Fungi Page 6
Sexual compatibility in the shaggy mane mushroom is controlled by a single gene or mating type locus. This is known as a bipolar or unifactorial incompatibility system. Complexity is introduced to this reproductive mechanism by the presence of different versions, or alleles, of the mating type locus (A1, A2, A3, etc.). When mycelia with the same mating type allele make contact they are incompatible (e.g. A1 × A1). But whenever a shaggy mane mycelium meets any mycelium with one of the other mating type alleles it has the potential to form a heterokaryon (e.g. A1 × A3 = A1A3). This favours outbreeding and maximizes the probability of sexual reproduction when colonies encounter one another. The more complicated tetrapolar incompatibility system relies on a pair of genes that control mating. A successful cross between two homokaryons, A1B1 × A2B2, produces a heterokaryon carrying both kinds of nucleus: A1A2/B1B2. Many different alleles of both genes have evolved in some mushrooms, so that the number of possible combinations of homokaryotic mycelia skyrockets. In another ink cap mushroom, Coprinopsis cinerea, there are tens of thousands of different mating types based on this tetrapolar system (e.g. A1B1, A1B2, A1B9, A2B7, and so on). This reproductive system seems mind-boggling when compared with the binary approach employed by our species. It makes sense for mushrooms whose survival relies on chance engagements with a colony of the same species growing in the same patch of soil. Coprinopsis cinerea is a model for mushroom research. Its genome encodes around 13,000 genes. The genomes of many other mushrooms are considerably larger.
16. The life cycle of the mushroom Coprinus comatus.
Once a heterokaryon forms, mushrooms develop according to an internal clock if soil moisture, temperature, and other environmental conditions are permissive. Mushroom development is a mysterious process. Mushroom formation begins with the growth of a knot of hyphae that expands into a rounded primordium. If the primordium is sliced from top to bottom an embryonic fruit body is often evident with miniature cap, gills, and stem. In the poisonous death cap, Amanita phalloides, the primordium develops into an egg-shaped structure in which the mushroom is surrounded by a protective layer of hyphae called the universal veil. As the mushroom expands, the universal veil is stretched and broken and the lower part of it persists as a cup called the volva that surrounds the bulbous base of the stem (Figure 17). Fragmentation of the upper part of the universal veil leaves white spots on the red cap of the fly agaric, Amanita muscaria, and scales on the caps of many other mushrooms. A second interior sheet of hyphae, called the partial veil, protects the gills in the primordium. Breakage of this membrane during cap expansion leaves a ring around the top of the stem in the death cap and the fly agaric.
The overnight appearance of mushrooms is one of the phenomena that have encouraged so much superstition about fungi. It is explained by the unusual way that fruiting occurs. Rather than relying on an increase in the number of cells in the fruit body, most of the rapid increase in the volume of mushroom is caused by the absorption of water and elongation of the millions of hyphae that form the stem and the cap. This hydraulic mechanism allows the fruit body to exert a pressure as it thrusts up through soil or rotting wood. In the urban environment, this exertion of force explains how clusters of mushrooms push their way through crevices in concrete and are even capable of rupturing a layer of asphalt.
17. Mushroom expansion in Amanita phalloides, the death cap.
While experiments have allowed mycologists to solve these mechanical aspects of mushroom development, we have few clues about the genetic controls of fruit body development. We do not know how the genome of the death cap always produces mushrooms with greenish or yellowish caps, evenly spaced white gills, and solid stems with a ring and a volva. Questions about the sensitivity of mushrooms to gravity are unanswered. The gravitropic growth of mushrooms aligns the cap horizontally and the gills vertically. This is essential to ensure that discharged spores fall between gills and escape into the air rather than becoming stuck on a gill surface. The way that mushrooms sense gravity is a mystery.
There is so much to learn about mushrooms, so many prospects for young investigators. The mushroom life cycle has been studied for more than one hundred years and the genetic control of mating remains an active research topic. As a young doctoral student, Elsie Wakefield (1886–1972) performed crosses between different mushroom strains to explore the genetics of the mating process. She went on to become Head of Mycology at Kew in London and is one of many women scientists who have dedicated their careers to mycology. Mycology is one of the branches of biology that has always attracted women and it is important to note Elsie Wakefield’s work to balance the overwhelming influence of Arthur Henry Reginald Buller (1874–1944), who is recognized as ‘The Einstein of Mycology’.
Buller was born in Britain and spent his career at the University of Manitoba in Canada. His seven-volume work, Researches on Fungi, is the foundation of the modern study of mycology. These books describe ingenious experiments on many aspects of mushroom biology and some of them exposed his eccentric personality. When he worked on bioluminescence, for example, he strapped leather horse ‘blinders’ to his head to avoid exposing his eyesight to bright street lamps. This strategy allowed him to preserve his sensitivity to the dim light thrown by mushrooms in his laboratory. He also made significant contributions to fungal genetics including a prediction, which was validated after his death, that outcrossing in mushrooms could occur through the transmission of nuclei from a heterokaryon into a homokaryon. This esoteric point is an important feature of mushroom biology and has practical applications in mushroom cultivation.
Buller worked on many kinds of fungi, including rusts that infect cereal crops. Rusts are examples of basidiomycete fungi that do not form mushrooms. They have the largest genomes of any fungi, with one species packing more than seventy times more raw sequence than yeast and encoding over 25,000 genes. For further comparison, there are an estimated 20,000 to 25,000 genes in the human genome. The reproductive mechanisms of the rusts will be discussed in Chapter 5 when we consider fungi that cause plant diseases.
Genetic research on the arbuscular mycorrhizal fungi (Glomeromycota), zygomycetes, and aquatic fungi that produce swimming zoospores is very limited in comparison with the investment made in understanding the reproductive biology of ascomycetes and basidiomycetes. Part of the explanation for this research bias lies in the difficulty of working with these fungi in culture. Fungi in the Glomeromycota that form arbuscular mycorrhizas with plants do not grow on their own in culture at all, but they can be grown in association with cultured roots. Asexual reproduction in these fungi is accomplished by the formation of huge spores. Some have a diameter of almost 1 mm and are filled with hundreds or thousands of nuclei. They are formed in clusters called sporocarps in some species. The genome of Rhizophagus irregularis, a fungus that forms arbuscular mycorrhizas with poplar trees, is even larger than the truffle genome and encodes 28,000 genes. Like the truffle genome, the chromosomes of Rhizophagus incorporate multiple copies of genes. It is possible that the entire genome of this fungus has been duplicated during its evolution. A lot of the protein-encoding genes seem to be involved in the communication between the fungus and its plant host. Sexual reproduction has not been observed in arbuscular mycorrhizal fungi, but it may occur in nature. Evidence for this is found through annotation of the Rhizophagus genome, which reveals that the fungus has genes that control meiosis and genes that specify different mating types.
Zygomycetes produce sexual spores called zygospores. Zygospores resist germination in culture, which makes it very difficult to assess the outcome of crosses between different strains. Sexual reproduction in these fungi has been studied in detail, however, and is often used as an illustration of a simple life cycle in biology textbooks (Figure 18). Mycelia of these fungi, including species of Mucor and Rhizopus (not to be confused with Rhizophagus), are not separated into compartments by septa. Their nuclei are distributed throughout their branching hyphae. The sequenced genomes of species of Mucor and R
hizopus are comparable in size to the genomes of mushrooms.
Zygomycete colonies of compatible mating types use volatile pheromone molecules to detect one another before they make physical contact. When hyphae sense these chemicals they make contact and stick together tip-to-tip. After attachment, each cell produces a septum behind its tip isolating a compartment that contains multiple nuclei. The two cells merge when the barrier in the contact region dissolves, and a thick wall forms around the blended cytoplasm to produce a zygospore. Nuclei from the two mating types fuse inside this spore and then undergo meiosis to produce a new population of haploid nuclei. These are released in airborne spores when the zygospore germinates and the spores produce a new generation of feeding mycelia. Spirodactylon, whose coiled spore-producing stalks were described in Chapter 2, is another zygomycete. Zygospores of this fungus have smooth walls rather than the warty surfaces shown in Figure 18.
18. Mature zygospores formed between compatible strains of zygomycete fungi.
Aquatic fungi
Asexual reproduction in aquatic fungi is accomplished by the formation of zoospores. Like the conidia of ascomycetes, zoospores carry clonal copies of the genome of single parents. Unlike conidia, which are blown around by air currents and dispersed by water, zoospores are capable of swimming towards new sources of food using chemical cues in the environment. The pathogenic chytrid Batrachochytrium dendrobatidis uses zoospores to locate and infect its amphibian hosts. Its genome includes almost 9,000 protein-encoding genes, which is considerably more than yeast. Sexual reproduction has not been described in this fungus and there is no trace of genes that might control mating type in its genome.
Sexual cycles do occur in other groups of zoosporic fungi. Allomyces macrogynus is a species classified in the Blastocladiomycota that produces two types of swimming sex cells or gametes that resemble its asexual zoospores. One of the gametes is much larger than the other and releases a pheromone called sirenin, named after the sirens that tried to lure Odysseus and his shipmates to their doom in Homer’s Odyssey. Sirenin attracts the smaller type of gamete and fusion occurs when they make contact. After the development of a diploid stage in Allomyces, meiosis restores the haploid number of chromosomes in the following generation. The same shift from haploid to diploid, and diploid to haploid, occurs in all of the sexual life cycles described in this chapter.
It is interesting to compare fungal and animal life cycles. In every version of sexual reproduction in fungi and animals, the merger of two cells, or gametes, and fusion of their nuclei doubles the number of chromosomes, and this fertilization event is followed by meiosis that halves the number of chromosomes. The difference between the reproductive cycles of fungi and animals lies in the way that the gametes are produced. Adult fungal cells have a single set of chromosomes; they are haploid and can function directly as gametes. Adult animal cells have two sets of chromosomes; they are diploid and gametes are produced by meiosis. The reason that sexual reproduction occurs in either group of organisms is that it allows for the reshuffling of the genome during fertilization and meiosis. This spreads different versions of genes (alleles) and different combinations of genes across populations, which is the basis for the origin ofnew species.
Chapter 4
Fungal mutualisms
Symbiosis
Symbiosis is a catch-all term that describes close biological interactions between two or more species. It embraces damaging relationships as well as life-sustaining coalitions. A bracket mushroom protruding from the trunk of a dead tree may be the last stage in a parasitic symbiosis. The fungus was supported by this symbiosis at the expense of the infected tree. Relationships from which both the fungus and its partner benefit are called mutualisms. Commensalism lies between these extremes, referring to situations in which one player is advantaged without having any discernible negative or positive effects on the other (Figure 19). Trichomycetes are fungi that live inside the guts of invertebrates and benefit from the stable environment and nutrients available in this habitat. They are commensals that do not seem to impact the health of their hosts. This chapter is devoted to mutualisms.
Mutualisms with insects
Scale insects engage in curious relationships with fungi that trap them beneath mats of tissue made from interlaced hyphae (Figure 20). Female scale insects are limbless plant dwellers that clamp themselves to plants. They feed through slender sucking tubes that penetrate the plant and probe its vascular tissues containing sugary sap. The arrangement between the insect and the plant is like the connection between an oil rig, its drill, and a deep well. The scale insect is a parasite that enjoys all of the nutritional benefit at the plant’s expense. Fungi that specialize in relationships with scale insects are basidiomycetes classified in a family called the Septobasidiaceae. There is no common name for this group. The fungus grows around the body of the insect, penetrates its exoskeleton, and feeds on the animal’s soft tissues. The fungus seems to be operating as a parasite that lives on a parasite. But rather than harming the insect, the fungus hides its captive from predators in the early stages of the association, allowing it to feed in safety. For this reason, the interaction between the fungus and scale insect is regarded as a mutualism. Rust fungi that cause epidemic plant diseases are close relatives of the Septobasidiaceae. There is no equivocation in identifying rusts as parasites, because their growth does not help the host plant at all.
19. Continuum of relationships between different species. Symbiosis is the general term for all of these interactions.
Ambrosia beetles cultivate fungi in galleries excavated in the wood of trees and shrubs. Mycelia of these ambrosia fungi grow on the walls of the galleries, digesting cellulose and other components of the wood. The beetles enjoy an exclusively fungal diet by consuming cells from the surface of the mycelium. Growth of the ambrosia fungi in the galleries is not left to chance. The beetles carry cells of the fungi in special pockets in their exoskeletons called mycangia. The mycangia of some beetles are protected by a rim of hairs that brushes fungal spores from the walls of rotting galleries into the pockets. Once inside the mycangia the fungi grow as yeasts rather than filamentous hyphae. These yeasts are nourished by secretions into the mycangia until they are deposited in fresh galleries and begin the process of wood decay. Ambrosia beetles favour damaged and dying trees, but some species attack healthy trees and cause considerable economic damage. The Asian ambrosia beetle is an invasive insect that was introduced to North America in the 1970s. It attacks 200 species of trees, ornamental shrubs, and vines.
20. Scale insect trapped on a leaf surface within a blanket formed by the fungus Septobasidium fumigatum. The insect feeds from the plant through its long sucking tube and the fungus feeds from the insect.
Fungus-stealing or ‘mycocleptic’ ambrosia beetles exploit the work of the ‘normal’ ambrosia beetles, by tunnelling into wood adjacent to existing galleries. Ambrosia fungi spread from the first set of galleries into the passages made by the mycocleptic beetles and provide them with food. This process reduces the quantity of wood available to the beetles that invaded the trees in the first place. The mycocleptic beetles still excavate their own galleries, but the benefit of dispensing with the active cultivation part of the symbiosis is evident from the fact that some of them no longer produce mycangia. Dispensing with the formation of these organs and their secretions must save some energy for the thieves.
A similarly complex symbiosis has evolved between Sirex woodwasps and the fungus Amylostereum areolatum. Female woodwasps drill holes into the sapwood of pine trees, lay one egg at the bottom of each hole, and add spores of the fungus on top. Like the ambrosia beetles, Sirex woodwasps have special pockets for carrying fungal spores. These mycangia are close to the insect’s ovipositor, stationing them for injection into the wood along with the eggs. The wasps also release a dose of toxin carried in mucous that weakens the defence response by the tree. The spores are embedded in wax when they are expressed from the mycangia. This is dissolved
by the wasp’s mucous, allowing the fungus to commence growth on the sapwood. When the eggs hatch, the larvae feed on the fungal mycelium. Amylostereum is a basidiomycete related to species of Russula that produce umbrella-shaped mushrooms. It forms a crusty fruit body on the surface of decaying wood and benefits from its mutualism by its injection through the tough tree bark by the wasp.
Symbioses between fungi and scale insects, ambrosia beetles, and woodwasps are highly developed relationships that have involved substantial structural, biochemical, and behavioural adaptations in the participating organisms. Mutualisms between fungi and social insects reach the height of complexity and are described as a form of mushroom farming. Leaf-cutter ants cultivate mushrooms in underground nests in South America and North America. Millions of insects can occupy the largest nests and support hundreds of fungus gardens that resemble honeycombs. Sterile female workers move between gardens through tunnels and feed the fungi with a pulp made from leaf fragments. Forty ant species engage in this symbiosis across the Americas with a single mushroom species called Leucoagaricus gongylophorus. The mycelium of this mushroom digests the leaf pulp using enzymes that convert cellulose and starch into sugars, and break down proteins into amino acids. The ants feed on buds produced on the surface of the mycelium. By depositing faecal fluid containing active enzymes obtained from the buds, the ants jump-start the composting of new leaf material before it becomes colonized by new fungal growth.