks and minerals is probably better understood than bacterial roles ( 12 ), and this ability is of prime importance in the weathering of human structures in the built environment and cultural heritage ( 13 – 15 ). On the positive side, the geoactive properties of fungi can be used for human benefit, and several aspects may contribute to providing solutions to several important global challenges. Geomycology is relevant to reclamation and revegetation of polluted habitats, bioremediation, nuclear decommissioning and radionuclide containment, biorecovery of important elements, and the production of novel biomaterials. This chapter outlines important geoactive properties of fungi in relation to important environmental processes, their positive and negative applications, and their impact on human society.

Plant pathogens are parasites that live at the expense of their host. While fungal pathogens are the largest group of plant pathogens, other important plant pathogens include bacteria, protists, chromists, nematodes, and even plants. Although this wide variety of pathogens share many aspects in epidemiology and management, here we deal only with fungal plant pathogens. The economic importance of fungal plant pathogens in the production of food, feed, materials, and ornamentals is undisputed ( 1 ). Direct costs include yield loss and use of resistant cultivars or pesticides. Indirect costs include the inability to grow certain crops or cultivars at a given location. Inspection and quarantine protocols to prevent the dispersal of pathogens ( 2 ) are indirect costs that are rarely taken into account. In contrast, as will be shown in this review, plant pathogens in nature are regarded as crucial contributors to the maintenance of biodiversity, similar to the role major animal predators play in wildlife.

The “Aquatic Phycomycetes” (sensu Sparrow) ( 1 ) constitutes an ecologically and economically important assemblage of eukaryotic microorganisms that share many morphological traits and ecological functions and interact with each other in the same aquatic ecosystems. There is molecular and structural evidence that the aquatic phycomycetes is a diverse, polyphyletic assemblage of species. For many years little research has been conducted with the aquatic phycomycetes, possibly because they were thought to be ecologically and commercially insignificant, but this perception has recently changed. Many of these species have been found to play key roles in biomass conversion in food webs ( Fig. 1 ) and in the carbon cycle ( 2 ).

To respond to the changing environment, cells must be able to sense external conditions. This is important for many processes including growth, mating, the expression of virulence factors, and several other regulatory effects. Nutrient sensing at the plasma membrane is mediated by different classes of membrane proteins that activate downstream signaling pathways: nontransporting receptors, transceptors, classical and nonclassical G-protein-coupled receptors, and the newly defined extracellular mucin receptors. Nontransporting receptors have the same structure as transport proteins, but have lost the capacity to transport while gaining a receptor function. Transceptors are transporters that also function as a receptor, because they can rapidly activate downstream signaling pathways. In this review, we focus on these four types of fungal membrane proteins. We mainly discuss the sensing mechanisms relating to sugars, ammonium, and amino acids. Mechanisms for other nutrients, such as phosphate and sulfate, are discussed briefly. Because the model yeast Saccharomyces cerevisiae has been the most studied, especially regarding these nutrient-sensing systems, each subsection will commence with what is known in this species.

Sunlight, harvested by photosynthetic organisms (plants, algae, and some bacteria) and used to synthesize energy-rich molecules (sugars) from carbon dioxide and water, provides the energy required to sustain life on Earth. In addition, sunlight properties such as intensity, duration, polarization, and spectral composition are used as sources of information ( 1 ). Indeed, all forms of life are continuously obtaining and decoding information from their environment. In fungi sunlight, ranging from ultraviolet (UV) to infrared wavelengths, regulates a diversity of biological processes including circadian rhythms, morphogenesis, tropism, and synthesis of pigments, among others (reviewed in reference 1 ). UV light can be harmful, since DNA modification products of photochemical reactions may be transmitted to the next generation as a mutation. Visible light appears not only to provide early warning of the presence of impending UV radiation and further damage, but also seems to contribute to the capacity of these organisms to deal with abiotic stress in general ( 2 – 5 ). Thus, the ability of most fungi to perceive and respond to light has very likely contributed to their survival and fitness.

Planet Earth plays host to an extravagantly diverse range of fungal species. Recent estimates suggest the probable existence of as many as 3 million fungal species ( 1 ), and the circa 75,000 of these that have been characterized to date display a wide range of lifestyles. Many fungi occupy specific niches within natural environments, playing essential roles in nutrient scavenging and recycling. Some thrive in close harmony with species from other kingdoms, a superb example being the mycorrhizal fungi, which display mutualistic interactions with plants. Other fungi are pathogenic, causing devastating infections of plants or animals. Indeed, the global threats that fungi pose to human health and food security are being increasingly recognized ( 2 ). Fortunately, a relatively small number of fungal species cause infections in humans (circa 400 species are described in the Atlas of Clinical Fungi [ 3 ]). Some of these fungi normally occupy environmental niches but are capable of colonizing and damaging human (or animal) tissues, whereas other fungi appear to be obligately associated with their host.

The growth and development phases of most fungi take place on a two-dimensional surface or within a three-dimensional matrix. The fungal sense of touch is therefore critical for fungi in the interpretation of their environment and is often involved in the switch to a new developmental state. Contact sensing, or thigmo-based responses, include thigmo differentiation, such as the development of invasion structures by plant pathogens; thigmonasty, where contact with a motile prey rapidly triggers its capture; and thigmotropism, where hyphal growth direction is guided by physical features in the environment. Like plants and some bacteria, fungi grow as walled cells. How do fungi, as walled organisms, not only sense contact, but also interpret the signal in a developmental context?

Melanins are dark pigments that are made by diverse fungi ( 1 , 2 ). Even fungi that produce white colonies, such as Candida albicans, have the ability to make melanins ( 3 , 4 ). Melanins have elicited considerable interest in microbial pathogenesis because they are important virulence factors for many pathogenic microbes, and their presence is associated with reduced susceptibility to antifungal drugs ( 5 , 6 ). Melanins are multifunctional molecules that give cells structural strength as well as reduced susceptibility to temperature extremes, heavy metals, and molecules produced by the immune system such as oxygen- and nitrogen-derived oxidants and microbicidal proteins ( 2 , 7 – 10 ).

Despite years of study and a great deal of exposure in the scientific press, there is still sometimes confusion regarding what are the defining characteristics of a rhythmic process that makes it truly circadian. The essence of this is that the rhythm must be shown not to depend in any way on continued environmental cycles, and to have a period of about a day. So, in this way, it is obvious that in petri dish cultures grown under a light-dark cycle, a rhythmic change in orange color in Neurospora crassa, or in melanin in Aspergillus fumigatus, could not be categorized as circadian, because the change in pigmentation would be seen as the result of light induction of, respectively, carotenoid or polyketide synthesis. As humans, our pupils constrict in proportion to the amount of light to which the eye is exposed, so a daily rhythm in pupil size would not be circadian but would instead reflect a diurnal cycle in ambient light. This part of the definition identifies the rhythm as truly endogenous in nature, in this way being the overt expression of an internal circadian oscillator. Beyond this, the definition is more subtle and reflects the biology of rhythms.

A universal feature of all organisms is their ability to respond to nutrient availability by regulating growth and developmental programs. The identification of the target of rapamycin (TOR) pathway was a seminal discovery in the quest to understand the molecular mechanisms that govern such processes. TOR is an evolutionarily conserved serine/threonine kinase belonging to the family of phosphatidylinositol kinase-related kinases. Other members of this family include the mammalian DNA damage checkpoint kinases ATM and ATR,which are conserved from yeast to human, and DNA-PK and SMG1, which are not found in yeasts. TOR regulates growth (accumulation of mass), proliferation (accumulation in cell number), and survival in response to nutritional changes by diverse mechanisms that include regulation of anabolic and catabolic metabolism, nutrient uptake, protein translation and turnover, gene transcription, and the epigenome (reviewed in 1 – 3 ).

DNA is the master instruction set, encoding everything that makes cells work. So it is no surprise that faithful duplication of DNA and proper distribution of new copies to daughter cells is highly choreographed and tightly controlled. The appearance of nuclei changes in predictable patterns during the cell cycle, passing through the morphologically uneventful interphase into the dramatic rearrangement of mitosis. The core cell cycle machinery is highly conserved among eukaryotes. Many of the mechanisms that control the cell cycle were worked out in fungal cells, taking advantage of their powerful genetics and rapid duplication times.

Chromosomes of fungi are linear segments of DNA, covered by a diverse assembly of RNA and proteins. They contain three landmarks required for function, namely, origins for DNA replication, centromeric DNA as attachment points for kinetochores, and telomere repeats to circumvent the end replication problem for linear DNA. Because fungi have been excellent model organisms for trail-blazing basic research since the adoption of Neurospora crassa as one of the workhorses for genetics in the 1940s ( 1 ), much of the foundation for general knowledge of eukaryotes was first uncovered with fungi, specifically the four species uniquely suited for genetics, biochemistry, and genomics: Schizosaccharomyces pombe, Saccharomyces cerevisiae, Aspergillus nidulans, and N. crassa. This has also been true for studies on chromatin and chromosomes.

Cellular ploidy is the number of complete sets of chromosomes in a cell. Many eukaryotic species have two (diploid) or more than two (polyploid) sets of chromosomes ( 1 ). These diploid and polyploid states are often the result of ancient whole-genome duplication (WGD) or hybridization events that occurred throughout the evolution of plants, animals, and fungi ( 2 – 4 ). Ploidy changes also occur during the development of many organisms and can vary within different tissues of the same organism and between individuals of the same species. For example, ploidy changes occur during the sexual cycle of eukaryotes, from haploid gametes to diploid somatic cells. Additionally, some cells continue to increase in ploidy during development, resulting in somatic tissues that have a mixture of diploid and polyploid cells, including human hepatocytes and megakaryocytes ( 5 – 7 ). These ongoing, developmentally programmed changes in ploidy are important for viability and are beneficial to many organisms ( 8 ), but the mechanisms controlling ploidy and the physiological significance of each ploidy level are not well characterized.

Studies of fungal evolution require an understanding of the phylogenetic relationships and relative evolutionary divergence of organisms. The first approaches to organizing fungi into related groups relied on morphological characteristics ( 1 ). These approaches provided a broad framework to organize fungal organisms for taxonomic classification based on recognizable morphological characteristics such as spore shape, asexual and sexual structures, and in mushroom-forming fungi, the shape and presence/absence of gills, veil attachments, and spore color. In zoosporic chytrid fungi the characteristics seen by scanning electron microscopy of zoospores reveal that the ultrastructure of the kinetosomes and flagellum are all diagnostic for the classification of many lineages ( 2 ). However, the microscopic nature of many fungi and especially of yeast-forming fungi with limited visible differences, and the prevalence of convergent evolution to homoplasies or similar characteristics across a tree, has made taxonomic classification of groups of fungi difficult or easily misled. The invention and application of DNA sequencing ( 3 ) and PCR ( 4 ) and the development of primers to amplify fungal rRNA enabled a new era of molecular phylogenetic studies in fungi ( 5 ). These approaches provided invaluable information that was used to resolve the major fungal lineages ( 2 , 6 – 20 ) and the delineation of species ( 21 – 24 ). Using DNA approaches to study the entire fungal tree of life provided new insight into the order of branching of major groups and the timing of morphological changes such as the loss of the flagellum found in zoosporic fungi ( 14 , 17 , 18 , 25 ).

Genetic variation is the stuff of evolution: if there is no variation, there can be no evolution. This review of fungal genetic variation begins with a survey of its sources and then discusses means of associating natural genetic variation with phenotypic variation, including phenotypes that are important to fungal adaptation or those that interest cell and developmental biologists engaged in basic or translational research. The tale of the study of fungal genetic variation is also a tale of advances in genomic science, and it is appropriate to note that the first Eukaryote to have its genome sequenced was a fungus, the model yeast Saccharomyces cerevisiae ( 1 ). Shortly thereafter, yeast was joined by filamentous Ascomycota, e.g., Neurospora ( 2 ), and Basidiomycota, e.g., Phanerochaetae ( 3 ). Not long after, three Aspergillus species were sequenced—Aspergillus oryzae ( 4 ), Aspergillus fumigatus ( 5 ), and Aspergillus nidulans—and the comparisons of their genomes ( 6 ), along with those of yeasts closely related to S. cerevisiae ( 7 ), represent landmarks in the field of comparative fungal genomics. A survey taken in April 2017 of fungal genomicists associated with the Joint Genome Institute and FungiDB estimated the number of fungal species with sequenced and assembled genomes at 2,000 and estimated that another 1,000 to 1,500 genomes represent multiple individuals from species that are studied by population genomics. No other group of eukaryotes enjoys as deep a genomic database as is seen for the fungi.

RNA interference (RNAi) is a mechanism conserved in eukaryotes that represses gene expression by means of small noncoding RNAs (sRNAs) of about 20 to 30 nucleotides (nt). Before the identification of the RNAi mechanism, its effects were observed in different organisms and described as independent processes. These effects were first observed in plants, in which the introduction of sequences intended to increase the production of floral pigments had the opposite effects, resulting in albino transformants. Hence, this phenomenon was called cosuppression ( 1 , 2 ). The same phenomenon was also observed in the fungus Neurospora crassa, in which an albino phenotype was obtained after transformation with extra copies of the gene al-1, which is involved in the production of carotenoids ( 3 ). The characterization of this phenomenon revealed reversibility of the albino phenotype, since some descendants of the original albino transformants reverted to the wild type phenotype. At that time, this phenomenon was not linked to the process of cosuppression observed in plants and was named “quelling.” The discovery of quelling in N. crassa represented a milestone in the field of RNAi, because this fungal model allowed the use of classic genetic tools to unravel the machinery of RNAi. The molecular mechanism of RNAi was finally uncovered by Fire et al., who discovered the central role of double-stranded RNA (dsRNA) in the RNAi of Caenorhabditis elegans ( 4 ). This central role of dsRNA was soon established in all the organisms harboring a functional RNAi mechanism, including plants, fungi, and animals. Cosuppression, quelling, and other posttranscriptional gene-silencing-related phenomena were integrated into the same conserved mechanism of RNA interference ( 5 ).

In 1994, a paper signed by a single author based on genetic approaches opened a decisive breach leading to a dramatic expansion of our perception of the biological significance of the prion phenomenon ( 1 ). In the following years, biochemical reconstitution and transformation established that these biological entities, originally identified and defined in the context of mammalian diseases such as Kuru or Creutzfeldt-Jacob disease, also exist in yeast as “protein-based genes” and correspond to previously described non-Mendelian genetic elements ( 2 , 3 ). It is now clear that in most known cases the physical basis for prion propagation is the formation, growth, and fragmentation of an amyloid aggregate. Amyloids are ordered protein polymers with a so-called cross-β structure ( 4 ). The original definition of prions as “infectious proteinaceous particles” is imprecise enough to still be operational today but as a consequence embraces a variety of biological phenomena and structural features ( 5 ). Defining prions thus remains a nontrivial task. While a more restrictive definition is perhaps neither possible nor desirable, this general term induces some confusion and controversy. In an attempt to clarify discourse, at some point investigators in the mammalian disease-related field denied the fungal “infectious proteinaceous particles” the name of prions and proposed instead to term them “prionoids” ( 6 ). These semantic battles should not be disregarded as sterile, but rather should be taken as an indication of the variety of the biological realities that the term covers.

Mobile DNA, comprising both active and decaying copies of transposable elements (TEs), is present in nearly all living organisms. Although fungal genomes tend to be significantly smaller than the genomes of plants and animals, they still can vary dramatically with respect to their TE loads ( 1 ). While TEs have been proposed to provide some beneficial functions to their hosts, e.g., by promoting genetic diversity and accelerating adaptive evolution ( 2 – 5 ), their overall impact is considered deleterious ( 6 ). Insertional mutagenesis, gene misexpression, and genome instability represent some well-known examples of the deleterious effects associated with TEs. Importantly, by being able to move between vertical genetic lineages, TEs can still proliferate in a population of sexually reproducing individuals despite causing substantial fitness defects ( 6 ).

Fungi have developed a plethora of strategies to colonize plants, and these interactions result in a broad spectrum of outcomes ranging from beneficial interactions to death of the host. With respect to plant pathogens, fungi represent probably the most diverse group of ecologically and economically relevant threats. Fungal plant pathogen species are primarily in the phyla Ascomycota and Basidiomycota. Among ascomycetes, plant pathogens are in various classes such as the Dothideomycetes (e.g., Cladosporium spp.), Sordariomycetes (e.g., Magnaporthe spp.), or the Leotiomycetes (e.g., Botrytis spp.). Basidiomycetes are represented by the two largest plant pathogen groups: the rusts (Pucciniomycetes) and the smuts (spread among the subphylum of Ustilaginomycotina).

Mycorrhizal fungi are a heterogeneous group of diverse taxa associated with the roots of over 90% of all plant species, from liverworts to angiosperms. Although they can spend part of their life cycle in the rhizosphere, mycorrhizal fungi always associate with the roots of plants, including forest trees, wild grasses, and many crops, and colonize environments such as alpine and boreal zones, tropical forests, grasslands, and croplands. Both partners benefit from the relationship: mycorrhizal fungi improve the fitness of their host plants by influencing mineral nutrition and water absorption and by increasing tolerance to biotic and abiotic stresses. The host plant rewards the fungal symbiont with carbon compounds derived from the photosynthetic process ( 1 ).

Lichens were generally accepted as an independent group of organisms when Schwendener ( 1 ) discovered that they are the result of the association of fungi and algae. This insight was not widely accepted by contemporaries but initiated a scientific revolution as lichens later became the prime example of a mutualistic symbiosis and, indeed, the phenomenon for which the term symbiosis was originally introduced in biology as “symbiotismus” ( 2 , 3 ). The lichen symbiosis has proved to be one of the most important lifestyles in the Ascomycota and is also known from a few Basidiomycota. Approximately 20,000 currently known fungal species live as lichens, mostly in species-rich lineages of Ascomycota ( 4 ). The traditional view of lichens as a mutualistic symbiosis of a fungus and one or several green algae or cyanobacteria has always been debated ( 5 ), but it has recently been challenged more than ever by the discovery of numerous additional microorganisms that potentially occur as obligatory participants in the symbiosis.

The interactions between fungi and plants encompass a spectrum of ecologies ranging from saprotrophy (growth on dead plant material) through pathogenesis (growth of the fungus accompanied by disease on the plant) to symbiosis (growth of the fungus with growth enhancement of the plant). We consider pathogenesis in this article and the key roles played by a range of pathogen-encoded molecules that have collectively become known as effectors.

While fungi can make positive contributions to ecosystems and agro-ecosystems, for example, in mycorrhizal associations, they can also have devastating impacts as pathogens of plants and animals. In undisturbed ecosystems, most such negative interactions will be limited through the coevolution of fungi with their hosts. In this article, we explore what happens when pathogenic fungi spread beyond their natural ecological range and become invasive on naïve hosts in new ecosystems. We will see that such invasive pathogens have been problematic to humans and their domesticated plant and animal species throughout history, and we will discuss some of the most pressing fungal threats of today.

In its short history of a century and a half, of which the last half-century brought the most dramatic advances, scientific medicine has found ways to cure or to treat millions of ill people who 200 years ago would have died early. Paradoxically, these successes of modern medicine have given rise to large groups of people at risk for fungal infections. Life-saving treatments may now breach normal immune functions, or susceptible patients such as premature newborns now survive long enough to become infected by a fungus. Invasive fungal infections have been very rare over most of our species’ history ( 1 ), and the fungi that infect healthy humans are a small, if fascinating, group. Many more invasive fungal infections now occur in patients with an underlying serious illness.

Research efforts by mycologists over the past few years finally established the fungal community and its previously overlooked members as crucial components of the microbiome. It is now undeniable that the commensal fungal microorganisms, alongside the other components of the microbiota, play a central role in association with the human host. Ongoing research is describing the fungal community and is providing new insights into biological mechanisms by which multidirectional interactions between the microbiome, their genomes (metagenome), metabolites (metabolome), and the human host ultimately affect health and/or disease states.

Humans are exceptional among vertebrates in that their living tissue is directly exposed to the outside world. In the absence of protective scales, feathers, or fur, the skin has to be highly effective in defending the organism against a gamut of opportunistic fungi surrounding us. Most (sub)cutaneous infections enter the body by implantation through the skin barrier. On intact skin, two types of fungal expansion are noted: (A) colonization by commensals, i.e., growth enabled by conditions prevailing on the skin surface without degradation of tissue, and (B) infection by superficial pathogens that assimilate epidermal keratin and interact with the cellular immune system. In a response-damage framework ( 1 ), all fungi are potentially able to cause disease, as a balance between their natural predilection and the immune status of the host. For this reason we will not attribute a fixed ecological term to each species, but rather describe them as growing in a commensal state (A) or in a pathogenic state (B).

We focus this article on turning a biofilm inside out. The “inside” of the biofilm comprises the individual biofilm-related phenotypes, their environmental drivers and genetic determinants, and the coordination of gene functions through transcriptional regulators. Investigators have viewed the inside of the biofilm through diverse approaches, and this article will attempt to capture the essence of many. The ultimate goal is to connect the inside to the “outside,” which we view as biofilm structure, development, pharmacological attributes, and medical impact.

The immune system has over millennia evolved strategies to discriminate between what is “self” and what is foreign (nonself) as well as “normal self” and “injured or altered self,” with the ultimate purpose being to defend and repair self. First, to fully appreciate the framework that governs most of our current understanding of how the immune system operates, it is imperative to be cognizant of some of the immunological models that have laid the foundation on which we stand.

Disease and death caused by fungal infections have transitioned from a rare curiosity to a major global health problem. Because the vast majority of fungi capable of causing life-threatening infections target individuals with impaired immunity, recent decades have witnessed a stark rise in fungal infections due to medical interventions such as chemotherapy for cancer and immunosuppression for organ transplantation and due to the prevalence of HIV infection ( 1 ). Pathogenic fungi are the causative agents of billions of infections annually, with ∼1.5 million attributable mortalities ( 2 ). The public health burden is comparable to that observed with more notable infectious diseases such as tuberculosis and malaria, yet the impact of fungal infections on human health has been largely overlooked ( 3 ).

Estimates of the number of arthropod species vary between 1,170,000 and 10 million, accounting for over 80% of all known living animal species. One arthropod subgroup, insects, is the most species-rich member of all ecological guilds in land and freshwater environments ( 1 ). As arthropods were emerging as the dominant animals they are today, fungi were also colonizing the land. Over the past 400 million years fungi and insects have coevolved a wide array of intimate interactions ( 2 , 3 ). These interactions include mutualistic endosymbiosis ( 4 ); fungi as obligate food sources, such as those found in fungus-gardening ants ( 5 ); sexually and behaviorally transmitted parasites, such as Laboulbeniales ( 6 ); and the most common disease-causing agents of insects ( 7 ). Entomopathogenicity has evolved independently and repeatedly in all the major phyla of the Kingdom Fungi ( 3 ). The heterogeneity of entomopathogenic fungi probably derives from both they and their hosts having short generation times, i.e., rapidly driving new diversity with each generation, and from their occupation of a wide range of habitats, with near ubiquity in the soil and on plants. Interactions among fungi, hosts, and the environment are therefore diverse and dynamic, which complicates comparisons between different fungi infecting different insects since their interactions may be necessarily disparate. Historically, this quandary was dealt with by intensively studying the host pathogen interactions of a couple of experimentally tractable fungal species, and then extrapolating these results to distantly related species. Consequently, most of what we know about the biochemical and molecular basis of interactions between fungi and insects has been determined with the experimentally tractable hypocrealean ascomycete genera Metarhizium (family Clavicipitaceae) and Beauveria (family Cordycipitaceae). Metarhizium, in particular, has also emerged as an excellent model to explore a broad array of questions in ecology and evolution, host preference and host switching, and the mechanisms of speciation.

The classic work of Paul Buchner in the first half of the 20th century established the common occurrence of symbiotic associations between microbes and animals, often a fungus association hidden from view within an animal gut (see reference 1 ). Interest in the fungi associated with insects continues ( 2 – 9 ), and recently renewed interest comes from the molecular techniques available to study the interactions.

There are about 700 species of taxonomically diverse fungi that are be able to attack living nematodes (juveniles, adults, and eggs) and use them as a nutrient source ( 1 , 2 ). The most important genera include Purpureocillium, Pochonia, Hirsutella, Nematophthora, Arthrobotrys, Drechmeria, Fusarium, and Dactylellina ( 3 , 4 ). Among these nematophagous fungi, only a few species are obligate parasites of nematodes, but the majority are facultative saprophytes ( 5 – 7 ). Based on the mechanisms by which they attack nematodes, these nematophagous fungi are usually divided into four general groups: (i) nematode-trapping fungi that use specialized trapping structures differentiated from hyphae; (ii) endoparasitic fungi that use their spores; (iii) the opportunistic fungi that invade or colonize nematode eggs, females, or cysts with their hyphal tips; and (iv) toxin-producing fungi that immobilize nematodes before invasion ( 1 , 8 ). Nematophagous fungi are important natural enemies of nematodes in soil ecosystems. In recent decades, environmental and health concerns over the use of chemical nematicides have greatly increased the demand for the development of biological control agents in plant protection. The reason for the growing interest in nematophagous fungi is mostly their potential as biocontrol agents against plant- and animal-parasitic nematodes. So far, a substantial number of myconematocides have been developed worldwide ( 3 , 4 , 6 ).

The nematode Caenorhabditis elegans is a highly tractable organism that has been studied in the laboratory for several decades and has provided novel insights into questions of development, neurobiology, and cell biology, among other basic biological processes ( 1 ). Until recently, however, little was known about its natural ecology. Now, because of greatly increased sampling of C. elegans from the wild, we are learning more about its life outside the laboratory ( 2 ). In particular, it has become clear that C. elegans and other Caenorhabditis nematodes are commonly infected by microsporidia in the wild ( 3 – 5 ). Microsporidia comprise a phylum of over 1,400 species of fungal-related parasites that can infect nearly all animal hosts ( 6 – 8 ), so it is perhaps no surprise that these pathogens commonly infect the nematode C. elegans.

From the saprotrophs that decay plant material to the pathogens and mutualists that shape plant population dynamics at local and regional scales, fungi are major drivers of ecosystem health, plant productivity, and sustainability in all major biomes ( 1 – 5 ). The ecological roles of such fungi are driven by their own genomic and epigenetic architecture, as well as that of their hosts, often with strong influences from the environmental context in which such interactions occur ( 1 – 9 ).

Mycoparasitism is a lifestyle where a living fungus (host or prey) is parasitized by and acts as a nutrient source of another fungus (mycoparasite or predator). Mycoparasitic interactions are common among fungi and, similar to plant pathogens, comprise both biotrophic as well as necrotrophic relationships. Biotrophic mycoparasites obtain their nutrition from living fungal host hyphae, with which they can live in a balanced relationship for extended stages of their life cycles, and usually show a narrow host range ( 1 ). Because of their nature, however, only few species are studied in detail and applied as biocontrol agents. Examples are Ampelomyces quisqualis (Ascomycota, Pezizomycotina), a natural enemy of powdery mildews, whose genome sequence has recently been released, as well as haustorial biotrophs from the Zygomycota ( 2 , 3 ). In contrast to biotrophs, necrotrophic mycoparasites are more destructive and often rather unspecialized ( 4 ). They usually have wide host ranges, often comprising fungal plant pathogens, and kill their fungal prey by invasion and secretion of damaging molecules followed by feeding on the released nutrients ( 5 , 6 ). These traits render these fungi useful in agriculture for the biological control of plant diseases.

Fungi are now widely used in industrial biotechnology, for example, as production hosts for technical and food and feed processing enzymes, as gene donors for such enzymes, as production hosts for organic acids and cholesterol-lowering drugs (the statins), and as starter cultures and probiotics ( 1 ). Around half of the industrial enzymes used globally are of fungal origin; the other half are of bacterial origin. However, this balance is now moving toward the use of more enzymes from a wider spectrum of families of the fungal kingdom. There are several reasons for this. Fungal enzymes are efficient, compatible, and suitable for industrial processing: they have sufficient protein stability to give the enzyme products an acceptable shelf life; they provide customer solutions, meet regulatory approval demands, and fulfill end user needs.

Lignin is an aromatic polymer with a heterogeneous structure. Varied, nonhydrolyzable linkages between subunits present a significant constraint to efficient exploitation of plant lignin. The different types of biomass and pretreatment used in biomass conversion processes result in a large variety in the molecular weight, structures, modifications, amount, and aqueous solubility of the lignin in the remaining product side streams. While polysaccharides are deconstructed biologically mostly by glycoside hydrolase and esterase hydrolysis, the ether and carbon-carbon bonds of lignin are attacked by fungal oxidative enzymes and small-molecule oxidative species, adding heterogeneity to the products.

Since ancient times, humans have used fungi as food sources ( 1 , 2 ). The edible sexual structures of basidiomycetes and ascomycetes (e.g., truffles), the so-called mushrooms, are produced mostly in wood because many fungi are tree symbionts or decayers of tree tissues. These fruiting bodies represent a rich source of proteins, with low fat content and otherwise nutritionally quite poor. In some soils, they accumulate pollution (heavy metals and radioactivity) and should only be eaten in moderate quantities. Some mushroom species are considered delicacies (e.g., truffles, boletus, morels), but cultivation attempts have been unsuccessful, with a few exceptions (e.g., Morchella rufobrunnea). Only a few saprobic species can be industrially produced, such as Agaricus bisporus ( 3 ), Lentinus edodes (shiitake), and Pleurotus ostreatus, with production having mainly taken place in Asia for thousands of years ( 4 ). Some other fungi, while not really cultured, are inoculated on trees grown in appropriate natural habitats to increase the production of fruiting bodies, such as for shiitake and oyster mushrooms, with, however, sometimes unpredictable success, as is the case for truffles ( 5 ).

Fungi, plants, and bacteria are the major kingdoms of life with well-developed secondary metabolism. About 500,000 secondary metabolites (also referred to as natural products) have been described to date. About 100,000 of these are derived from animals, 350,000 are from plants, and 70,000 are from microbes ( 1 , 2 ). Roughly 33,500 bioactive microbial metabolites have been described ( 2 ). Of these 33,500 microbial metabolites, about 12.5% (4,200) are metabolites of unicellular bacteria and cyanobacteria, 41% (13,700) are products of Actinomycete fermentations, and about 47% (15,600) are of fungal origin ( 1 ). Furthermore, the rate of discovery of new fungal metabolites has accelerated significantly in the past two decades relative to the rate of discovery in the actinomycetes, filamentous bacteria that traditionally have been the richest source of microbial natural products ( 2 ). This complex and rich secondary metabolism is highly developed in the filamentous Ascomycota and Basidiomycota, while it is underdeveloped in the unicellular forms of the Ascomycota and Basidiomycota and in the Zygomycota, Blastocladiomycota, and Chytridiomycota ( Fig. 1 ). The diversity of fungal species, particularly in the Ascomycota and Basidiomycota, and the accompanying diversification of biosynthetic genes and gene clusters points to an almost limitless potential for metabolic variation. In fact, one can argue that much of the ecological success of the filamentous fungi in colonizing virtually all habitats on the planet is owed to their ability to deploy arrays of secondary metabolites in concert with their penetrative and absorptive life forms. This dependence of the fungi on secondary metabolites to conquer diverse habitats and sustain their existence within them is evidenced by the facts that most species make multiple types of secondary metabolites, their expression is orchestrated with the life cycle and environment, and significant portions of their genomes are devoted to encoding and regulating the production of these products.