16 Deuteromycota: The Imperfect Fungi Richard E. Baird CHAPTER 16 CONCEPTS • Taxonomy of the Deuteromycota is based on asexual spore formation or no spores produced. • Sexual stages are primarily the in Ascomycetes, but there are a few in the Basidiomycetes. • Species can be parasitic or saprophytic. • Asexual spores called conidia are nonmotile. • Conidia are formed on conidiophores either singly or grouped in sporodochia, pycnidia, acervuli, or synemmata.
Species of the Deuteromycota, also known as the imperfect fungi, are among the most economically destructive group of fungi. These fungi cause leaf, stem, root, fruit, and seed rots; blights; and other diseases. The Southern corn leaf blight epidemic in the 1970s, which caused a damage of one billion dollars, was incited by Helminthosporium maydis, the anamorph or the asexual form of the ascomycete Cochliobolus heterostrophus. Other deuteromycetes, such as Aspergillus flavus, produce mycotoxins (aflatoxins) in infected corn kernels. Mycotoxins when ingested by humans or animals can cause cancer of the digestive tract or other serious illnesses or death. The deuteromycetes were called imperfect fungi in the early literature because they were thought not to produce sexual spores like those by species of the Ascomycota (Chapter 13 and Chapter 15) and Basidiomycota (Chapter 19). Descriptions and classifications of these fungi were based solely on production of conidia or on mycelial characteristics, or both. Deuteromycetes are now known to be the anamorphic stage of members of the Ascomycota and Basidiomycota. For example, Fusarium graminearum is the imperfect (asexual) stage of Gibberella zeae. The ubiquitous pathogen Rhizoctonia solani, which does not produce asexual spores, is the anamorph of the basidiomycete Thanatephorus cucumeris.
DEUTEROMYCOTA OR FUNGI IMPERFECTI A brief history of this group may be helpful in understanding why a fungus may be known by two scientific names. During the 1800s, fungal identification was based strictly on morphological characters. The object of these studies was to identify pathogenic fungi when very little was known about their anamorph–teleomorph (sexual spore) 0-8493-1037-7/03/$0.00+$1.50 © 2003 by CRC Press LLC
relationship. The asexual fruiting bodies were often the only structures present on infected host tissues, and scientists were unaware that sexual reproductive states existed. Early mycologists, such as Persoon (1801), Link (1809), and Fries (1821), described genera and species of imperfect (lacking a known sexual stage) fungi that were later classified as Fungi Imperfecti. These studies initiated the description of many deuteromycetes and other fungi and are considered the starting point for fungal classification. Saccardo (1899) compiled descriptions of the known fungi into one unified source in his Sylloge Fungorum series. Taxonomic keys and descriptions to the genera and species of the Deuteromycota by using spore shape, size, presence of cross-walls (septa) in the hyphae, and fruiting body type were provided by numerous scientists over the next half century. Conidiospore development (ontogeny) was used as a basis to identify genera and species after the 1950s. Asexual spore development on conidiophores and within fruiting bodies was considered a more natural classification for these fungi. Although considered a more natural classification scheme than those previously based on spore and conidiophore morphology, the earlier systems continue to be used by plant pathologists and disease diagnosticians because of the ease in identifying the genera and species. Since these early works, hundreds of papers and monographs were developed that include additional genera and species with information on their associated sexual stages. Many of the imperfect fungi do not readily form a sexual stage in culture or on host tissue. Therefore, artificial systems for identification were developed and are currently being used. Recognizing that the majority of deuteromycetous fungi with a teleomorphic stage also have a second name for the sexual stage may be critical in understanding how to identify these fungi. 133
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LIFE HISTORY The deuteromycetes are primarily terrestrial in distribution, but can occur in salt or fresh water. They survive by deriving nutrients as saprophytes on plant debris or as parasites on living hosts. The degree of parasitism or pathogenicity varies depending on the fungus, the isolate, and host they invade. Many species of deuteromycetes are not only parasitic on plants but also infect animal cells. Deuteromycetes may cause damage directly by infecting a host or indirectly by producing toxins. Direct infections by deuteromycetes to living hosts, other than plants, are termed mycoses and approximately 15 types are known. The more common mycoses include candidiosis (Candida albicans), and superficial infections called dermatophytosis, which include athlete’s foot or dandruff caused by several Tinea species. A common species of deuteromycete associated with animal, avian, and human disease is Aspergillus, which causes aspergillosis. Infections may induce lesions on the surface of the skin or may damage internal organs such as lungs and liver. Infection frequently occurs when hosts are under stress, and immunosuppressive situations result from the poor health (e.g., AIDS virus). Indirect damage from some of these fungi results from ingestion of infected food (feed) or through inhalation of particles containing mycotoxins that are either carcinogenic or cause other health problems. Mycotoxins are produced by fungi during the growth of the crop, or during transportation, processing, and storage. For example, aflatoxin levels in corn increase during drought and when high levels of nitrogen fertilizers are applied. Different types of mycotoxins are produced by species of Aspergillus, Fusarium, and Penicillium. One of the most important groups of toxins is the aflatoxins produced by Aspergillus species and occur primarily on crops such as corn, cotton seed, and peanuts. Another group of mycotoxins are fumonisins that are produced by F. verticillioides ( = moniliforme) and F. proliferatum (Gelderblom et al., 1988). Fumonisin is primarily associated with F. verticillioides, which can routinely be cultured or identified from corn tissues (Bacon and Nelson, 1994). Several forms of the toxin exist, but FB1, FB2, and FB3 are the most common and important that are typically associated with food and feed (Gelderblom et al., 1988). If ingested, fumonisin can cause a neurological disorder in horses, called leukoencephomalacia, pulmonary edema in pigs, and esophageal cancer in humans.
TAXONOMY OF THE DEUTEROMYCETES The size, shape, and septation pattern of conidia are used as the primary characters for the practical and working identification of deuteromycetes genera and species.
FIGURE 16.1 Condia or asexual spores can be simple or complex and can be single or multicellular. (Drawing courtesy of Joe McGowen, Mississippi State University).
Conidia are defined as asexual, nonmobile spores that belong to the anamorphic stage of a fungus life cycle. The Saccardian system, which used spore type to identify the deuteromycetes, was the first major tool employed by mycologists to identify genera (Saccardo 1899). The Saccardian system was incorporated into other systems that included more natural classification schemes based on conidial ontogeny or development. This more advanced system is usually referred to as “The Hughes-TubakiBarron System of Classification.” The work by Barnett and Hunter (1986) includes keys and descriptions for identification of genera. Finally, Hennebert and Sutton (1994) identified subtle differences in spore development on conidiophores for identifying genera and species.
MORPHOLOGICAL STRUCTURES Conidia (Figure 16.1) are produced on specialized hyphae called conidiophores. Because there are a large number of deuteromycete fungi, much variation can occur in their reproductive structures. Conidia vary in shape and size and can be one- or two-celled or multicellular, depending on the number of septa present. Septa within the spores vary, from transverse (across) to longitudinally oblique. Shapes range from filiform (thread-like), ovoid (egg-shaped), clavate (club-shaped), cylindrical (cylinder-shaped), stellate
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FIGURE 16.2 Fungi that produce conidia borne on loosely spaced conidiophores are called Hyphomycetes (Drawing courtesy of Joe McGowen, Mississippi State University).
(star-like), or branched. Conidia can be ornamented with appendages and appear hyaline to colored. Conidiophores are specialized hyphae, branched or unbranched, bearing specialized conidiogenous cells at the points where conidia are produced. Conidiophores may occur singly (separate) or in organized groups or clusters. If conidiophores are formed individually (Figure 16.2) and not enclosed in specialized structures, then the fungi that produce these forms are called Hyphomycetes. The hyphomycetes are subdivided into two groups based on color of hyphae and spores. The dematiaceous group has dark hyphae and spores, whereas the moniliaceous species possess light or pale-colored hyphae and spores. Dematiaceous genera of deuteromycetes include plant pathogenic species such as Alternaria, Aspergillus, Bipolaris, and Penicillium. An example of an economically important dematiaceous hyphomycete is Alternaria solani, the causal organism of early blight of tomato. The disease caused by A. solani is considered by many to be the most economically damaging to tomatoes in the U.S. The disease occurs each year because the pathogen overwinters in plant debris in soil as chlamydospores (thick-walled survival spores). As the temperature warms in the spring, the chlamydospores germinate within the plant debris or soil, and hyphae continue to grow saprophytically, forming conidia on individual conidiophores. Conidia produced during the saprophytic stage are disseminated by wind, rain, and insects or are transported in soil on farm machinery. For long-distance dissemination, infected seed is the primary source. Under wet and warm conditions, the conidia
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present on the tomato plant tissue germinate and hyphae can either penetrate the host through stomata or directly through the cuticle. Infections usually occur first on the mature foliage. Lesions develop and conidia are produced within the necrotic areas of the tomato foliage or stems. Secondary infections can occur from the local dissemination of new condia formed on the host and plants can become completely defoliated. The fungus can develop chlamydospores that remain dormant in the dead plant tissues to start the cycle over for the following season. Symptoms of early blight can be observed on all aboveground tomato parts. Following germination, pre- and postemergence damping-off of plants can occur. Because lesions are generally first observed on mature leaves, the disease appears to progress from the lower portion, moving upward to the top of the plant. Infections increase forming circular lesions up to 4 to 5 mm in diameter, and the lesions become brown, with concentric rings giving the necrotic area a target-shaped appearance. Leaves that are infected often are observed with yellowing areas. As multiple lesions occur from secondary infections, the leaves turn brown and die. The entire plant can become defoliated and die at this stage. Controls for the disease include avoiding purchase of infected seed and soil for transplants that harbor the fungus; crop rotation with other solanaceous plants, such as potatoes, eggplant, and peppers; removal or burial of crop residue; and use ofdiseasefree plants, fungicides, and resistant varieties.
DEUTEROMYCETE CONIDIOMATA If condiophores are grouped together into organized clusters, then they are formed within specialized structures called conidiomata. The different types of conidiomata include acervuli, pycnidia, sporodochia, and synnemata. Modern references assign acervuli and pycnidia to the group Coelomycetes. Sporodochia-forming species are considered under the dermatiaceous or moniliaceous subgroups of hyphomycetes. Synnemata have been placed under the hyphomycetes in the subgroup stilbaceous fungi (Alexopoulus et al. 1996). Conidiophores and conidiomata on hosts or in culture are used for identification. The ability of the deuteromycetes to form these structures in culture vs. that on a host varies per genus and species. A structure that is routinely formed on a host may not often be observed when grown in culture media. An example is setae (sterile hyphae or hairs) that are associated with acervuli of Colletotrichum spp. Setae (sterile-like appendages) routinely form in the acervuli produced on living hosts, but may be absent when growing on selective medium. As most identification keys are based partially on the morphology of the pathogen on host tissue, proper identification may require direct observation of conidiomata development on plant tissue rather than on artificial media.
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FIGURE 16.3 Sporodochia form on the surface of the host plant containing clusters or groups of conidiophores. (Drawing courtesy of Joe McGowen, Mississippi State University.)
Sporodochia are similar to acervuli except that the cluster or rosettes of conidiophores form on a layer or cushion hyphae on the host surface (Figure 16.3), whereas acervuli are imbedded in epidermal tissue or the plant cuticle. Sporodochia also appear as mat-like cottony structures due to the clustering of condiophores. However, in culture, sporodochia that resemble those on host materials are rarely observed, making species identification difficult. Examples of specific genera of fungi forming sporodochia include Epicoccum, Fusarium, and Strumella. Fusarium oxysporum f. sp. vasinfectum, causal agent of Fusarium wilt of cotton, is a good example of a sporodochia-forming fungus. The conidia of this pathogen overwinter in plant debris or can be introduced into fields by infected seed or in soil transported by farm equipment. The fungus forms chlamydospores in the soil or in plant debris. Under optimum weather conditions, the conidia or chlamydospores germinate and the fungus grows saprophytically, producing conidia on conidiophore rosettes. Germinating conidia or hyphae that come in contact with host root tissue can invade by direct penetration or indirectly through wounded areas on the root. Root wounding is often increased by the root-knot nematode, Meloidogyne incognita, and the occurrence of this pest has been directly associated with fields with increased levels of Fusarium wilt. Following invasion into the root, the hyphae then grow inter- and intracellularly through the cortex and endodermis. The fungus penetrates the vascular system, and conidia are rapidly produced and distributed systemically into the transpiration stream of the cotton plants. The fungus physically obstructs the lumens
FIGURE 16.4 Synnemata consist of fused condiophores at the base, forming conidia at the apex or on the sides of the structure. (Drawing courtesy of Joe McGowen, Mississippi State University.)
of the xylem tissue, preventing water movement, which eventually results in wilting and death of the plants. Chlamydospores form in dead host tissues and overwinter in the plant residue. Control practices include use of clean seed from uninfested fields, use of resistant varieties, and reduction of root-knot nematode levels through chemical control, rotation, or with nematode-resistant varieties. Synnemata conidiomata form conidiophores that are fused and the conidia often form at or near the apex (Figure 16.4). Species that produce conidia in this fashion belong to the Stilbellaceae group of deuteromycetes. Conidiophores of the synnemata-forming species are usually elongate and easy to identify from cultures because of their upright, whisker-like appearance. Synnemata are generally formed in culture unlike the sporodochial-forming species. Common genera that form synnemata are Graphium, Arthrosporium, Isaria, and Harpographium. Graphium ulmi, the anamorph for the causal agent of Dutch elm disease, produces synnemata during the asexual
Deuteromycota: The Imperfect Fungi
or conidial stage of the life cycle. In the spring, the pathogen, which overwinters in plant debris, grows saprophytically and forms mycelia where the synnemata are produced. The spores of the pathogen can then infect healthy trees by an insect vector, and if roots of healthy plants are grafted to an infected tree, the pathogen can be transmitted to the healthy tree. The conidia are primarily disseminated by the elm bark beetle (Scolytidae) carried on their body parts. As the insects feed on the tree tissues, they deposit conidia into the feeding wound sites. The fungus germinates, becomes established, and grows into the xylem tissue. Blockage of the vascular system occurs from the fungus and by defense responses of the host tree. Symptoms of Dutch elm disease include yellowing and wilting on one to many branches early in the season, depending on when infection occurs. Leaves of infected trees turn brown and die in portions of the tree or the entire tree may be affected. If the tree survives the first years following invasion, death will occur sometime during the second year of infection. If stems of the tree are sectioned, a brown discoloration is observed in the outer xylem of twigs, branches, and sometimes roots. Also, in dead and dying trees, insect larval galleries from the elm bark beetle can be observed under the bark of the tree trunk. Controls for Dutch elm disease include methods to eliminate the vector and pathogen by removing dying and dead wood (sanitation — Chapter 32). Fungicides injected into the will stop the spread of the pathogen, but treatments must be repeated continuously from one year to the next.
COELOMYCETES CONIDIOMATA Acervuli (sing. acervulus) contain a defined layer of conidiophores and conidia formed just below the epidermal or cuticle layer of plant tissues (Figure 16.5). Conidiophores and conidia erupt through the host epidermis or cuticle exposing the acervulus. Conidiophores within these structures are generally short and simple compared to the hyphomycetes, such as Aspergillus and Penicillium. Once exposed, acervuli are usually saucer-shaped in appearance (Barnett and Hunter, 1986). In culture, fungi that typically form acervuli on host tissue can often appear to produce sporodochia. The eruption of host cuticle or epidermis, which defines an acervulus, cannot be observed in culture. An example of an acervulus-forming pathogen is Colletotrichum lagenarium, the causal agent of anthracnose of cucurbits. Colletotrichum lagenarium overwinters on plant debris and in seeds obtained from infected fruits. The fungus grows saprophytically in the soil and produces conidia on dead tissue. The conidia are disseminated locally by rainfall and soil transported on farm equipment and over longer distances by infested seed. When weather conditions are suitable for infection, conidia germinate on the host. The germ tubes (hyphae) form appressoria at the
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FIGURE 16.5 Acervulus embedded in host tissue containing clusters or groups of conidiophores (Drawing courtesy of Joe McGowen, Mississippi State University.)
point of contact with the host cell, quickly followed by formation of infection pegs, which allow direct entry into the host. Hyphae then grow intracellularly, killing host cells. As the fungus continues to grow, angular light to dark brown or black lesions are formed between the leaf veins. The lesions are elongate, narrow, and water-soaked in appearance and become sunken and yellowish to brown. When conditions are favorable, acervuli form on stromal tissue and conidiophores containing conidia erupt through the cuticle of the host. Spores are exposed to the environment and disseminated. Girdling of the stems or petioles can occur and defoliation results. Control practices include the use of disease-free seed, crop rotation with resistant varieties, cultural practices that remove or bury plant debris, fungicide sprays, and use of resistant cucurbit varieties when available. Pycnidia (sing. pycnidium) differ from acervuli by the formation of the flask-shaped structures composed of fungal tissue that enclose the conidia and conidiophores (Figure 16.6). Pycnidia shapes described by Alexopoulus et al. (1996) include the following: papillate, beaked, setose, uniloculate, and labyrinthiform. Conidiophores that form within the pycnidium can be extremely short as in Phoma or larger as in Septoria or Macrophoma. Pycnidia resemble perithecia, which are sexual reproductive structures of some species of the Ascomycota (Chapter 15). If observed microscopically, the spores of the ascomycetes are borne in asci and not on conidiophores. Other important considerations when identifying pycnidia-producing deuteromycetes is the presence or absence of an ostiole, which is located at the apex and where conidia are exuded in a thick layer or cirrhus. Keys to the identification of deuteromycetes often refer to imbedded pycnidia within the host
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FIGURE 16.6 Pycnidium, a flask-shaped structure composed of fungal tissue, can be either embedded in or superficial on host tissue. Large numbers of conidiophores and conidia formed within the structure. (Drawing courtesy of Joe McGowen, Mississippi State University).
material, but in pure culture, pycnidia of the same fungal species will occur superficially on artificial media. Septoria glycines, the causal agent of brown spot of soybean, produces conidia within pycnidia. The fungus can reinfect plants within the same field during subsequent years since the conidia or mycelium overwinter on debris of host stem and leaf tissues. During warm and moist weather, sporulation occurs as the fungus is growing saprophytically and the conidia are disseminated by wind or rain. The spores germinate and hyphae invade by passing through stomata. The pathogen grows intercellularly, killing adjacent cells. Lesions and the new conidia form that can serve as a source for secondary infections on the same host plants. The pathogen primarily invades foliage, causing a flecking appearance on mature leaves, but infections can also occur on stems and seed. If environmental conditions are optimum, secondary infections can occur, resulting in defoliation that generally moves from the lower leaves and progresses upward. Lesions are usually irregular, becoming dark brown and can be up to 4 mm in diameter. During initial development of the pathogen, lesions often coalesce to form irregular-shaped spots. On young plants, leaves turn yellow and abscise, but late in the growing season, infected foliage can be rusty brown before falling off. Control methods are limited to the use of resistant varieties, rotation to nonhost crops, and fun-
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gicides. The latter methods are generally not considered to be economically feasible. Species in the Mycelia Sterilia are traditionally included in the Deuteromycota as a group that does not form asexual spores. Identification of these fungi is based on hyphal characteristics, absence or presence of sclerotia (survival structures) and number of nuclei per hyphal cell. Rhizoctonia solani and Sclerotium rolfsii have teleomorphs that place them into the Basidiomycetes (Chapter 19). Both of these fungi are important plant pathogens that occur worldwide and attack agronomic, vegetable, and ornamental crops. Rhizoctonia solani has many morphological and pathogenicity forms called anastomosis groups. Rhizoctonia species, which are responsible for brown patch of turfgrass, survive as sclerotia in plant debris in the soil. Over a wide range of temperatures, the sclerotia germinate and the fungus grows saprophytically until a suitable host becomes available. Once the fungus becomes established in the host, circular lesions develop. Leaves and sheaths lose their integrity and appear water-soaked. The damaged tissue at first has a purplish-green cast, which then becomes various shades of brown depending on weather conditions and the type of grass. Often, darkpurplish or grayish-brown borders can be observed around the infected areas. Fungicide applications are effective in preventing or reducing severity of the brown patches. The disease may also be controlled by the following cultural practices: avoiding excessive nitrogen applications that enhance fungal growth, increasing surface and subsurface drainage, removing any sources of shade that reduce direct sunlight and increase drying of the leaf surface, and reducing thatch build-up when possible. In summary, the deuteromycetes are a very diverse group based on the presence of conidiophores and conidia, except for the fungi that do not produce asexually. The reader should keep in mind that when mycologists first created this artificial group, the sexual reproductive stages were unknown. Although the teleomorphs have now been determined for many of the deuteromycetes, these sexual stages are often rare or almost never seen on a host or in culture. As many deuteromycetes are plant pathogens, the group has been maintained for identification based on their asexual reproductive structures and cultural characteristics when identifying members of the Mycelia Sterilia.
REFERENCES Alexopoulus, C.J., C.W. Mims, and M. Blackwell. 1996. Introductory Mycology, 4th ed. John Wiley & Sons, New York, 869 pp. Bacon, C.W. and P. E. Nelson. 1994. Fumonisin production in corn by toxigenic strains of Fusarium moniliforme and Fusarium proliferatum. J. Food Prot. 57: 514–521.
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Barnett, H.L. and B.B. Hunter. 1986. Illustrated genera of Imperfect Fungi, 4th ed. Burgess, Minneapolis, MN, 218 pp. Fries, E.M. 1821. Systema Mycologicum 1. Gryphiswaldiae, Lund, Sweden, 520 pp. Gelderblom, W. C. A., K. Jaskiewicz, W. F. O. Marasas, and P. G. Thiel. 1988. Novel mycotoxins with cancer-promoting activity produced by Fusarium moniliforme. Appl. Environ. Microbiol. 54: 1806–1811.
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Hennebert, G.L. and B.C. Sutton. 1994. Unitary parameters in conidiogenesis. In: Ascomycetes Systematics: Problems and Perspectives in the Nineties. Plenum, New York, pp. 65–76. Link, J. H. F. 1809. Observationes in ordines plantarum naturales. Mag. Ges. Naturf. Freunde, Berlin, 3: 3–42. Persoon, D.C.H. 1801. Synopsis Methodica Fungorum. H. Dieterich, Göttingen, 706 pp. Saccardo, P.A. 1899. Sylloge Fungorum Omnium Hueusque Cognitorum, Vol. 14. Self- published, Pavia, Italy.