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  Powdery mildews are biotrophs like rusts and cannot be grown in culture. Compared with the enormous genomes of rusts (described in Chapter 3), Blumeria graminis functions with far fewer protein-encoding genes. The reason for this difference is not clear, but the loss of genes is characteristic of many parasites that establish obligatory relationships with single hosts. One reason that rust genomes are so big is that these fungi need to overcome the defences of two unrelated plant species. The formation of so many types of spore is another characteristic that may be related to genome expansion.

  The chains of spores produced by powdery mildews are asexual conidia. Mycelia of compatible mildew strains associate on the surface of leaves and produce sexual ascospores inside pinhead-sized fruit bodies. The fruit bodies are decorated with long hairs that bend as they dry. In some powdery mildew, these appendages act as stilts that lever the fruit body above the tangled mycelium on the leaf surface. This allows the sexual stage to fall free from the leaf and overwinter on surrounding twigs. In the spring, the fruit bodies crack open along a line of weakness, exposing the asci within. Explosive discharge of the asci blasts the ascospores into the air where they are dispersed by wind.

  Mildew ascospores are infectious and can establish a new round of disease in a crop. In grape powdery mildew, for example, the fruit bodies develop at the end of the summer in wine-growing regions and remain dormant during the winter. Ascospores are released when the fruit bodies open in the spring, and these initiate infections when they land on young leaves. As the disease develops, the fungus colonizes shoots and ripening berries and its asexual conidia spread the mildew throughout the vineyard. Powdery mildews are among the enchanting illustrations published by Charles and Louis-René Tulasne in 1861 (Figure 23). The Tulasne brothers discovered the way that single species of ascomycetes rotate between the formation of asexual and sexual spores (Chapter 2).

  Powdery mildew is the most damaging disease of grapes, but other ascomycetes produce black rot, bitter rot, and ripe rot in vineyards. Adding to the challenges posed by multiple pathogens, a single fungus is capable of producing several diseases depending on the timing of the infection. Botrytis cinerea is an ascomycete that grows as a necrotroph and produces bunch rot when it develops on fruit damaged by insects or high winds, and grey rot in wet weather. The same fungus also causes noble rot when its growth is stimulated by rainfall, followed by weeks of hot, dry weather. Noble rot results in the drying or partial raisining of the grapes, which concentrates their flavour and sweetness. Grapes with noble rot are picked individually to produce expensive dessert wines like Sauternes from Bordeaux.

  23. The powdery mildew fungus Microsphaera penicillata, illustrated by Charles Tulasne.

  In rice blast disease, caused by the ascomycete Magnaporthe grisea, conidia stick to the leaf surface, germinate to produce a short hypha, and form a swelling at the end that serves as a pad from which the infection proceeds. This pad is called an appressorium (Figure 24). It is a rounded cell that fastens itself to the rice leaf with an adhesive O-ring and blackens itself with melanin pigment. The pigment prevents leakage of molecules from the cell as it swells and becomes pressurized. This pressure comes from the absorption of water and rises to ten times the level that inflates a bicycle tyre. Once pressurized, the fungus produces a thin peg on the underside of the cell that pierces the plant surface. In some diseases, fungi release enzymes to weaken the plant surface to allow easier access. The rice blast fungus is sufficiently strong to work by pressure alone. It deforms metal films in laboratory tests and pushes holes through plastics including the bulletproof vest material called Kevlar.

  24. Appressorium of the rice blast fungus, Magnaporthe grisea.

  Rice blast can be controlled with fungicides, but recent epidemics have ruined millions of hectares of crops. The devastating effects of rice blast fungus have led to concerns that it might be used as a biological warfare agent. Before the United States halted research on bioweapons during the Nixon administration, field trials were conducted in the 1960s to gauge the potential impact of the fungus on rice production in China and SouthEast Asia.

  Rice is attacked by many other ascomycetes at the seedling stage and later in plant development. Infection by Fusarium fujikuroi causes plants to elongate, topple over, and die without producing any grain. These unusual disease symptoms are induced by the production of a growth hormone called gibberellic acid. Plants synthesize this hormone to stimulate seed germination and control their growth rate, but the production of an excess of the compound by the fungus has disastrous effects on rice. One thousand species of Fusarium have been described and many of them are plant pathogens. Banana growers are very concerned by a virulent strain of Fusarium oxysporum called Tropical Race 4. The global banana crop is exceedingly vulnerable because the Cavendish cultivar, which has served as the commercial banana since the 1960s, is a clone derived from a single plant grown in Asia. The previous cultivar of banana was wiped out by Fusarium wilt, also known as Panama disease, in the 1950s, and Cavendish has worked well since then. But with the emergence of the TR4 strain of the fungus the entire crop is threatened again.

  Rusts, smuts, and ascomycetes are responsible for most fungal diseases of plants. A handful of zygomycetes cause soft rots of flowers, fruits, and bulbs. The zoosporic fungus Olpidium brassicae, described in Chapter 2, infects cabbage roots, but, like zygomycetes, these basal groups of fungi are more important in decomposition than plant disease. It is worth revisiting the fact that many plant diseases are due to microorganisms that look like fungi but are not fungi. For example, the potato blight pathogen, Phytophthora infestans, is an oomycete water mould (Chapter 1). Pathogens called downy mildews are also oomycetes. Grapes are infected by the ascomycete powdery mildew Uncinula necator, and the oomycete downy mildew Plasmopara viticola. A practical consequence of this taxonomic nuance is that some fungicides that affect powdery mildews do not work against downy mildews.

  Fungicides

  Bordeaux mixture, which contains copper sulphate and slaked lime (calcium hydroxide), was developed as a preventative spray against grape diseases in the 1880s. The effectiveness of copper as a fungicide was probably recognized much earlier than this and it is still used today. Bordeaux mixture prevents spore germination on the leaf surface and can also be applied to soil to control root disease. Fungicides range from blends of simple compounds containing copper and sulphur, to generations of synthetic compounds that disrupt specific metabolic pathways in the fungi. Contact fungicides are active on plant surfaces and systemic fungicides are absorbed by the plant and transmitted throughout its tissues. Mancozeb is used as a protectant in the same way as Bordeaux mixture. This broad-spectrum fungicide interferes with lipid synthesis and energy production in fungi. Systemic fungicides include carboxin that inhibits protein synthesis, and triadimefon that is applied as a seed treatment and damages the cell membranes of fungi.

  The development of resistance to fungicides has affected the usefulness of many compounds within a few years of their introduction. Benomyl, which disrupts the protein skeleton inside fungal cells, was used against fungal diseases from the late 1960s until its withdrawal from the market in 2001. It was a very effective treatment for powdery mildews and a wide range of other diseases, but resistance spread very swiftly and there were concerns that human exposure to benomyl was associated with birth defects. The limited ‘shelf life’ of fungicides, coupled with their expense and potential toxicity towards humans and other animals, explains why there is so much investment in breeding programmes to exploit natural defence mechanisms of plants against fungi.

  Chapter 6

  Fungi and decomposition

  Fungi and the carbon cycle

  Most people associate fungi with rotting fruit and decaying wood, and, more generally, regard their presence as a symptom of death. This is not unreasonable. Fungi that do not form supportive or parasitic relationships with plants and animals feed on the debris of life. These saprotrophs decompose de
ad roots, leaves, flowers, fruits, seeds, twigs, branches, upright tree trunks, and fallen logs. They rot animal faeces and the tissues of invertebrates and vertebrates. Some of these decomposers produce mushrooms and these become a source of nutrients for other organisms. Fungi also clear up our mess, breaking down every natural product used in a lifetime of consumerism and destroying synthetic materials made by industry. Fungi spin the carbon cycle.

  Coprophilous fungi

  Cows and other ruminants have a complex digestive system that is adapted for processing fibrous plant materials. After chewing and rumination (rechewing), forage is digested by a combination of enzymes secreted by the cow and by microorganisms inside the animal’s stomach compartments and its gut. Anaerobic fungi participate in cellulose breakdown in the rumen. When the dung is dropped on the ground, the immediate availability of oxygen stimulates the growth of ‘coprophilous’ fungi that permeate the dung and absorb nutrients from the residue of plant materials.

  Cow dung does not seem like a promising place to look for beautiful fungi, but inspection of its malodorous surface with a hand lens reveals some stunning organisms. After deposition, the dung of grass-fed animals becomes covered with Pilobolus stalks that sparkle with fluid droplets and resemble cut crystal glassware. These pressurized structures, described in Chapter 1, function as squirt guns that blast their spore-filled capsules away from the dung. The cup fungus Ascobolus stercorarius is another brief occupant. The Latin name of this species means ‘of or relating to dung’. This fungus fashions five-millimetre-wide yellow discs that support a layer of thousands of explosive asci. Each disc is a fruit body covered with conspicuous dots and each dot is a single ascus coloured by its violet ascospores. When spores are released by a group of asci in one part of the disc, the disturbance causes nearby asci to detonate and these mini-explosions surge from one side of the little fruit body to the other. Discharge of spores from individual asci happens too quickly to be visible without recording the mechanism with a high-speed camera running at more than 100,000 frames per second. But the salvo of hundreds of exploding asci is apparent as a wave of disappearing dots. The resulting puff of ascospores is visible as smoke rising from the fruit body.

  Tiny gilled mushrooms also grow from the dung. These include the ink cap species Coprinopsis cinerea, and a mottlegill called Panaeolus semiovatus. Some hallucinogenic mushrooms are associated with animal dung. The liberty cap, Psilocybe semilanceata, grows in pastures fertilized by sheep, horses, and cows, but does not fruit directly on the dung.

  Rather than reproducing at the same time, different fungi produce their spores according to a programme that plays out over several days. Zygomycetes, including Pilobolus, are the first fungi to emerge from cow dung, followed by cup fungi and other ascomycetes, and mushrooms appear last. This ecological succession illustrates the complexity of the decomposition process. The spores of Pilobolus pass through the digestive system of an animal before they are deposited in its fresh dung. They germinate immediately, forming an extensive mycelium within the dung that absorbs relatively simple nutrients before the development of spores. The cup fungi may also pass through a herbivore and begin growth immediately, but they invest more time in breaking down more complicated molecules in the dung before fruiting. Mushroom colonies take the longest to digest the most resilient polymers in the dung and generate enough mass to support the development of their gilled fruit bodies.

  Sporormiella species grow on dung from many different animals. Historical accumulations of the spores of this ascomycete in ancient sediments have been linked to changes in the abundance of herbivores. Palynological records from Queensland, Australia, show an abrupt drop in the deposition of Sporormiella spores 41,000 years ago. The regional climate was stable at this time and human hunters are identified as the cause of the collapse of animal populations and consequent disappearance of their dung and the fungus. A similarly precipitous decline in spore numbers in sediment samples from lakes and wetlands in North America is found towards the end of the Pleistocene (beginning 15,000 years ago). This was caused by the eradication of the woolly mammoth, mastodon, and rhinoceros by Palaeo-Indians at the end of the last ice age. Changes in the concentrations of Sporormiella ascospores have also been linked to the disappearance of Madagascan megaherbivores 1,700 years ago and the 17th-century extinction of giant flightless birds called moa in New Zealand.

  Some of the coprophilous fungi whose dispersal mechanisms are geared towards passage through animal guts are also found on plant products that have been processed for gardening including wood chips and wood mulch. They are opportunists that have taken advantage of these refined sources of cellulose. Woody garden supplies also attract a few of the saprotrophic fungi that decompose the tissues of dead and dying plants. Plant decomposition involves a succession of fungi and is affected by moisture levels, temperature, and other environmental variables. Broad patterns of decomposition can be determined from the microscopic study of fungi growing on rotting vegetation. Species that form their spores on the plant surface can be identified directly and other fungi can be isolated from the decomposing tissues and grown in culture. The usefulness of these experimental approaches is limited, however, because they tend to exaggerate the importance of fungi with conspicuous fruit bodies and species that grow well in culture. More recent research in which fungi are identified from DNA extracted from leaves and other samples of rotting vegetation have revealed a more detailed picture of the decomposition process.

  Leaf decay

  Leaf decomposition is one of the processes examined using molecular methods. Deciduous leaves are primed for breakdown by endophytes that grow inside them before leaf fall as well as by fungi that populate the leaf surface. These internal and external leaf residents are yeasts and species of filamentous ascomycetes. They do not damage the leaves until they are shed, but begin consuming sugar molecules and other simple nutrients as soon as the host defences are shut down. They provide another example of the changes in fungal behaviour that can accompany an alteration in the growth conditions offered by plants (Chapter 4). After this early surge in fungal activity, decomposition proceeds more slowly as different ascomycetes colonize the leaves and digest cellulose and other complex polysaccharides. Basidiomycetes tend to be latecomers in the succession of fungal communities on decaying leaves. They are the most effective in destroying lignin, which is the complex organic molecule that strengthens plant cell walls.

  Lignin is linked to cellulose and other compounds in leaves and these polysaccharides are exposed for decomposition once the lignin is dissolved. This process may uncover cellulose for ascomycetes that cannot remove lignin for themselves, allowing them to grow in the leaf litter along with the basidiomycetes. Metagenomic methods have revealed the DNA sequences of hundreds of different fungi involved in the breakdown of leaves of single species. DNA sequencing methods can also be used to examine the variety of fungal genes encoding enzymes in the leaf samples. This is useful because variations among these genes can be linked to specific groups of fungi, providing a snapshot of the fungi involved in the breakdown of different leaf components.

  The remarkable complexity of the decay process is evident from DNA sequences amplified from decomposing conifer needles. Breakdown of these stiff leaves can take several years and deep layers of needles accumulate in evergreen forests. Endophytes act as pioneer decomposers in the first year. These fungi give way to new communities of saprotrophs that arrive and digest the more resistant components of the needles. After four or five years the shape of individual leaves is no longer recognizable in the deeper layers of the leaf litter and humus is produced at the end of the decomposition process. Humus resists further breakdown for hundreds of years and is an essential component of soil structure that controls moisture content and nutrient retention.

  Endophytes are also crucial in the decomposition of leaves that fall into freshwater streams. Some of these fungi are ascomycetes called ‘Ingoldians’, after C. T. Ingold, who discovere
d them in the 1930s. Ingold was an influential British mycologist whose studies on fungi spanned more than seventy years. His fungi produce exquisite spores shaped as stars, crescents, sigmoids, commas, and miniature cloves (Figure 25). These develop on the surface of leaves, float downstream, and attach to submerged leaf fragments and twigs. Ingoldians, along with other kinds of fungi, bacteria, and invertebrates break down leaves very swiftly in unpolluted streams. The extended shapes of the Ingoldian spores favour their attachment to leaves and they also become concentrated in foam bubbles that accumulate around rocks and fallen logs. Spores trapped in the foam may become airborne as the bubbles collapse. This would explain how these aquatic fungi establish themselves as endophytes in plants growing above the water.

  25. Aquatic spores produced by Ingoldian fungi.

  Wood decomposition

  Fungal decomposition of wood is more noticeable than the activities of microscopic fungi on herbivore dung and leaves. Logs in forests become decorated with the brackets of basidiomycetes that digest the masses of cellulose and lignin in the rotting heartwood. At the same time, the remaining bark becomes covered with black blobs and solid crusts of saprotrophic ascomycetes. The biochemistry of wood decay involves many different enzymes that act on cellulose, other polysaccharides called hemicelluloses, and lignin. This mixture of polymers that form the dry matter of plants is called lignocellulose. Lignin is a huge, branching molecule of aromatic rings linked into a rigid three-dimensional framework that is very resistant to breakdown. Its decomposition is carried out by fungi that secrete enzymes called peroxidases and laccases that weaken different parts of the structure of lignin. White rot fungi use these enzymes to oxidize the lignin in fallen trees and then feed on the cellulose exposed by removal of the lignin. The loss of lignin accounts for the bleaching of the rotten wood. White rot fungi include Phanerochaete chrysosporium, whose fruit bodies grow as thin crusts on the wood surface, and Trametes versicolor, known as ‘turkey tail’ for the concentric patterns of coloured stripes on its little brackets. The cultivated shiitake mushroom, Lentinula edodes, is another white rot fungus. Its capacity for growth on hardwood logs is the basis for a global industry with an annual production of more than two million tons of the flavourful mushroom.