The Classification of Fungi and Slime Moulds into Major Groups

For an organism to be formally recognized by taxonomists, it must be described

in accordance with internationally accepted rules and given a Latin binomial,
namely a generic name followed by a specific epithet. Man, for example, is Homo
sapiens, and the cultivated mushroom, as indicated in the previous chapter,
Agaricus bisporus. It was estimated in 1983 that about 64 000 fungal species
were known, and in 1995 the estimate was 72 000, suggesting that about 700 new
species are discovered each year. The number of species so far discovered,
however, is probably only a small proportion of those that exist, as few habitats
and regions have been intensively studied. Various approaches have been used in
trying to estimate the number of fungal species in the world. For example, in well-
studied regions fungal species can be six times as numerous as those of flowering
plants. On this basis, since about 270 000 flowering plants are known, there may
be about 1.6 million fungal species. Other approaches suggest similar or larger
numbers.
A genus may contain one, a few or many species. Since there are such a large
number of fungal species, there are also a large number of genera, most of which
will be unfamiliar to any one mycologist. Hence categories intermediate in status
between genus and kingdom are needed. Genera can be grouped into families,
families into orders, orders into classes, and classes into phyla (sing., phylum)
(Table 2.1). Mucor mucedo, for example, is in the class Zygomycetes, which is in
the phylum Zygomycota. In a formal classification the ending -mycetes indicates
a class, and -mycota a phylum. Standard endings are also used for most of the
other formal taxonomic categories. Informal terms are also used. The
Zygomycota, for example, are often referred to informally as Zygomycetes,
although formally the Zygomycetes are the larger of two classes within the
Zygomycota. In addition there are informal groupings, such as the yeasts, which
do not correspond to any formal grouping.
Formal classifications and nomenclature change as new information becomes
available, and at any one time there are differences of opinion as to classification
and the names of categories. There is greater stability as regards informal
nomenclature which will therefore generally be used in this book, although
formal categories will be mentioned as necessary, and an outline formal
classification is provided in Appendix 2.
Table 2.1 Categories in classification illustrated by reference to two Zygomycetes,
Mucor mucedo (page 39) and Glomus macrocarpum (page 397)
Kingdom Fungi
Phylum Zygomycota
Class Zygomycetes
Order Mucorales a Order Glomalesb
Family Mucoraceae Family Glomaceae
Genus Mucor Genus Glomus
Species mucedo Species macrocarpum
a There are several familiesin the order Mucorales, and many genera in the familyMucoraceae. The
genus given illustrates the use of the same name with different endings at several levels in a
hierarchy.
b The order Glomaleshas only one family, but the familyhas severalgenera.

It is now clear that the fungi studied by mycologists include organisms from
three Kingdoms, the Chromista, the Protozoa and the Fungi. The features that
distinguish these three Kingdoms are set out in Table 2.2. Only the Kingdom
Fungi consists exclusively of fungi. The Chromista are aquatic and mainly
photosynthetic organisms. The latter, the algae, are studied by plant biologists
and include the seaweeds, as well as filamentous and microscopic unicellular
forms. The group of fungi informally called Oomycetes or water moulds have
structural, genetic and biochemical features that have now established them as a
phylum, the Oomycota, within the Chromista. They are common microfungi
with many species and include important plant pathogens such as the organism
that causes potato blight. They have motile spores which swim by means of two
flagella, and grow as hyphae with cellulose-containing walls. In addition to the
Oomycetes there are two other Chromistan groups of fungi which are also
aquatic or parasitic and with motile spores; the Hyphochytriomycota and the
Labyrinthulomycota. They have few species and are dealt with briefly in
Appendix 2. The Protozoa are diverse unicellular organisms with separate lines of
descent from a unicellular ancestor. The fungi known as slime moulds all belong
to this kingdom. They do not form hyphae, lack cell walls during the phase in
which they obtain nutrients and grow, and are capable of ingesting nutrients in
particulate form by phagocytosis. The slime moulds hence fail to meet any normal
definition of the fungi, and are now established as members of the Kingdom
Protozoa, but they produce fruiting bodies which have a superficial resemblance
to those of fungi. This resulted in slime moulds early attracting the attention of
mycologists, and to their inclusion in most textbooks on mycology. Two of the
best studied groups of slime moulds are the cellular slime moulds and the
plasmodial slime moulds or Myxomycetes.
The Kingdom Fungi consists solely of species that are hyphal, or clearly related
to hyphal species. Throughout most or all of their life cycle they possess walls
which normally contain chitin but not cellulose, and they are exclusively
absorptive in their nutrition. They are divided into four divisions as shown in
Fig. 2.1. The Chytridiomycetes are the only group with motile cells (known as
zoospores). The form taken by the sexual phase of the life cycle is an important
criterion in the classification of fungi that lack zoospores. The sexual process
leads to the production of characteristic spores in the different groups. The fungi
Table 2.2 Important groups of fungi
Features that may be shared by several groups Group Kingdom

Trophic phase a lacking cell walls. 1. Cellular slime moulds.

Able to ingest particulate food. Amoebae aggregate to
The slime moulds form a 'slug' which gives
rise to a fruit body.
Example: Dictyostelium
Protozoa
2. Plasmodial slime moulds
(Myxomycetes).
Amoeboid phase followed
by a multinucleate
plasmodial phase.
Example: Physarum

3. Oomycetes. Zoospores
biflagellate b, sexually
produced spores are oospores. Chromista
Walls contain cellulose.
Many Example: Saprolegnia
species
produce 4. Chytridiomycetes.
zoospores
Zoospores uniflagellate c.
Sexual process may involve
fusion of motile gametes,
walls contain chitin.
Example: Allomyces
Trophic phase with
cell walls; nutrition
exclusively absorptive.
The fungi
(in the
informal sense) 5. Zygomycetes. Sexually
produced spores are zygospores.
Example: Mucor
Fungi
6. Ascomycetes.
Sexually produced spores are
ascospores. Example: Pyronema
Zoospores
not 7. Basidiomycetes (mushrooms,
produced toadstools, rusts and smuts).
Sexually produced spores are
basidiospores. Example:
Agaricus
8. Mitosporic fungi. No
sexually produced spores.
Example: Penicillium
a The phase concerned with nutrition and growth.
b Having two flagella.
c Having a single flagellum.
that form zygospores are classified as Zygomycetes, those that form ascospores as
Ascomycetes, and those forming basidiospores as Basidiomycetes. The hyphae of
Ascomycetes and Basidiomycetes have numerous cross-waUs. Another feature
widespread in Ascomycetes and Basidiomycetes is that when hyphae within a
fungal colony come into contact they may fuse with each other. This hyphal
anastomosis, if frequent, can convert the radiating hyphae of a colony into a
three-dimensional network. Hyphal anastomosis, as indicated later, may be a
major factor in permitting the mycelium of some Ascomycetes and
Basidiomycetes to produce large fruit bodies. Cross-walls and hyphal
anastomoses are largely lacking in the Zygomycetes and Chytridiomycetes. These
organisms are sometimes termed the 'lower fungi', in contrast to the 'higher
fungi', the Ascomycetes, Basidiomycetes and related forms. There is some
justification for a loose distinction of this kind, in that the potentialities of hyphal
and mycelial organization have been more fully exploited in the latter groups.
Many Ascomycetes and Basidiomycetes, in addition to producing spores by a
sexual process, form other types of spores asexually. There are also many species,
recognizable as higher fungi through the presence of cross-walls in their hyphae,
that produce asexual spores but lack a sexual phase. These are known as
mitosporic fungi, as all their spores are produced following mitosis but none by
meiosis. They were formerly termed the Deuteromycetes or Fungi Imperfecti, and
a Deuteromycete was reclassified as an Ascomycete or Basidiomycete if a sexual
phase was discovered. However, analysis of DNA sequences now allows these
asexual fungi to be classified with their closest sexual relatives, and it appears that
they have arisen from many different groups of fungi by the loss of sexuality.
They thus do not constitute a natural group, and ultimately they could all be
assigned to Ascomycete or Basidiomycete groups.
Some important features of the fungal groups mentioned above are indicated in
Table 2.2. Fig. 1.2 shows the relationships between the Kingdoms to which fungi
belong, and other living organisms. The way in which the four major divisions of
the Kingdom Fungi are thought to have diverged is shown in Fig. 2.1.
These groups also form the subject of the rest of the chapter, along with two
additional groups, the yeasts and the lichens. Yeasts are fungi that are normally
unicellular and reproduce by budding, although some will, under appropriate
conditions, produce hyphae, just as some normally hyphal fungi may produce a
yeast phase. Many yeasts have a sexual phase that enables them to be classified as
Ascomycetes or Basidiomycetes, although some do not. It is useful, however, to
deal with the yeasts, which have much in common with each other with respect to
form, habitat, practical importance and methods of identification as a group, and
many books are devoted to the subject of yeasts, as are some institutes. The
lichens are intimate symbiotic associations of a fungus, nearly always an
Ascomycete, with an algal or a cyanobacterial species. The fungal components of
lichens can be assigned to order within the Ascomycetes, but it is often useful, on
the basis of morphology, physiology and ecology, to consider the lichens as a
group.
The eight groups listed in Table 2.2 along with the yeasts and lichens will now
be considered in more detail.
 j ~ Ascomycetes
I i
! I
t i

I i
I
Basidiomycetes
h
/

Zygomycetes

! [- } Chytridiomycetes

I 1 I I I 1 I
600 500 400 300 200 100 0
Millions of years

Figure 2.1 The pattern of divergence of fungal phyla. The tree was constructed from
comparisons of 18S RNA gene sequences of over 30 species of fungi. The dates of divergences
are based on the earliest appearance of key structures in the fossil record. For example, the
oldest fossils showing clamp connections, diagnostic of Basidiomycetes, are almost 400
million years old. Dates of divergence available from the fossil record were used to deduce
that the average rate at which the rDNA base sequence changes is 1% per hundred million
years. Hence divergence times could also be theoretically assigned to branches in the
phylogenetic tree for which there was no fossil evidence available. Both fossil and molecular
data indicate that the diversification of Ascomycetes and Basidiomycetes has occurred in
parallel with the diversification of a land flora during the last 500 million years, and that the
first terrestrial fungi evolved about 550 million years ago. Fossil Ascomycete fruiting bodies
resembling present day Pyrenomycetes have been found on a 400 million year old fossil plant.
Based on Berbee, M.L. & Taylor, J.W. (2000), The Mycota, Springer-Verlag, Berlin;
Taylor,T.N., Hass, H. & Kerp, H. (1999) Nature 399, 648.

The Cellular Slime Moulds

The slime moulds are Protozoa that have been much studied by mycologists. The
cellular slime moulds are so designated to contrast them with slime moulds that
produce plasmodia. Two phyla are recognized, the Acrasiomycota and the
Dictyosteliomycota. Members of the Acrasiomycota occur mainly on dung and
decaying vegetation and will not be considered further. Members of the
Dictyosteliomycota occur in soils throughout the world, especially in the surface
soil and leaf litter of deciduous forests. Although they are common, only about
fifty species are known. One of these, Dictyostelium discoideum, has been studied
intensively by biologists and biochemists interested in cellular interaction and
developmental processes. The life cycle of D. discoideum (Figs. 2.2, 2.3) will now
be described.
 MACROCYST~.

,~, 'k~ Mating,with

" Meiosis' 2n.. ._2,_, _ _ 4 _ cell fusion
~ n n I fundonnUClear

and amoeba f ,~.~..! ;:::~ i ' ~ Starvationa d

multipli eba aggregation

SPORE
(---

Migration
Di ~ and differentiation
FRUITING,~,,.,,'J
BODY

Figure 2.2 The life cycle of the cellular slime mould Dictyostelium discoideum. An
amoeba is shown with a central nucleus and a contractile vacuole, and is ingesting a rod-
shaped bacterium. When starvation occurs in a population of amoebae, aggregation gives
slugs, which may consist of thousands of amoebae and be I mm long. After migration to a
suitable site, slugs differentiate into a fruiting body consisting of a basal disc, a stalk, and
a sporangium containing spores. If a spore reaches a suitable substratum, it germinates and
an amoeba emerges. Amoebae that differ in mating type can mate to give diploid (2n)
macrocysts, with loosely textured primary walls and more dense, secondary, inner walls.
Macrocysts are capable of prolonged survival, and on germination undergo meiosis to give
haploid (n) amoebae once more.
Figure 2.3 Amoeba aggregation in the cellular slime mould Dictyostelium discoideum. A,
Centres of attraction have formed and are surrounded by bright zones of elongated
amoebae moving towards centres, and dark zones of roughly circular, temporarily
stationary amoebae. B, After about an hour the zones are breaking up into streams moving
towards the centres. C, After a further hour all the amoebae have joined streams. Dark-
field microscopy. (Reproduced with permission from Newell, P. C. (1981). Chemotaxis in
the cellular slime moulds. In Lackie, J. M. & Wilkinson, P. C., eds, Biology of the
Chemotactic Response, pp. 89-114. Cambridge University Press, Cambridge.)

The amoebae of D. discoideum can be grown readily in two-membered culture

with a variety of bacteria such as Escherichia coli. An agar medium that will
permit growth of the chosen bacterial species is spread with bacteria and
inoculated with Dictyostelium amoebae. The bacteria multiply and the amoebae
feed on them by phagocytosis, taking the bacteria into food vacuoles within
which the bacteria are digested. Under suitable conditions nuclear division
followed by cell division occurs about every 3 hours, so a large amoeba
population is soon produced. Mutant strains of D. discoideum have been
obtained that can be grown in pure culture on a complex soluble medium, but
growth is slower with a generation time of about 9 hours. The nutrition of
cellular slime mould amoebae is, however, mainly ingestive, particulate food
normally being taken by phagocytosis.
The amoebae of D. discoideum are about 10 gm in diameter. In addition to
food vacuoles a contractile vacuole is present, expelling excess water that has
entered the cell by osmosis. There is a single haploid nucleus with seven
chromosomes. The haploid DNA content is low in comparison with most
eukaryotes, being about 12 times that of E. coli, or about 50 million base pairs.
Genetic studies can be carried out by making use of the parasexual cycle.
Occasional cell fusion followed by nuclear fusion produces diploid amoebae. This
is a rare event, occurring once in a population of 10 s to 10 6 amoebae, but selective
procedures have been devised for the ready isolation of such diploids. Such
diploids can lose chromosomes one at a time to give aneuploids and finally the
haploid condition. Such haploidization can be encouraged by selective
procedures. The loss of whole chromosomes during haploidization makes the
parasexual cycle very useful for assigning genes to linkage groups. Rather
infrequent mitotic crossing over within linkage groups can also occur.

OH 0 COOH
_~ II I

CN2
' I
N2N CN2
I
COON

_ ~ 0 ~ denine OH 0

o~ I OH CH30
ON CI

N N

COOHf J ~

Figure 2.4 Componds regulating cell movement and differentiation in the cellular slime
mould Dictyostelium discoideum. A, Folic acid, a bacterial metabolite that attracts trophic
phase amoebae. Folic acid also acts as a vitamin for the many organisms in which a
derivative of folic acid, tetrahydrofolate, is an enzyme co-factor in the metabolism of C 1
componds. B, Cyclic AMP (3',5'-cyclic adenosine monophosphate) is emitted by
aggregating amoebae, attracting further amoebae into the aggregate. It also has an
important role in metabolic regulation in both prokaryotes and eukaryotes. C, DIF,
differentiation inducing factor, brings about the differentiation of amoebae at the anterior
end of the slug into the stalk cells of the fruiting body. D, Discadenine prevents the
premature germination of D. discoideum spores.
 Efficient location of bacteria by amoebae is facilitated by chemotactic
responses. Amoebae repel each other by a factor, as yet unidentified, that they
release. They hence avoid high concentrations of amoebae, where there will be
few surviving bacteria, and are dispersed to areas where bacteria are more likely
to be present. They also show positive chemotaxis to a product released by
bacteria, folic acid (Fig. 2.4A), and themselves release an enzyme which destroys
folic acid. This presumably prevents the building up of a uniform background
concentration of folic acid, which would not give any indication as to the
direction of the folic acid source and the bacteria.
Ultimately amoebae exhaust the supply of bacteria in their vicinity. Their
behaviour then changes. They cease to repel each other and cease to respond to
folic acid. Instead some begin to emit cyclic AMP (3',5'-cyclic adenosine
monophosphate (AMP), Fig. 2.4B) and others respond to this substance by
positive chemotaxis. Attractant centres are formed. These centres emit pulses of
cyclic AMP every few minutes. Nearby amoebae respond by moving towards the
centre for about 100 s, covering about 20 ~m and also releasing a pulse of cyclic
AMP which attracts amoebae further from the centre. After a refractory period
amoebae recover cyclic AMP sensitivity and become ready to respond to a further
pulse of cyclic AMP. Thus, a relay system operates that can attract amoebae a
centimetre or more from a centre. Each centre becomes surrounded by a field of
amoebae moving towards the centre. With dark field microscopy alternate zones
of moving amoebae, which are elongated and bright, and stationary amoebae,
roughly circular and dark, can be recognized (Fig. 2.3). Subsequently, the field
breaks up into streams moving towards the centre. Finally, all the amoebae within
range of a centre's influence reach the centre to produce an aggregate which,
depending on the amoeba population at the time food was exhausted, may
contain from a few hundred to a few hundred thousand cells.
The aggregation process in D. discoideum has been the subject of intensive
study, and the sensory transduction path, from the binding of cyclic AMP at the
surface of the plasma membrane to the movement of the amoeba in the direction
from which a cyclic AMP pulse originated, is gradually being elucidated.
Amoebae produce phosphodiesterases which destroy cyclic AMP. One is released
into the medium and presumably prevents the build-up of a uniform background
of cyclic AMP, the other is membrane bound and perhaps frees receptors from
cyclic AMP thus permitting response to further pulses.
In some cellular slime moulds attractants other than cyclic AMP are
responsible for aggregation. Cyclic AMP is, however, the attractant for several
Dictyostelium spp. other than D. discoideum. There is hence the possibility that
a single centre may attract more than one species. If this occurs a sorting process
takes place, resulting in aggregates consisting of cells of only one species. This is
the result of species specificity in cell adhesion, resulting from the release of
species-specific proteins. A molecule of such a protein has two sites able to bind
to surface polysaccharides on the amoebae of the producer species, and can thus
cause adhesion between such cells but not those of other species. The specific
proteins involved in cell adhesion in D. discoideum are called discoidins.
The mass of cells resulting from aggregation develops into a slug-like
organism (sometimes termed a grex or pseudoplasmodium) which is enclosed in
a slime sheath. The 'slug', depending on the number of cells in the aggregate from
which it originated, may be minute or as much as 1 mm in length. It can migrate
for several days, and is positively thermotactic and phototactic, moving towards
warmth and light. In nature this will help the slug to move through leaf litter or
soil to a site on the surface suitable for the development of a fruiting body from
which spores can readily disperse.
The fruit body consists of a basal disc (hence the specific epithet, discoideum),
a multicellular stalk and a roughly spherical mass of spores, the sporangium. The
stalk consists of cell wall materials, largely cellulose, secreted by the stalk cells
before they die. During slug migration the cells that will become the stalk (the pre-
stalk cells) are at the tip of the slug. The conversion of amoeboid pre-stalk cells
into the vacuolate, walled, stalk cells is brought about by a differentiation
inducing factor, DIF (Fig. 2.4C) produced at the tip of the slug. The fruiting
bodies, as they rise from the substratum, avoid colliding with each other. This,
and the adequate spacing of fruiting bodies, is due to the emission by the
developing fruiting bodies of a volatile factor, ammonia, which repels other
fruiting bodies. The spores can remain viable for several years, their premature
germination either within the sporangium or in a dense mass being prevented by
a germination inhibitor, discadenine (Fig. 2.4D). When spores are well dispersed
the inhibitor is lost by diffusion. Germination is stimulated by amino acids, which
will be encountered if a spore arrives in the vicinity of bacteria. A germinating
spore swells, the spore wall ruptures, and an amoeba emerges and begins to feed
on bacteria. Under unfavourable conditions the amoebae of some cellular slime
moulds, but not D. discoideum, can develop cell walls to become microcysts.
These cells, more resistant than amoebae, germinate when favourable conditions
return.
The life cycle so far described can be accomplished by amoebae that remain
haploid and constitute a clone, that is have originated from a single haploid cell.
A sexual process, however, can be initiated by bringing together amoebae that
differ in mating type. The mating type of a cell is determined by which of two
alleles, m a t A or m a t a, is present. Cell clumping is brought about by a volatile
factor, ethylene, released by m a t A cells and acting on m a t a cells. Cyclic AMP is
then released attracting more cells into the clump. Within the clump, cell and
nuclear fusion occurs between two cells of different mating type. The resulting
zygote ingests and digests many of the surrounding cells to produce a large cell
which develops a thick wall to become a macrocyst. Under suitable conditions the
macrocyst germinates with meiosis occurring and haploid amoebae being
released.

The Plasmodial Slime Moulds (Myxomycetes)

Another protozoan phylum studied by mycologists is the Myxomycota

(plasmodial slime moulds). Two classes are recognized, the Protosteliomycetes,
which will not be considered further, and the Myxomycetes. Members of the
latter, much larger, class produce fruit bodies visible to the naked eye. They are
usually found on dead plant materials and have been collected by naturalists for
over a century. Although not obtrusive, they are common, and an observant
collector can obtain a dozen or more species in a visit of a few hours to woodland.
The fruiting bodies are made of durable materials, and museum specimens can
remain in good condition for many years.
The special feature of the Myxomycete life cycle is the plasmodium, a
multinucleate mass of protoplasm not subdivided into cells. It is a transient stage
in the life cycle, so is less often seen in nature than are the fruiting bodies. Some
species have only minute plasmodia, but in others the plasmodium can reach the
size of a dinner plate; such plasmodia are occasionally seen in nature as a slimy
yellow mass on decaying wood. As with the cellular slime moulds, there is an
amoeboid phase in the life cycle. Myxomycete amoebae, however, differ from
those of the cellular slime moulds in that they can produce flagella and swim. The
amoebae of Myxomycetes are common in soil and in decaying timber. Both
amoebae and plasmodia are phagotrophic, in this resembling protozoa rather
than true fungi.
About 700 Myxomycete species are known, and of these about 300 are in the
order Physarales, which have large plasmodia. A few isolates of one species,
Physarum polycephalum, have been extensively used for research in cell and
molecular biology. Studies on genetic variability in nature have been carried out
mainly with a second species, Didymium iridis. An account of the life cycle (Figs.
2.5, 2.6) of P. polycephalum follows.
The amoebae of P. polycephalum feed on bacteria, and like those of
Dictyostelium are readily grown in two-membered culture with Escherichia coli.
They have been grown in pure culture on a complex medium, but multiply more
slowly. The nucleus has a prominent nucleolus, and food vacuoles and a
contractile vacuole are present. If a culture is flooded with water, the amoebae
elongate and turn into flagellates. The flagella are smooth and emerge from the
anterior end of the cell. There are usually two flagella, but whereas one, pointing
forward, is active, the other commonly points backward, is held close to the cell
surface, and is inactive. The flagellates neither feed nor undergo cell division, and
when free water disappears, revert to the amoeboid state. When all the bacteria
present have been consumed, the amoebae turn into thick-walled cysts, which can
survive for a long period in the absence of nutrients. In the presence of bacteria a
cyst will germinate and an amoeba emerge. The amoeboflagellate phase is hence
able to multiply in the presence of bacteria, swim when flooding occurs, and
survive periods without nutrients. It can be maintained indefinitely in two-
membered culture and could probably similarly persist in nature;
amoeboflagellate protozoa are common in soil and water.
Initiation of the plasmodial phase usually requires mating between two
genetically different strains. Plasmodium initiation occurs with the highest
frequency if the two strains differ at two loci, designated mat A and mat B. At
each locus many alleles have been found so the number of possible mating types
is large. If two amoebae differ at the mat B locus the probability that they will
fuse to give a diploid cell is greatly increased. The probability that a diploid cell
(zygote) will develop into a plasmodium is greatly increased if it contains two
different mat A alleles. Although plasmodium formation is most likely if amoebae
differing at both mating type loci are brought together, plasmodium formation
can occur at a much lower frequency when there are differences at only one of the
Figure 2.5 The life cycle of the Myxomycete Physarum polycephalum. An amoeba is
shown with a contractile vacuole and a nucleus with a prominent nucleolus, typical of
Physarum. Starvation results in cyst formation, and free liquid in conversion to flagellates.
Cell and nuclear fusion between haploid (n) amoebae of different mating type initiates the
formation of diploid (2n) plasmodia, which grow large and show rapid protoplasmic
streaming in prominent veins. Starvation of plasmodia may cause the formation of
sclerotia consisting of multinucleate spherules, but accompanied by exposure to light,
results in fruit body development. Meiosis occurs during spore formation, and under
suitable conditions spores germinate to give haploid amoebae. The life cycle of the
apomictic strain is identical except that there is no mating or ploidy change.

loci, and very occasionally, within a clone. P. polycephalum is hence normally

self-sterile (heterothallic), and has diploid plasmodia, since zygote production is
involved in their origin. There are also, however, facultatively apomictic strains.
In these a mutation at the mat A locus enables amoebae to give rise, without
mating, to plasmodia which are haploid. The amoebae of these strains can also
Figure 2.6 Some stages in the Myxomycete life cycle. A-C, Physarum polycephalum. A, Plasmodium
migrating along and ingesting a streak of washed yeast cells on water agar. Plasmodia in pure culture on an
agar medium are shown in Fig. 5.5. B, Microplasmodia in shaken liquid culture. C, Microsclerotia, consisting
of numerous multinucleate spherules, formed in liquid culture after nutrient exhaustion. Phosphate storage
granules are present in the spherules. D, Fruit bodies of Physarum viride on dead plant material. Tracks left
by plasmodia as they migrated to exposed sites suitable for fruit body formation and spore dispersal are
visible. (A-C from Carlile M. J. (1971). Myxomycetes and other slime moulds. In Booth, C., ed., Methods in
Microbiology vol. 4, pp. 237-265. Academic Press, London. D, John and Irene Palmer.)

mate with other strains to produce diploid plasmodia. Studies on other

Myxomycetes have demonstrated self-fertile (homothallic) species, in which
mating occurs within a clone. The sexual behaviour of Myxomycetes is thus very
flexible. Not only are self-sterile, self-fertile and apomictic species known but all
three forms of behaviour have been found among strains of a single species,
Didymium iridis.
 A zygote grows and undergoes mitosis without cell division occurring. Since
nuclear division is synchronous, the young plasmodium has in turn 2, 4, 8 and 16
nuclei. Uncountably large numbers are soon reached and plasmodia may cover
many square centimetres and contain millions of nuclei. Plasmodial nuclear
divisions differ from those of amoebae in that the nuclear membrane remains
intact. Perhaps in a multinucleate cell the open type of mitosis might risk 'mix-
ups' between nuclei. Plasmodia increase in size by fusion with each other as well
as by growth. As the size of plasmodia increases protoplasmic streaming becomes
more pronounced, and ultimately develops into the shuttle streaming in well-
defined channels ('veins') characteristic of the mature plasmodium. Observation
of a 'vein' with the microscope shows a torrent of protoplasm moving in one
direction at speeds of up to 1 mm s-1 for about a minute, after which flow occurs
in the opposite direction for about a minute. Presumably it is this shuttle
streaming in the branching pattern of veins, partly overcoming the limitations of
diffusion as a means of transporting oxygen, nutrients and wastes, which makes
possible the development of a 'cell' as enormous as the plasmodium.
Plasmodia can be grown in pure culture on a soluble medium containing a
suitable carbohydrate (e.g. glucose, starch), nitrogen source (e.g. peptone),
mineral salts and vitamins (thiamine, biotin and haem), either on an agar medium
or in shaken liquid culture. Provided with adequate nutrients a plasmodium
grows, with a doubling time of about 8 hours with respect to mass, and slowly
spreads. If starved, a plasmodium migrates, and is attracted by various nutrients
including many carbohydrates. In nature plasmodia can surround and digest large
fungus fruit bodies, and in the laboratory phagocytosis of smaller organisms can
be demonstrated. Shaken liquid culture, however, shows that plasmodia are
capable of absorbing nutrients and in nature plasmodial nutrition is probably
partly ingestive and partly absorptive. The plasmodium, as a result of its size, is
able to move further and attack far larger organisms than can amoebae.
Plasmodia are surrounded by a mucopolysaccharide slime sheath which probably
gives some protection from desiccation and aids locomotion.
When genetically identical plasmodia meet, they fuse to form a single
plasmodium. If, however, the plasmodia belong to different strains, then fusion
may not occur, or if it does, the nuclei of one strain may be eliminated, sometimes
with considerable destruction of protoplasm. This vegetative incompatibility and
its genetic basis, is considered in more detail later (page 259, Fig. 5.5).
Starvation of plasmodia can have two consequences, depending on whether or
not light is present. In darkness a starved plasmodium becomes a sclerotium,
which consists of numerous spherules, each thick-walled and containing several
nuclei and protoplasm. The sclerotia can survive for some years, but in the
presence of nutrients the spherules germinate and the protoplasts merge to form
a plasmodium. A starved plasmodium in light gives rise to a fruit body containing
spores. Meiosis occurs during sporulation to return the organism to the haploid
state; in apomictic strains there is a pseudomeiosis which does not involve a
reduction in ploidy. Spores are capable of prolonged survival and are readily
dispersed, and if they arrive at a suitable site germinate to release amoebae.
The remarkable properties of the Myxomycete plasmodium have been
extensively exploited in fundamental biological research. The synchronous
division of millions of nuclei provides unequalled opportunities for studying
DNA, RNA and protein changes throughout the nuclear cycle, and the control of
mitosis. The rapid protoplasmic streaming has led to investigations on the
molecular basis of cell motility, and the complex life cycle to a variety of studies
on genetics and developmental biology.

The Oomycetes

The group of organisms informally known as Oomycetes are now recognized as

constituting a phylum, the Oomycota, in the kingdom Chromista. A remarkable
product of parallel evolution, they so resemble the true fungi in structure and life
style that they have always been studied by mycologists and were until recently
regarded as fungi. About 700 species are known. Their sexual phase has a clear
differentiation into large female and small male structures, termed oogonia (sing.
oogonium) and antheridia (sing. antheridium), respectively. Within an oogonium
meiosis occurs and, depending upon the species, one or a few oospheres

Figure 2.7 Electron micrographs (shadow cast) of zoospores of the Oomycete

Phytophthora palmivora. A, Zoospore with a short anterior flagellum bearing
mastigonemes (stiff lateral hairs) and a long, nearly smooth posterior flagellum. B, Details
of flagella. The anterior bears prominent mastigonemes and the posterior has just
perceptible fine hairs. (Reproduced with permission from Desjardins, P. R., Zentmyer,
G. A. & Reynolds, D. A. (1969). Electron microscopic observations of the flagellar hairs of
Phytophtbora palmivora zoospores. CanadianJournal of Botany 47, 1077-1079).
Table 2.3 Important differences between the Oomycetes and Fungi
Oomycetes The Fungi
Zoospores Biflagellate; an anterior tinsel Uniflagellate; a posterior
and a posterior smooth smooth flagellum in the
flagellum Chytridiomycetes
Lysine biosynthesis Via diaminopimelic acid Via ~-aminoadipic acid
Mitochondria Cristae tubular Cristae plate-like
Wall polysaccharides Cellulosepresent; chitin also in No cellulose; chitin usually
some species present
Wall proteins Hydroxyproline present Proline present

(unfertilized eggs) are produced. Each contains, when mature, a single haploid
nucleus. Meiosis also occurs in antheridia, and the antheridia grow towards
oogonia. From the antheridia fertilization tubes penetrate oogonia, and a single
fertilization tube enters each oosphere. A single haploid nucleus passes from the
antheridium through the fertilization tube, and fuses with the haploid nucleus in
the oosphere. The oosphere then develops into the oospore (fertilized egg)
characteristic of the class. Each oospore has a single diploid nucleus. When the
oospore germinates it gives rise to a mycelium that is diploid, in contrast to the
haploid mycelium of most other fungi.
Another characteristic of the Oomycetes that distinguishes them from the Fungi
is the biflageUate zoospore (Figs. 2.7, 4.15). This has a forwardly directed tinsel
flagellum - a flagellum with a row of hairs on each side when viewed with the
electron m i c r o s c o p e - as well as a posteriorly directed smooth flagellum.
Ultrastructural and biochemical studies have shown that the Oomycetes differ
from the Fungi in further fundamental ways (Table 2.3).
Three Oomycete orders, the Saprolegniales, Pythiales and Peronosporales, have
attracted considerable attention, and will be considered further.

The Saprolegniales (Water Moulds)

Members of the order Saprolegniales, often referred to as the water moulds, are
common in fresh water and also occur in damp soil. The life cycle (Fig. 2.8) of the
common genus Saprolegnia will be described, along with features of interest from
other genera.
Saprolegnia is common in soil and fresh water. Many species are saprotrophic,
living on animal and plant remains, but a few are parasites, killing fish and
destroying fish eggs. Hyphae of Saprolegnia are sometimes seen emerging from
Figure 2.8 The life cycle of the Oomycete Saprolegnia ferax. When the diploid (2n)
mycelium ceases growth through nutrient exhaustion, septa form delimiting sporangia at
hyphal tips. A sporangium is shown just prior to the breakdown of its tip and the discharge
of zoospores. Both primary and secondary zoospores have a smooth and a tinsel flagellum,
but the point of insertion differs. Secondary cysts have hooks which probably facilitate
attachment to suitable surfaces. Meiosis occurs in antheridia and oogonia to give haploid
(n) nuclei, and oospheres are delimited within oogonia. Antheridia grow on to the surface
of oogonia, and produce fertilization tubes that penetrate oogonia and fuse with
oospheres. Here a single fertilization tube is shown contributing a nucleus to an oosphere.
Fusion between antheridial and oosphere nuclei results in the formation of diploid (2n)
oospores.

sick or dead goldfish in aquaria. Crude cultures of Saprolegnia and other water
moulds can be obtained by placing boiled split hemp seeds, ants' 'eggs' or other
forms of 'bait' in pond water samples; the bait soon supports growth of a water
mould. After contaminating bacteria are eliminated, most water moulds grow
readily in pure culture on common liquid or agar media, such as glucose-yeast
extract-peptone-mineral salts agar. Water moulds cannot reduce sulphate, so in
defined media sulphur is best supplied in a reduced organic form such as
methionine. Glucose is a good carbon source, and nitrogen can be supplied as an
ammonium salt or as amino acids. Vitamins need not be supplied.
The hyphae of Saprolegnia are, compared with most fungi, large and extend
rapidly. They taper towards the tip, and the maximum width is only attained at
some distance behind the tip. The hyphae lack cross-walls, as do those of other
Oomycetes, and nuclei are scattered through the cytoplasm. Prominent
protoplasmic streaming in the direction of the tip occurs. The number of branches
depends on nutrition, high nutrient concentrations increasing branch frequency.
Old cultures may develop resting structures termed gemmae (sing. gemma) which
germinate under suitable conditions.
Starvation conditions can initiate sporangium production, and ultimately
release of zoospores. In the laboratory this can be achieved by replacing a liquid
nutrient medium by distilled water or a dilute salts medium; after sporangia have
developed zoospore discharge can often be brought about by a further
replacement of the distilled water or salts medium. A sporangium is a swollen
hyphal tip, separated from the rest of the hypha by a cross-wall; zoospores form
inside the sporangium. Zoospores when first discharged from Saprolegnia
sporangia are pear-shaped with flagella emerging at the front. These primary
zoospores swim rather sluggishly and soon develop thin cell walls to become
spherical primary cysts. The cysts germinate to release kidney-shaped secondary
zoospores with laterally emerging flagella. These swim much more vigorously and
may form secondary cysts only after some hours. Secondary zoospores show a
chemotaxis towards salts which is enhanced by the presence of amino acids.
Perhaps the role of the primary zoospore is to swim away from the sporangium
and the static water adjacent to the fungus, so that the primary cyst may be
dispersed by currents. The chemotaxis of the secondary zoospore may assist in
reaching hosts or nutrient materials and it appears that the secondary cysts,
copiously provided with spines and hooks as revealed by electron microscopy,
may be well adapted for attachment to a host. Secondary cysts normally
germinate by producing a slender hypha, the germ-tube, which shows
chemotropism (oriented growth) towards amino acids, facilitating penetration of
nutritious substrata.
There is much variation in the Saprolegniales, both within and between species
and genera, in the pattern of zoospore activity and encystment indicated above.
In Saprolegnia the secondary cyst may sometimes germinate to produce a
zoospore instead of a germ-tube. Such a 'tertiary' zoospore resembles the
secondary zoospore in form and activity. The primary zoospores of Achlya
commonly encyst immediately on discharge, forming at the exit pore of the
sporangium a cluster of cysts from which secondary zoospores emerge. In
Thraustotheca the primary zoospores encyst within the sporangium and are
released by sporangium rupture. In Dictyuchus the primary zoospore phase is
suppressed; the developing zoospores encyst within the sporangium and the cysts
germinate to discharge secondary zoospores, each through a separate pore in the
sporangium wall. In Pythiopsis it is the secondary zoospore that is suppressed, the
primary cyst germinating to give a germ-tube. Some fungi, known to be
Saprolegniales from their sexual morphology, do not produce sporangia, and
others produce within their sporangia non-motile spores that have presumably
evolved from cysts. The motile phase of the zoospore life cycle is clearly one that
shows flexibility within a species according to requirements, and in the course of
evolution is eliminated or modified in response to natural selection.
Most water moulds are hermaphrodite or monoecious, bearing both male and
female reproductive structures on a single diploid mycelium. Oogonia are usually
sited singly, often at the tips of hyphal branches, and depending on species may
contain one or a few oospheres. Antheridia may develop on the same hypha as an
oogonium and close to it, or on other hyphae. Monoecious species are self-fertile
(homothallic). The oospores that develop from fertilized oospheres are thick
walled and can survive for long periods. They germinate by means of a germ-tube,
which may develop into a mycelium, or may terminate in a small sporangium
from which zoospores are discharged.
There are a few species that are self-sterile (heterothallic), with the cooperation
of two mycelia being needed for the sexual process to occur. When two
compatible mycelia are brought together, one produces male, and one female
structures. Such species are referred to as dioecious. The dioecious condition
facilitates research on the sexual process, and two dioecious species, Acblya
bisexualis and Acblya ambisexualis, have been employed in such studies. In these
species mycelium that has arisen from a single oospore will not form sexual
structures. If, however, culture filtrate from a 'female' strain is added to a 'male'
strain, antheridial initials (incipient antheridia) develop. This is due to a steroid
sex hormone, hormone A or antheridiol (Fig. 2.9A) which is produced by the
female strain and is active at very low concentrations. When antheridial initials
have been induced in the male strain, it in turn releases a steroid sex hormone,
hormone B or oogoniol (Fig. 2.9B), which induces oogonial initials in the female
strain. Both antheridiol and oogoniol are synthesized by the fungus from
fucosterol, the most abundant sterol in Acblya and other Oomycetes. Antheridial
initials grow towards oogonia by chemotropism up a concentration gradient of
antheridiol. At one time the occurrence of hormones additional to hormones A
and B and acting later in the sexual process was postulated; the occurrence of
such hormones is now thought unlikely. The distinction between male and female
strains is not absolute. A comparison of many strains demonstrates the existence
not only of 'strong males' that act only as males, and 'strong females' that act only
as females, but also of intermediate weak male and weak female strains. Such
strains will act as male when paired with a strong female, or female when paired
with a strong male. The physiological basis for this series of strains seems to be in
the amounts of hormone A produced, with a strong female producing most
hormone A and a strong male the least. Relative sensitivity to hormone A may
also be important.
Monoecious species of Acblya and Tbraustotbeca have been shown to respond
to hormones from Acblya ambisexualis and A. bisexualis, and to produce factors
initiating antheridium development in dioecious species. It is likely, therefore,
that sexual development in monoecious species is controlled by the same (or
similar) hormones as in dioecious species. Under some conditions of nutrition and
temperature a normally dioecious species may develop both oogonia and
antheridia on a single mycelium. It seems likely that monoecious and dioecious
species are closely related and that the evolution of one type from the other could
 OH A .
I
I

o./
o

HO 22 o

OH
HO

RO o
Figure 2.9 The sexual hormones of the Oomycte Achlya. A, Antheridiol ( C 2 9 H 4 2 0 5 9
MW 470) is a sterol produced by the female strain. It induces antheridial initials in the
male strain at concentrations as low as 2 • 10-ll M. B, The sterol oogoniol and its esters
(R = H, acetate, propionate or isobutyrate residues) are produced by a male strain that has
developed antheridial initials in response to antheridiol. The major component of the
hormone mixture is the isobutyrate ester (C33Hs406: MW 546). Oogoniol and its esters
induce oogonial initials in the female strain. They are less potent than antheridiol, with the
most active component of the mixture inducing sexual development in the female at
concentrations of ca 10.7 M.

readily occur. Although good progress has been made in understanding hormonal
and physiological aspects of sexuality in water moulds, the genetic basis of their
sexuality remains to be clarified.

The Pythiales

One of the families in the order Pythiales, the Pythiaceae, includes two of the best
known fungal genera, Pythium and Phytophthora.
Species of Pythium occur in flesh water and soil and can live as saprotrophs on
plant debris, but some species under suitable conditions will attack plants, and
are hence facultative parasites. Pythium ultimum and Pythium debaryanum can
attack a wide variety of seedlings if they are overcrowded and under too moist
conditions, causing 'damping-off' and extensive death. The hyphae of Pythium in
their invasion of plant tissues are both intercellular and intracellular, passing
between and penetrating through plant cells, and pectic enzymes diffuse ahead of
the hyphae, dissolving the middle lamella of cell walls and softening the tissues.
The life cycle of Pythium is similar to that of Saprolegnia. Sporangia are formed,
commonly at the tips of hyphal branches, and zoospore maturation occurs in a
thin-walled vesicle which emerges from the sporangium under appropriate
conditions. The zoospores resemble the secondary zoospores of Saprolegnia and
in some species have been shown to be attracted by root exudates. After a period
of swimming, or on arrival at a suitable substratum, encystment and germ-tube
emergence occurs. Many Pythium species are self-fertile, forming antheridia and
oogonia and successfully producing oospores in cultures that have been derived
from a single zoospore. Others, however, tend to be self-sterile. In Pythium
sylvaticum there are predominantly male and predominantly female strains and
the sexual process occurs most readily when the two types are brought together,
although most of the male and a few of the female strains can if necessary be self-
fertile.
Phytophthora (Greek: phyton, plant; phthora, destroyer) is a genus of
outstanding importance to plant pathologists, killing or otherwise damaging a
wide variety of economically important plants. There are some aquatic species,
but it is the terrestrial species that are of economic importance. Many species can
attack a broad range of hosts. Phytophthora cinnamomi, for example, can infect
many woody plants, including conifers, and in recent years has devastated the
native eucalypt forests of Australia. At the other extreme Phytophthora infestans
is limited to the Solanaceae (the potato and tomato family) and is best known as
the cause of late blight of potato, a disease responsible for a famine in Ireland in
the 1840s and still of economic importance. Phytophthora hyphae differ in their
behaviour from those of Pythium in that they are intercellular, passing between
plant cells but producing short specialized branches, haustoria, which penetrate
the cells. The sporangia of Phytophthora are borne on specialized branched
hyphae, the sporangiophores, which typically emerge from the infected plant and
extend ca 200 pm into the air. This facilitates the dispersal of the sporangia which
are detachable in terrestrial species when mature. Such sporangia may behave as
conidia and produce a germ-tube (direct germination) or may release zoospores
(indirect germination). Which occurs depends on environmental conditions, low
temperatures favouring zoospore release. Thus in P. infestans zoospores are
produced at 15~ and germ-tubes at 20~ behaviour which is understandable as
at the lower temperature, free w a t e r - from dew or rainfall - in which zoospores
can swim will persist longer. Chemotaxis of zoospores to plant exudates has been
shown in several Phytophthora species. In some species the sexual phase can be
produced in cultures derived from single zoospores whereas other species are
normally self-sterile and it is necessary to bring together two mating types,
designated A~ and A z. Oospores are thick-walled and probably are capable of
prolonged survival in soil at times when there are no suitable hosts for infection.
Mycelium too has some capacity for survival on plant debris.
Pythium and Phytophthora, like members of the Saprolegniales, can be grown
in pure culture, although they tend to be nutritionally more demanding. A
remarkable feature of the Pythiaceae, and one in which they are apparently
unique among eukaryotic organisms, is that although they cannot synthesize
sterols, vegetative growth can occur in the absence of added sterols. There is,
however, a requirement for sterols for both sporangium production and the
sexual process. Sterols are thought to endow plasma membranes with greater
strength; perhaps this is essential for the zoospore membrane but not for the
membranes of hyphal cells that are protected by a tough cell wall. The role of
sterols in the sexual process may be as precursors for sex hormones similar to
those known in Acblya.

The Peronosporales

This order includes two families, the Peronosporaceae (downy mildews) and the
Albuginaceae (white rusts), that are obligate parasites of vascular plants.
Examples of downy mildews are Plasmopara viticola, which causes an important
disease of the grape vine, and Peronospora parasitica, which attacks members of
the Cruciferae (the cabbage family). The white rust Albugo candida also attacks
Cruciferae. The downy mildews and white rusts are biotrophs, obtaining their
nourishment only from living cells, in contrast to the Pythiaceae, which are
usually necrotrophs, killing cells and then obtaining nourishment from them.
Although downy mildews and white rusts cannot be grown in pure culture, some
species have been grown in association with cultured cells of the relevant host
plant. Zoospores remain of importance in some species, but in others, such as
P. parasitica, the sporangia function solely as conidia.
The Peronosporales can be arranged as a series with respect to their parasitic
activity. Pythium has considerable saprotrophic ability, and as a parasite has a
wide host range and causes rapid death. Phytophthora has little saprotrophic
ability, but as a parasite has a greater degree of host specialization and some
capacity for obtaining nourishment from living cells. Finally, the downy mildews
and white rusts are obligately biotrophic parasites, usually with a limited host
range and rarely killing host plants. A parallel series can be traced from aquatic
to fully terrestrial species. These series probably also reflect an evolutionary trend
from aquatic saprotrophs, through facultative necrotrophs with a considerable
dependence on damp conditions, to fully terrestrial biotrophs.

The Chytridiomycetes

The Chytridiomycota, more informally the Chytridiomycetes, are a phylum in the

Kingdom Fungi. The main feature that occurs in Chytridiomycetes, and is absent
from the other fungi that remain to be considered, is the zoospore, which has a
single smooth posterior flagellum. This could well be a primitive feature of the
fungi, lost in the groups that adopted a more terrestrial life style.
The Chytridiomycota, of which about 800 species are known, are classified
into five orders. Three of these orders, the Blastocladiales, the Chytridiales
(chytrids) and the Neocallimastigales (anaerobic rumen fungi) will be considered
further.
The Blastocladiales

Members of the order Blastocladiales are mainly saprotrophs living on plant or

animal debris in fresh water, mud or soil. Most produce true mycelium, which
branches dichotomously and is divided into compartments by occasional
partitions, pseudosepta, differing in chemical composition from the hyphal wall.
The sexual process consists of the fusion of a pair of uninucleate gametes, both of
which are motile. Two genera, Allomyces and Blastocladiella, which have been
used extensively in fundamental research, are considered in more detail below.
Another genus, Coelomomyces, which will not be considered in detail, is an
obligate parasite of arthropods. In the course of its life cycle it has to infect in turn
mosquito larvae and adult copepods; it may thus prove useful for the biological
control of mosquitoes and hence malaria.
Allomyces macrogynus, which occurs on organic debris in ponds and in soil, is
readily grown in agar or liquid culture on a yeast-peptone-soluble starch medium
or on suitable defined media. The life cycle (Figs. 2.10, 2.11C) has both haploid
and diploid mycelial phases. The latter is a convenient point at which to begin a
description of the life cycle. The diploid mycelium is often termed the sporophyte
('spore-producing plant'), since when nutrients are exhausted it develops
sporangia. Zoospores are formed within the sporangia, which hence may be
termed zoosporangia. The zoosporangia are of two types. In one type, the thin-
walled mitosporangia, all nuclear division is by mitosis, and diploid zoospores
(mitospores) are produced. Zoospore release can be obtained by immersing
sporangia in pond water or in very dilute mineral salts solution- distilled water
is harmful to the zoospores, which are not protected by a cell wall. The zoospores,
which swim by means of a single posterior flagellum, have a nuclear cap,
consisting of a store of ribosomes enclosed within a membrane. This feature,
confined to the Blastocladiales, presumably facilitates prompt protein synthesis
and rapid growth when germination occurs. The zoospores show chemotaxis to
amino acids, and on reaching a suitable substratum immediately encyst and
germinate. Germination is bipolar. A rhizoid emerges, penetrates the substratum
and then repeatedly branches giving finer rhizoids. Then from the opposite side of
the cyst a stout hypha emerges and by repeated dichotomous branching produces
a new sporophyte mycelium.
The other type of zoosporangia produced by the sporophyte mycelium is
thick-walled meiosporangia. Within these meiosporangia, which remain dormant
for some weeks, meiosis occurs, and when germination takes place, haploid
zoospores (meiospores) are released. These behave similarly to mitospores, but
following encystment give rise to haploid mycelium which is referred to as the
gametophyte ('gamete-producing plant') because, on nutrient exhaustion, it gives
rise to gametangia within which gametes develop. The male gametangia occur at
the end of hyphae, with the female gametangia immediately below, whereas in
another species, Allomyces arbusculus, the opposite arrangement occurs. The
female gametangia are colourless, and the female gametes (Fig. 2.11C) resemble
zoospores in size and form. The male gametangia and the male gametes, which
are smaller than the female gametes, are bright orange with T-carotene. Exposure
to pond water or very dilute mineral salt solutions brings about gamete discharge.
Figure 2.10 The life cycle of the Chytridiomycete Allomyces macrogynus. The diploid
(2n), sporophyte mycelium branches dichotomously, and on nutrient exhaustion bears
thin-walled sporangia (mitosporangia) one of which is shown at the beginning of
zoospore discharge. Zoospores are propelled by a single posterior flagellum, and after a
period of motility form cysts. These germinate to give further diploid sporophytes. The
sporophyte mycelium bears thick-walled resting sporangia (meiosporangia), as well as
thin-walled sporangia. A detached meiosporangium, with its ornamented, nearly opaque
wall is shown. Meiosis occurs in the meiosporangium. The zoospores and cysts of the
haploid (n) phase closely resemble those of the diploid phase. Haploid mycelium, however,
bears male and female gametangia. The female gametangium shown has begun to
discharge female gametes, but the papilla on the male gametangium which normally
discharges first is still unopened. The large female and small male gametes are similar to
each other and to zoospores, and fuse to give motile zygotes. These, after encystment,
germinate to produce diploid mycelium.
Figure 2.11 Stages in the life cycle of
Chytridiomycetes. A, Phase contrast photomicro-
graph of a zoosporangium (width ca 100 lam) of
an anaerobic rumen Chytridomycete, Neo-
callimastix sp., isolated from faeces of a
Malaysian water buffalo. B, The same zoo-
sporangium viewed by fluorescence microscopy
after treating with a stain (DAPI) specific for
nuclei. The nuclei are confined to the zoo-
sporangium and are absent from the extensive
rhizomycelium. C, Electron micrograph of a
female gamete of Allomyces macrogynus. The
nucleus contains a single electron-dense nucleolus,
and is surrounded by a massive nuclear cap,
consisting of ribosomes. The cytoplasm contains
lipid droplets (spherical, grey) and mitochondria.
The base of the single posterior flagellum is visible
below the nucleus. The zoospores of Allomyces
closely resemble the female gamete. Scale bar,
1 lam. (A, B from Trinci, A. P. J., Davies, D. R.,
Gull, K., Lawrence, M. I., Nielsen, B. B., Rickers,
A. & Theodorou, M. K. (1994). Anaerobic fungi
in herbivorous animals. Mycological Research 98:
129-152. C, reproduced with permission from
Pommerville, J. & Fuller, M. S. (1976). The
cytology of the gametes and fertilization of
Allomyces macrogynus. Archives of Microbiology
109, 21-30.)
The male gametes, which are the first to be released, swarm around the female
gametangia. This is due to the sex attractant, sirenin (Fig. 2.12), which is released
by the female gametangia and gametes and which is effective at attracting male
gametes over a wide range of concentrations. Sirenin was named after the sirens,
females of classical Greek mythology having an attractant power over navigators.
The male gametes fertilize the female gametes as they emerge from the
gametangia, nuclear fusion rapidly following cell fusion. The zygote formed by
fertilization retains the flagella from both gametes and is hence the sole
biflagellate phase in the life cycle. The zygote differs from the male gamete and
resembles zoospores in being insensitive to sirenin but being attracted by amino
acids. On arrival at a suitable substratum it encysts and germinates to give rise to
the sporophyte mycelium.

CH2OH

CH2OH

Figure 2.12 Sirenin, the attractant released by female gametangia and gametes of the
Chytridiomycete Allomyces, and to which the male gametes respond. It is a bicyclic
sesquiterpene (ClsH2402:MW 236). It is active over a wide range of concentrations, from
10-l~ to 10-s M. In addition to being highly active, it is highly specific, most analogues and
the D-isomer being without activity. Sirenin is destroyed by the male gametes after it is
taken up. There is also evidence for a complementary sex hormone, parisin, released by
male gametes and attracting female gametes.

A. macrogynus, which has a basic haploid chromosome number of 14, and

A. arbusculus which has one of 8, are both hermaphrodite but self-fertile. There
are polyploid strains in both species. The two species can hybridize, and a third
species, Allomyces javanicus, is a naturally occurring hybrid between them.
Artificial hybrids between A. macrogynus and A. arbusculus have been produced,
and include some strains that are effectively female (i.e. almost wholly lack male
gametangia) and others that are effectively male. The isolation of sirenin was
achieved by using a female hybrid for sirenin production, and a male, yielding
only male gametes, for assay.
Zoospores of Blastocladiella emersonii (Fig. 4.15) closely resemble those of
Allomyces. On arriving at a suitable surface, a zoospore encysts, and soon
produces a rhizoid that penetrates the substratum and repeatedly branches.
Synthesis of cell constituents and repeated nuclear division occur, and what had
been the cyst enlarges and becomes an ovoid thallus, from which more rhizoids
emerge. The thallus then usually differentiates into a thin-walled zoosporangium
from which zoospores are discharged. Under optimal conditions the entire cycle
takes about 20 hours, during which time up to eight successive nuclear divisions
have occurred to yield about 256 (i.e. 28) zoospores. This cycle has been the
subject of extensive physiological and biochemical research. The thallus can also
develop into a thick-walled zoosporangium which is capable of prolonged
survival under adverse conditions. This is often termed the RS or resistant
sporangium. Such sporangia develop in the presence of high bicarbonate
concentrations, produced naturally by overcrowding and high carbon dioxide
production. Within resistant sporangia synaptonemal complexes, indicative of
meiosis, have been demonstrated by electron microscopy, and the zoospores
released from RS sporangia have been found to have half the DNA content of
those from the thin-walled OC (ordinary colourless) sporangia. This suggests that
the intensively studied cycle involving OC sporangia corresponds to the diploid
cycle in Allomyces, but the significance of the haploid zoospores is less clear. Two
other types of zoosporangia have been observed in B. emersonii, one of which is
orange from the presence of y-carotene. Perhaps this corresponds to the male
gametangium of Allomyces. It is surprising that the life cycle of B. emersonii,
parts of which have been so intensively studied, has not yet been elucidated fully.
The life cycle of another species, Blastocladiella variabilis, has been fully
elucidated, and in this sexuality has been clearly demonstrated, with two types of
gametangia, discharging gametes which although of equal size, are orange and
colourless, respectively.

The Chytridiales

Members of the order Chytridiales, often referred to as chytrids, lack a true

mycelium but may produce slender rhizoids penetrating the substratum or very
slender hyphae (the rhizomycelium) linking sporangia. They are mainly aquatic,
with some species saprotrophs on plant or animal debris, and others parasites on
algae or small aquatic animals. There are also species that live in soil. Some of
these are saprotrophs, but others are parasites of vascular plants, in a few
instances causing economically important diseases. The saprotrophic aquatic
species can be obtained in crude culture by baiting water samples with suitable
substrates such as cellulose (e.g. boiled grass leaves), chitin (e.g. shrimp skeletons)
or keratin (e.g. hair or moulted snake skin!). Many of the saprotrophic species
have been grown in pure culture but those attacking living organisms are obligate
parasites. The Chytridiales include a wide diversity of forms, and the more readily
handled species should provide interesting material for future physiological,
biochemical, genetical and ecological research.

The Anaerobic Rumen Fungi

Ruminant animals, such as sheep and cattle, are highly effective consumers of
plant biomass. This effectiveness is due to the activity of microorganisms in the
rumen, a specialized region of the gut. Conditions in the rumen are virtually
anaerobic, and the study of rumen microbes requires stringent exclusion of
oxygen, both in the sampling and processing of rumen contents and in the
subsequent culture of the rumen organisms. Obligately anaerobic bacteria and
protozoa have long been known to be abundant in the rumen, but the presence of
fungi was unsuspected until 1974. It was then realized that some organisms which
had been regarded as protozoa were in fact Chytridiomycete zoospores. Attention
which had hitherto been directed towards the liquid from strained rumen
contents was then switched to the residual plant debris, where abundant
obligately anaerobic Chytridiomycetes were found.
The anaerobic rumen fungi are now regarded as constituting the order
Neocallimastigales. The life cycles of several rumen fungi have been studied, and
that of one species, Neocallimastix hurleyensis, followed in axenic culture.
Zoospores of Neocallimastix are unusual in being multiflagellate, with 8-17
flagella. Zoospores are attracted to, and encyst upon, fragments of plant material.
Cyst germination follows, with the germ-tube penetrating the substratum to
become a rhizoid that branches profusely. As with Blastocladiella (page 36) a
thallus develops and becomes a zoosporangium (Fig. 2.11A), which in N.
hurleyensis yields an average of 88 zoospores. When this fungus is grown at 39~
the temperature of the rumen, about 30 hours elapse between a zoospore
encysting and the release of further zoospores from the mature sporangium that
develops.
The activities of anaerobic Chytridiomycetes in the herbivore rumen are
considered further in Chapter 7.

The Zygomycetes

The phylum Zygomycota consists of two classes, the Zygomycetes and the
Trichomycetes. Within these two classes the sexual process consists of the fusion
of two gametangia to give a resting spore, the zygospore. Whether the two classes
are closely related is not clear. The Trichomycetes, of which about 200 species are
known, are obligate parasites that live in the gut of insects and other arthropods,
and will not be considered further. There are about 900 species of Zygomycetes,
and members of one order, the Mucorales, are very widespread and abundant.
The Mucorales will now be considered, after which brief reference will be made
to other orders.

The Mucorales

Most members of the Mucorales are saprotrophs, and are common in soil and on
the droppings of rodents and large herbivores. Others cause rots of fruits and
some occur on the decaying fruiting bodies of mushrooms and toadstools. The
saprotrophic members of the Mucorales usually have little ability to attack
refractory substrates such as cellulose or chitin, but have large hyphae that spread
rapidly, and soon sporulate to produce spores that germinate easily. They hence
'get there first' and can exploit readily assimilable nutrients such as sugars before
other fungi arrive, and hence have been termed 'primary saprotrophic sugar
fungi'. Others, including Mucor hiemalis, have been found on rotting wood,
apparently obtaining sugars released from wood by Basidiomycete enzymes, and
hence are 'secondary saprotrophic sugar fungi'. A few members of the Mucorales
are mycoparasites, attacking other fungi, particularly saprotrophic Mucorales
growing on dung. Their hyphae are generally small and many are obligate
parasites and cannot be cultured. The largest genus in the Mucorales is Mucor
itself, many species of which are common in soil and on decaying plant materials.
The life cycle (Fig. 2.13) of Mucor and similar members of the Mucorales will
now be described.
Sporangiospores of the Mucorales may contain one or several nuclei. Such a
spore, placed on a suitable agar medium, will germinate. The spore swells, often
to many times the original volume, and a new inner wall is synthesized beneath
the original spore wall. After a few hours a hypha, the germ-tube, breaks through

Figure 2.13 The life cycle of the Zygomycete Mucor mucedo. Multinucleate
sporangiospores germinate to give mycelia of the same mating type (§ or-). Mycelia bear
sporangia on sporangiophores, as shown on the left. On the right a sporangium in section
shows the columella, sporangiospores, and oxalate crystals on the sporangium surface. If
colonies of different mating type come into proximity, zygophores (stout aerial hyphae)
differing in mating type grow towards each other and form gametangia when they come
into contact. Cell fusion and nuclear fusion occur to give thick-waUed diploid (2n)
zygospores. Meiosis precedes germination, which gives a sporangiophore terminating in a
sporangium containing haploid (n) sporangiospores.
the old spore wall. The cell wall of the germ-tube is continuous with the new inner
wall of the spore. Sometimes several germ-tubes are produced. These hyphae
grow and branch, and within a day a circular colony of vegetative mycelium is
established. The vegetative hyphae spread upon and penetrate the substratum,
and in some species aerial hyphae rising above the substratum are also produced.
Some genera, such as Rhizopus and Absidia, achieve rapid spread by means of
stout rapidly growing aerial hyphae known as stolons. When these touch a
suitable substratum slender hyphae, rhizoids, develop and penetrate the
substratum. The hyphae of the Mucorales are coenocytic with many nuclei and
few cross-walls.
Some members of the Mucorales are capable of growth under anaerobic
conditions (page 147). In these circumstances metabolism is fermentative and
alcohol is produced from sugar. In the absence of air and in the presence of high
carbon dioxide concentrations sporangiospores of some Mucor species, such as
Mucor rouxii and Mucor racemosus, germinate by budding, and a form
resembling yeast is produced. This mould-yeast dimorphism, controlled by
environmental conditions, has been studied in Mucor and Mycotypha. Some
Rbizopus species, such as Rhizopus oryzae, will grow under anaerobic conditions
but remain hyphal. Some other members of the Mucorales, such as Phycomyces
blakesleeanus, are incapable of anaerobic growth.
The most widespread mode of asexual sporulation in the Mucorales is the
production of sporangiospores. The details of sporangium (pl. sporangia) form and
their arrangement on the erect hyphae that bear them, the sporangiophores, vary
and are used in defining genera and species. Detailed studies of sporangiophore
growth have been carried out in a genus, Pbycomyces, in which the sporangiophores
- and also the mycelial hyphae, sporangia and zygospores- are very large. The
Pbycomyces sporangiophore is at first merely a stout hypha rising above the
substratum and elongating by means of wall extension in the apical region. Then
elongation ceases and the apex swells to form a spherical sporangium. The sporangium
is at first bright yellow from the presence of [5-carotene, but then turns black as the
carotene undergoes oxidative polymerization to form sporopollenin, a substance very
resistant to chemical and biological degradation. Within the sporangium the
protoplasm is cleaved and rounds off to give about 100 000 sporangiospores, each
containing a few nuclei. The walls of the sporangiospores contain sporopollenin.
Most of the sporangium is occupied by sporangiospores, but the sporangiophore
projects into the sporangium as a columella. After the sporangium is formed,
elongation of the sporangiophore is resumed, with extension occurring in a growth
zone just below the sporangium. The sporangiophore, both in the early and late
phases of elongation, displays a range of sensory responses. It shows positive
phototropism (Fig. 4.5), growing towards the light over a very wide range of light
intensities. Other sensory responses that enable the sporangiophore to rise above the
substratum (in nature usually dung) while avoiding obstacles are negative geotropism
(upward growth), negative hydrotropism (growth towards dry conditions), an
avoidance response when approaching obstacles including other sporangiophores,
and growth into air currents (positive anemotropism). The sporangial wall of
Phycomyces is thin, and readily ruptured when the sporangium is mature, to expose
the spores which are contained in a mucilaginous matrix. It is probable that such
'slime spores' are dispersed by rain splash.
 Mucor, like Phycomyces, has an approximately spherical sporangium
borne on an erect sporangiophore, but both sporangium and sporangiophore
are much smaller. In some species the sporangium wall dissolves at maturity
leaving a 'stalked spore drop'. The sporangium of Rhizopus resembles that
of Mucor but there is no mucilage in the sporangium; on rupture 'dry
spores' are released which are readily dispersed by air currents. In other
genera of Mucorales there is a wide variety of sporangium form. In
Thamnidium a sporangium similar to that of Mucor occurs at the end of the
sporangiophore, but about halfway up the sporangiophore there are short
branches carrying tiny sporangia (sporangioles) each containing only a few
sporangiospores. Thamnidium may, depending on conditions of light,
temperature and humidity, produce a sporangium only, sporangioles only,
or both. Cunninghamella apparently produces conidia, but developmentally
these, and similar structures in some other genera, appear to be sporangioles
containing a single spore. The sporangiospores of the Mucorales are mostly
classifiable into dry spores, dispersed by air currents, and slime spores,
dispersed by rain splash. The genus Pilobolus, however, members of which
grow on dung, has an active discharge mechanism. The sporangiophores are
phototropic and when mature, the osmotic rupture of a subsporangial
swelling 'shoots' the sporangium a metre or so away from the dung, to
alight on vegetation and perhaps to be consumed by a herbivore and thus to
arrive in flesh dung.
Some members of the Mucorales produce, in addition to sporangiospores, a
further type of asexual spore, the chlamydospore. These thick-walled spores are
produced within hyphae and have no dispersal mechanism, so probably they
remain at the site of production until conditions favourable for growth recur.
Mucor racemosus produces copious chlamydospores both in vegetative hyphae
and within sporangiophores. The sexual process has received detailed study in
Mucor mucedo. When vegetative growth brings two colonies that differ in mating
type into close proximity, both produce zygophores, specialized aerial hyphae.
Zygophores of differing mating type grow towards each other through the air, a
form of positive autotropism termed zygotropism and effective over several
millimetres. When two zygophores of different mating type come into contact the
walls firmly fuse to each other and zygophore elongation ceases. The two
zygophores then swell in the region immediately adjacent to the area of contact to
give two multinucleate progametangia. Each progametangium develops into a
gametangium by the production of a cross-waU which delimits it from the
adjacent region of the zygophore which is then termed the suspensor. The cross-
wall separating the two gametangia then breaks down and the fused gametangia
develop into a zygospore. The development of a thick zygospore wall containing
the black pigments melanin and sporopollenin soon renders the observation of
cytological events difficult. Limited cytological observations supplemented by
genetic analysis suggest that nuclei of different mating type pair and fuse, and that
unpaired nuclei degenerate. Meiosis occurs, but in M. mucedo only one of the
four recombinant types from a single diploid nucleus survives, and subsequently
multiplies. Zygospores do not readily germinate. In M. hiemalis there is no
germination for 30 days, and 1% of the zygospores germinate in the following 90
days. A sporangiophore emerges from the zygospore and terminates in a
sporangium. In M. hiemalis and M. mucedo all the resulting sporangiospores,
representing one of the four products of a single meiosis, are of the same mating
type. Many members of the Mucorales other than M. mucedo are self-sterile, with
two mating types designated plus (§ and minus (-), respectively. There are,
however, self-fertile species, such as Rhizopus sexualis, in which colonies
developing from a single uninucleate sporangiospore will produce zygospores.
Other details of the sexual process also vary.
In Phycomyces blakesleeanus parental haploid nuclei degenerate, and usually
only a single diploid nucleus survives and undergoes meiosis, to yield four
recombinant haploid nuclei which then multiply. Sometimes, however, more than
one diploid nucleus undergoes meiosis and hence a larger variety of recombinant
types are produced. Up to 80% of the zygospores have been shown to germinate,
from 80 to 120 days after their production.

9 0

OH

Figure 2.14 The main zygophore-inducing hormone in Mucor mucedo, trisporic acid C
(C18H2604:MW 306). It is active at about 10-8 M. M. mucedo also produces small amounts
of a second zygophore-inducer, trisporic acid B, which carries an oxygen instead of a
hydroxyl group.

The hormonal control of the sexual process in Mucor mucedo has been studied
in detail. It has been found that a mixed culture of a plus and a minus strain will
produce trisporic acid (Fig. 2.14), a substance which is not produced by either
plus or minus strains when grown alone. Trisporic acid, applied to a plus or
minus strain, induces zygophores. The production of trisporic acid is the result of
a remarkable collaborative biosynthesis involving both plus and minus strains
(Fig. 2.15). The pathway is known in outline although the identity of some
intermediates and steps are uncertain. Both plus and minus strains are able to
cleave f~-carotene to yield retinal. The minus strain can convert this compound by
a series of steps into trisporol, but trisporol can only be converted into trisporic
acid by the plus strain. The plus strain on the other hand, can convert retinal into
4-dehydrotrisporic acid, which can only be converted into trisporic acid by the
minus strain. Thus, the production of trisporic acid is dependent upon the
diffusion of trisporol and 4-dehydrotrisporic acid between plus and minus strains
either through the substratum or through a i r - the two compounds are both water
soluble and volatile. Although trisporic acid is the factor responsible for
zygophore induction it is not volatile and hence cannot account for the attraction
through air of the plus and minus zygophores to each other. This mutual
attraction appears to be due to the diffusion of volatile strain-specific precursors.
Figure 2.15 The collaborative synthesis of trisporic acid by plus (+) and minus (-) strains
of Mucor mucedo. There are two possible routes for the synthesis of trisporic acid from
retinal, the trisporol and the 4-dehydrotrisporic acid pathways. The minus strain has the
enzymes required for the first part of the trisporol and the second part of the
4-dehydrotrisporic acid path. Conversely the plus strain has the enzymes for the first part
of the 4-dehydrotrisporic acid and the second part of the trisporol path. Thus trisporic acid
synthesis is accomplished only when both strains are present.

In addition to inducing zygophores, trisporic acid stimulates the production of

fl-carotene and other intermediates in its own biosynthetic path, thus greatly
increasing its own rate of biosynthesis, an example of positive feedback or
metabolic amplification. Trisporic acid stimulates carotenoid production and
induces zygophores in a wide range of Zygomycetes, including self-fertile species.
It is likely, therefore, that the hormonal control of the sexual process
demonstrated in M. mucedo is widespread and perhaps universal in the
Zygomycetes.

Other Zygomycete Orders

The Mucorales are so widespread and abundant that they are frequently
encountered by almost all mycologists. The other Zygomycete orders are of a
more specialist interest, although in some environments they are of considerable
significance. The Glomales are important since they form a characteristic
mutualistic association with the roots of a wide variety of vascular plants (page
397). Their hyphae penetrate between the cells of roots where they send
haustoria, which branch like trees (arbuscules; from arbor, Latin for tree), into
cells, and in some groups also form swellings (vesicles) inside cells. Such fungi are
hence known as arbuscular or vesicular-arbuscular fungi, and the associations as
arbuscular or vesicular-arbuscular mycorrhizas (mycorrhiza; Greek, fungus-
root). The Zoopagales are predators or parasites of small animals such as
amoebae and nematodes (eelworms). The predatory forms have a sticky
mycelium which traps the prey which is then invaded by hyphae, and the parasites
infect their host by means of spores, which germinate after being ingested by the
host or sticking to its surface. The Entomophthorales include the large genus
Entomophthora (Greek, insect destroyer) whose members infect and kill a wide
variety of insects including house flies (page 428). Some fungi in this group are
used in the biological control of insect pests.

The Ascomycetes
Members of the phylum Ascomycota, commonly referred to as the Ascomycetes,
are those fungi in which the sexual process involves the production of haploid
ascospores through the meiosis of a diploid nucleus in an ascus (pl. asci). Most
Ascomycetes also carry out asexual sporulation, conidiospores (conidia) being

Figure 2.16 Diagrams of Ascomycete fruit bodies. A, Apothecium of Aleuria vesiculosa.

The upper surface is lined with asci and spacer hyphae, paraphyses. B, Asci and paraphyses
of A. vesiculosa (A, B after Buller, A. H. R., Researches on Fungi, vol. 6, Longman, Green,
London, 1934). C, Perithecium of Sordaria fimicola. An ascus is shown protruding from
the opening (ostiole) of the perithecium prior to discharging its ascospores and collapsing.
It will be replaced by a sequence of other asci, shown in various stages of development
(after Ingold, C. T. (1971). Fungal Spores: their Liberation and Dispersal, Clarendon Press,
Oxford): D; Cleistothecium of Eurotium repens. The ascus walls lyse and finally the
perithecium ruptures to release ascospores (after Webster, J. (1980). Introduction to Fungi;
2nd edn, Cambridge University Press, Cambridge).
Figure 2.17 Fruit bodies (ascocarps) of Ascomycetes. A-C,
Diverse forms in the order Pezizales. A, Apothecia of Aleuria
aurantia. Asci line the inside of the cups. B, The stalked
apothecium of the morel, Morchella hortensis. The pits are lined
with asci. C, Excavated fruit bodies of the truffle Tuber
aestivum, one intact and one sectioned. Animals dig up and eat
the fruit bodies, and in so doing, scatter the spores that they
contain. Tuber is thought to have evolved from Pezizales with
more usual fruit bodies. D, Several stalked perithecial stromata
(two in focus) have developed from a buried sclerotium of
Claviceps purpurea. The exits to the perithecia (ostioles) appear
as dots on the heads of the stromata. See also Fig. 8.17. (A-C,
John and Irene Palmer, D, Stephen Shaw and Peter Mantle.)
Further illustrations of fruit bodies are available at
http://www.wisc.edu/botany/fungi/html
produced on specialized aerial hyphae, the conidiophores, that rise above the
substratum. The sexual phase of an Ascomycete is now termed the teleomorph,
and the asexual phase the anamorph, the concept of 'perfect' and 'imperfect'
states having been abandoned. Isolates are often obtained that fail to produce a
teleomorph, but which from their asexual sporulation are clearly identical with a
known Ascomycete. Such anamorphic isolates are assigned a separate Latin
binomial. For example, strains of Eupenicillium brefeldianum that fail to produce
ascocarps are designated Penicillium dodgei. When both anamorphic and
teleomorphic phases are present, the teleomorph name is used.
Many Ascomycetes produce their asci in complex fruiting bodies termed
ascocarps (Figs. 2.16, 2.17). Such Ascomycetes were formerly regarded as
Euascomycetes ('true Ascomycetes'), and classified on the basis of ascocarp form.
The 'Discomycetes', for example, were those which had a disc-shaped ascocarp,
the apothecium, on which asci are exposed. The 'Pyrenomycetes' were those that
produced asci within a flask-shaped ascocarp, the perithecium. The
'Plectomycetes' were those in which the asci developed inside an approximately
spherical ascocarp, the cleistothecium. Ascocarp form is crucial in relation to
spore dispersal. An apothecium is ideal for the discharge of ascospores into the
air, but the asci are ill-protected during development. A perithecium gives some
protection, but limits the rate at which ascospores can be discharged. In a
cleistothecium, the asci are well protected, but can only be released by rupture of
the cleistothecium.
There are a very large number of saprotrophic and parasitic Ascomycetes, at
least 18 000, which have ascocarps. In addition there are many more such
Ascomycetes, perhaps a further 14 000, that have a mutualistic association with
phototrophic microorganisms and constitute the lichens (see page 76). Other
Ascomycetes, formerly termed the Hemiascomycetes ('Half Ascomycetes'), do not
have ascocarps, solitary asci being produced. Such Ascomycetes are not
numerous, but include many important yeasts (page 71).
There has been a major reclassification of the phylum Ascomycota. Grouping
into such classes as the Discomycetes, Pyrenomycetes and Plectomycetes on the
basis of ascocarp form is no longer accepted. This is because it brings together
fungi that on a range of other criteria are dissimilar, and separates ones that are
similar. Instead a wide range of characters, including details of ascus structure,
are being used to assemble species into genera, genera into families, and families
into orders. Some of the orders now recognized contain not only saprotrophic
and parasitic species but also lichens.
A few well-studied Ascomycetes, selected on the basis of importance, or
illustrative of Ascomycete diversity, will now be considered.

The Pezizales: Pyronema, Ascobolus, Morels and Truffles

The Pezizales commonly produce their ascocarps, which are often quite large, on
the surface of forest soil, dead wood or dung. A common species is Pyronema
omphalodes, formerly known as P. confluens, which is found on bonfire sites and
on sterilized soil in greenhouses. It is readily grown in pure culture on defined
media containing a sugar and mineral salts. Ascospores placed on the medium
germinate to produce a rapidly spreading and branching mycelium of large
hyphae. The hyphae show features which are lacking in the 'lower fungi'
discussed in earlier pages but are usual in 'higher f u n g i ' - Basidiomycetes,
Ascomycetes and mitosporic fungi. The hyphae have numerous cross-walls
(septa, sing. septum) but these are perforated by septal pores which permit rapid
protoplasmic streaming and even the passage of nuclei. A hypha may also
undergo fusion (anastomosis) with a neighbouring hypha. Usually each hypha
produces a slender side branch. These grow towards each other and fuse. Hyphal
anastomosis can convert a mycelium of radiating hyphae (as is seen in the lower
fungi) into a three-dimensional network characteristic of the higher fungi. Such a
network can facilitate the transport of protoplasm and nutrients to any point on
a mycelium and facilitate the production of the large fruiting bodies which are
widespread in the higher fungi.
The life cycle of Pyronema is outlined in Fig. 2.18. There is no asexual
sporulation. Induction of sexual sporulation requires carbohydrate limitation.
One procedure is to grow the fungus in a small Petri dish of glucose mineral salts
agar placed in a large Petri dish of water agar. The dense mycelium in the inner
dish spills over to form a thin mycelium with apothecia on the outer dish. This
procedure, which produces copious apothecia, also demonstrates the efficiency
with which nutrients and protoplasm are translocated from the inner dish.
Alternatively both moderate vegetative growth and moderate apothecium
production can be obtained in a single Petri dish by providing the slowly utilized
sugar lactose instead of the readily utilized glucose. Exposure to light is essential
for apothecium production, and apothecia and mycelium produced in daylight
are pink from the presence of carotenoids. It is, however, the ultraviolet
component of daylight that is responsible for apothecium formation and the
visible component for carotenoid induction, so it is possible, by exposure to
suitable ultraviolet radiation, to produce white apothecia.
Pyronema is self-fertile and apothecium production begins with the formation
of clusters of the male structure, the slender antheridium, and the female
structure, the approximately globose ascogonium. Both are multinucleate. The
ascogonium carries a projection, the trichogyne, which grows and curves to make
contact and fuse with an antheridium. Nuclei then pass from the antheridium into
the ascogonium. Ascogenous hyphae grow out from the fertilized ascogonium,
and nuclei enter these hyphae. It is likely that the nuclei pass into the ascogenous
hyphae in pairs, one member of each pair being derived from the antheridium. In
Pyronema there is no proof of this, but in Ascomycetes in which genetic analysis
has been carried out the results of hybridization experiments establish firmly that
pairing must occur. The growth of an ascogenous hypha is terminated by the tip
curving sharply back on itself to form a crozier. The two nuclei nearest the hyphal
tip then divide, thus giving a sequence of four nuclei, two of which will have come
from the antheridium and two from the ascogonium. Septa then form, isolating
one nucleus in a terminal cell, two nuclei, one 'male' and one 'female', in the
penultimate cell, and leaving the fourth nucleus in a stalk cell. Usually the
penultimate cell then elongates to form a tubular ascus. The two haploid nuclei
fuse to give a diploid nucleus which undergoes meiosis to yield four haploid
nuclei. These in turn undergo mitosis to form eight nuclei. Walls develop around
these nuclei and associated cytoplasm to yield eight ascospores.
 Meanwhile the terminal and stalk cells may have fused to bring together a male
and female nucleus to initiate the development of another crozier, and sometimes
penultimate cells will develop into croziers instead of asci. So a cluster of
ascogonia will produce many ascogenous hypha, and an ascogenous hyphae
(which sometimes may branch) can produce many croziers and many asci. The
resulting apothecium will contain a large number of parallel tubular asci
interspersed with other hyphae, the paraphyses, which perhaps can be regarded as
spacers, keeping the asci apart and allowing their optimal orientation. Finally the
asci discharge their ascospores. A small lid, the operculum, at the tip of the ascus
is forced open, and the ascospores are shot into the air where they can be
dispersed by air currents.
An important feature of Ascomycete meiosis is that the tetrad of four nuclei
produced in a single meiosis all survive and for a time are kept together in the
form of four or more, usually eight, ascospores in a single ascus. This permits
tetrad analysis, the isolation and genetic study of all four recombinant types
formed in the meiosis of a single hybrid nucleus. Moreover in Ascomycetes with
narrow tubular asci the nuclei cannot slip past each other so that the four
products of meiosis are represented by four successive pairs of ascospores
arranged along the ascus. These can be dissected out in order and cultured. Such
an ordered tetrad analysis yields information on whether two alleles segregated at
the first or second meiotic division. In those Ascomycetes in which broad asci
allow nuclei to slip past each other, ordered tetrad analysis cannot be carried out.
However, an unordered tetrad analysis in which the eight ascospores are
separately cultured without attempting to keep them in order is less laborious and
still yields much useful genetic information. Pyronema omphatodes has not been
utilized in genetic work, but the potentialities of tetrad analysis have been
exploited in several other Ascomycetes.
Another genus in the Pezizales, the coprophilous Ascobolus, has been the
subject of considerable study. A. crenulatus, formerly termed A. viridutus, like
Pyronema omphalodes is self-fertile. A. immersus, however, which has been
extensively used for genetical research, is self-sterile, and the sexual process only
occurs if the two mating types are brought together. Each strain then produces
ascogonia, and fertilization occurs through hyphae of the other strain acting as

Figure 2.18 The life cycle and stages in ascus production in the Ascomycete Pyronema
omphalodes. Here n + n indicates the association of as yet unfused nuclei from the
antheridium and ascogonium. In self-sterile Ascomycetes, however, n + n represents a
dikaryotic phase, with nuclei of different mating types within the same cell. Stages in ascus
production are illustrated as follows. A, A trichogyne growing from an ascogonium has
just fused with an antheridium. Antheridial nuclei are shown white and ascogonium nuclei
black. B, The contents of the antheridium have passed into the ascogonium and pairing of
nuclei has occurred. C, Ascogenous hyphae are developing at the surface of the
ascogonium. Paired nuclei move into the hyphae and divide repeatedly. D, Crozier
formation at the tip of an ascogenous hypha. E, Nuclear division has occurred in the
crozier. F, Septa have formed in the crozier. G, Nuclear fusion has occurred in the
penultimate cell, initiating ascus development, and the tip cell has fused with the stalk cell.
H, Meiosis has occurred in the developing ascus, and the fused tip and stalk cell have given
rise to a further crozier. I, Mature ascus with eight ascospores. J, Discharged ascus,
showing operculum still attached at ascus tip.
antheridia. The broad asci of A. immersus only allow the analysis of unordered
tetrads (Fig. 5.8C, D). The occurrence of a mutation affecting spore colour,
however, permits the analysis of the contents of very large numbers of asci
without having to culture the ascospores. When mutant and wild-type are
hybridized, the asci produced would be expected to contain four wild-type and
four mutant ascospores. Most asci do, but occasional asci are found with
different frequencies of wild-type and mutant. This results from the process of
gene conversion, which may be a very widespread phenomenon but is most
readily demonstrated in Ascomycetes in which tetrad analysis can be carried out.
A. fufuraceous (formerly called A. stercorarius) is another self-sterile species,
requiring the interaction of the two mating types for the sexual process to occur.
It also produces spores asexually by the fragmentation of aerial hyphae. These
spores, termed oidia or arthrospores (Fig. 2.19A), germinate to give a mycelium
if they fall on a suitable substratum. If they fall on a mycelium of the same mating
type they do not germinate, but on the mycelium of the opposite mating type an
oidium will induce the formation of an ascogonium. The trichogyne of the
ascogonium grows towards the oidium and fuses with it. Apothecium
development follows. The asci of Ascobolus and probably most Pezizales are
phototropic, curving to point in the direction of maximum light intensity and
hence shooting their ascospores in the direction with least obstruction.
The Pezizales also include the morels, members of the genus Morchella
(Fig. 2.17B). Morels grow in woodlands and their fruiting bodies, edible and
much prized, are stalked and up to 15 cm high. Some success has been achieved
both in the field cultivation of morels, and the production of mycelium, but not
fruiting bodies, in pure culture. Other Pezizales are known as cup-fungi, because
the disc bearing the asci is curved into a saucer or even a cup-shape (Fig. 2.17A).
In Pezizales with subterranean fruiting bodies, the truffles (Fig. 2.17C), this
tendency has gone further so that the asci line the interior of a roughly spherical
fruiting body. Here there is no discharge mechanism, and it is likely that spores
are dispersed by rodents which, attracted by the smell of mature fruiting bodies,
dig them up and eat them. A few species, such as Tuber melanosporum, are highly
prized by gourmets and are found with the help of trained dogs or pigs. They
form mycorrhizal associations with the roots of beech and oak, and recently some
success has been achieved in promoting the mycorrhizal association between
T. melanosporum and the oak.

The Sordariales: Neurospora, Podospora, Sordaria and Chaetomium

The Sordariales include one of the most intensively studied of all fungi,
Neurospora crassa. It is readily grown in pure culture and has large hyphae which
spread rapidly. Two types of asexual sporulation occur. The aerial mycelium
rising above the substratum consists largely of branched chains of macroconidia
(Figs. 2.19B, 4.1B), roughly ellipsoidal cells containing several nuclei.
Macroconidia are pink due to the presence of carotenoids, and are produced in
enormous numbers. They are readily detached and dispersed by air currents and
on landing on a suitable substratum germinate readily. In the tropics burnt
vegetation is often festooned with the pink mycelium and macroconidia of
Neurospora, and in the laboratory care is needed to avoid the massive release of
macroconidia with resulting contamination of other cultures when Petri dishes
are opened. Tiny uninucleate microconidia (Figs. 2.19C, 4.1C) are also produced,
but do not germinate so readily as macroconidia and do not survive so long. It is
probable that the role of microconidia is participation in the sexual process.

Figure 2.19 Asexual sporulation in Ascomycetes. A, Formation of arthrospores in

Ascobolus furfuraceus, by the separation of the constituent cells of an aerial hypha. B,
Macroconidia of Neurospora crassa. Alternate bulging and constriction occurs throughout
the aerial hypha, septa develop at the constrictions and the resulting macroconidia detach
at the slightest disturbance. C, Microconidia of Neurospora crassa. The tiny microconidia
are budded from the conidiophore and form sticky clusters. D, Conidiophore of
Aspergillus (Emericella) nidulans bearing primary and secondary sterigmata (phialides).
From the latter conidia are being produced, the basal ones not yet swollen to full size or
separated from the phialides. E, Conidiophore of Penicillum expansum bearing metullae
on which phialides are producing conidia. D and E are diagrammatic, phialides and
conidia being more numerous than shown.
N. crassa is self-sterile with two mating types designated A and a. Mating type is
controlled by a single locus with the alternative alleles A and a. A single strain of
either mating type will produce protoperithecia consisting of ascogonia bearing
trichogynes but further development occurs only if a macroconidium or
microconidium from the other mating type reaches the trichogyne. Then a
perithecium is produced containing numerous asci. Each ascus contains eight
initially uninucleate ascospores within which mitosis occurs to give a
multinucleate condition. The asci elongate through the neck of the perithecium to
discharge their ascospores. Neurospora ascospores are dark and thick-walled and
can survive for many years. They germinate only if dormancy is broken by heat
shock (60~ for 20 minutes) or treatment with furfural.
Genetic studies on N. crassa, including analysis of ordered tetrads and
chromosome mapping, were already being carried out in the 1930s. At the end of
the decade the organism was chosen by Beadle and Tatum to examine the
possibility that genes act by controlling the production of enzymes, a view that
can be precisely formulated as the one-gene one-enzyme hypothesis. They showed
that wild Neurospora strains could be grown on inorganic media containing a
sugar and the vitamin biotin. Conidia were then treated with X-rays, mated with
wild-type cultures, and single ascospores isolated with a view to obtaining mutant
cultures derived from single haploid nuclei. Some cultures obtained in this way
were shown to have nutrient requirements additional to those of the wild type,
and it was established that such a deficiency could result from the mutation of a
single gene. The study of a range of mutations resulting in the same nutritional
deficiency led to the elucidation of the biosynthetic pathway for tryptophan and
for ornithine. Neurospora crassa thus had a major role in the foundation of
biochemical genetics and is now genetically and biochemically one of the most
intensively studied organisms.
Hyphal anastomosis occurs readily in N. crassa, as in many Ascomycetes. It is
therefore possible to obtain heterokaryons, mycelia that contain genetically
different nuclei. This can be done by bringing together two mutant strains each
with a different nutritional deficiency on a medium that lacks both nutrients.
Under these circumstances heterokaryosis leads to an organism with a selective
advantage as compared with either constituent strain. Heterokaryosis has been
extensively utilized in genetical research, but the formation of heterokaryons
through hyphal anastomosis in nature may be rare. This is due to vegetative
incompatibility. Isolation of strains from nature has demonstrated the occurrence
of many incompatibility groups, heterokaryon formation being able to occur
within a group but not between groups. A genetic difference at any one of a large
number of loci, termed het loci, results in somatic incompatibility.
Neurospora sitophila and N. intermedia resemble N. crassa in being self-sterile
and having eight-spored asci. N. tetrasperma has, as the name indicates, four
spores in each ascus. Each spore contains two nuclei, one carrying the A and one
the a mating type allele. This is the result of the precise way in which meiosis and
ascospore wall delineation occur in this species. The presence of both A and a
alleles in single ascospores, yields a culture capable of producing perithecia. The
consequences of possessing a mating system are thus evaded and apparent self-
fertility results. These four species with a mating system are sufficiently closely
related to be self-fertile. There are also five Neurospora species known which
have eight-spored asci, are self-fertile without any indications of a mating system,
and lack conidia.
The genus Podospora, which grows on herbivore dung, is closely related to
Neurospora. In some species repeated mitosis following meiosis results in large
numbers of ascospores in the ascus. P. decipiens, for example, has strains
characterized by 8, 16, 32 or 64 ascospores per ascus. The most intensively
studied species, P. anserina, resembles N. tetrasperma in normally having four
binucleate ascospores per ascus, with the two nuclei in each ascospore carrying
nuclei of opposite mating type. The precise meiotic manoeuvres and spore wall
delineation that result in this condition are, however, different from those in
N. tetrasperma and must have evolved independently. Some asci of P. anserina
contain both large binucleate and small uninucleate ascospores. Germination of
the small ascospores yields self-sterile cultures. P. anserina does not produce
macroconidia, and the microconidia, although capable of fertilizing ascogonia of
the opposite mating type, hardly ever germinate to yield a mycelium. P. anserina
has been employed extensively in studies on vegetative incompatibility.
Sordaria (Fig. 5.8A, B) is another genus that is closely related to Neurospora
and is found on dung. In the self-fertile species S. fimicola extensive studies on the
physiological control of perithecium formation have been carried out and
mutants have been obtained that block various steps in perithecium production.
Ascospore discharge and its control have been studied in detail. Each ascus in turn
elongates through the neck of the perithecium. The ascus then bursts, discharging
the ascospores through the ruptured tip, and collapses. The perithecium neck is
capable of phototropic orientation to point in the direction of maximum
illumination. Ascospore discharge is stimulated by warmth, light and low relative
humidity and hence tends to occur during daytime. Ascospore colour mutants
have been used for the analysis of linear tetrads by direct spore counting and have
been valuable for studies on gene conversion.
Another genus, Chaetomium, is common in soil and on dead vegetation. It
produces cellulase which enables it to attack cellulose-rich substrates, causing
deterioration of paper, straw and wood under damp conditions. The perithecia
lack any obvious neck and are hairy. The asci lyse outside the perithecium
liberating the ascospores which ooze through the aperture of the perithecium like
toothpaste from a tube as a 'spore tendril'. Dispersal is probably by rain splash.

Ophiostoma and Claviceps

Ophiostoma ulmi (formerly Ceratocystis ulmi), a member of the order

Ophiostomatales, causes Dutch elm disease. The epidemic that has devastated the
elms of Western Europe in the last few decades is due to an aggressive strain that
has recently been given specific status as Ophiostoma novo-ulmi. The fungus
produces asexual spores (conidia) by budding at the tips of single hyphal branches
(conidiophores), at the tips of bundles of such hyphae, known as coremia (sing.
coremium) or synnemata (sing. synnema) or by the budding of hyphae growing
within an older hypha. The conidia themselves may subsequently multiply by
yeast-like budding. As in Chaetomium the asci lyse within the perithecium,
releasing ascospores. These ooze out of the tiny perithecium through its long
(ca 1 mm) slender straight neck to give a stalked spore drop. This, as well as the
droplets of conidia produced at the tips of conidiophores and coremia, are viscous
and are probably carried from tree to tree by the bark-boring beetles which by
tunnelling beneath the bark initiate infections. Spread within a tree occurs by
mycelial growth and the transport of conidia through the water-conducting
vessels of the sapwood. The fungus produces toxins and also causes blocking of
the vessels by gum production.
The ergot fungi, forming the genus Claviceps in the order Hyopocreales, are
parasites of grasses and cereals. Infection is limited to the ovary, which is
penetrated by hyphae from germinating ascospores or conidia. The ovary is
extensively colonized by hyphae, which produce large numbers of conidia at the
surface of the ovary. The infected ovary produces an exudate rich in sugars and
amino acids, 'honeydew', which accumulates in the floral cavity and may even
overflow. The exudate supports further fungal growth and attracts insects which
feed on honeydew and the conidia it contains. The conidia are dispersed to other
florets and plants by contact, rain splash or on insects. Sporulation and honeydew
secretion ceases in a few weeks and the infected ovary develops into a hard black
mass of hyphae, the sclerotium (pl. sclerotia) (Fig. 8.17) instead of the seed that it
should have become. Sclerotia, which are capable of prolonged survival, are
widespread in higher fungi. Those of Claviceps are conspicuous and have been
known for centuries as ergots. The ergots fall to the ground in autumn and
survive the winter. In early summer one or a few fruit bodies emerge (Fig. 2.17D)
and on them numerous perithecia form. Long needle-shaped ascospores develop,
are discharged into the air, dispersed by air currents and initiate a fresh season's
round of infections.
The best known ergot fungus is Claviceps purpurea (Figs. 2.17D, 8.17), the
ergot of rye. In past centuries the contamination of rye bread with ergot was
responsible for horrifying outbreaks of ergotism involving gangrene, loss of limbs
and death. The disease is now of mainly veterinary interest, as cattle or sheep
eating ergot-infected grasses may suffer serious effects including abortion. Ergot
contains a wide variety of alkaloids that stimulate the central and sympathetic
nervous systems in various ways producing a range of secondary effects. Ergot
preparations have been used for centuries for hastening childbirth and controlling
subsequent bleeding, and in the twentieth century chemists and pharmacologists
identified many active components and obtained safer and more reliable
preparations. Therapeutically useful ergot alkaloids are now produced on a large
scale by the fermentation industry (page 521).

The Eurotiales: AspergiUus, Penicillium and their Teleomorphs

Members of the order Eurotiales produce their asci within cleistothecia,

approximately spherical closed ascocarps. The ascus walls lyse, releasing
ascospores (Fig. 4.1F) into the ascocarp cavity. Ultimately the ascocarp is
ruptured, but there is no specific mechanism for ascospore dispersal. The
Eurotiales are of interest because of the abundance and importance of the
anamorphic states of some genera. Several teleomorphic genera, including
Eurotium and Emericella, have their anamorphic states in the genus Aspergillus,
and the teleomorphic genera Eupenicillium and Talaromyces in the genus
Penicillium. Teleomorphic states have not been demonstrated for many species of
Aspergillus and Penicillium, and they are usually infrequent even in those species
where they do occur. The designations Aspergillus and Penicillium are hence
much more widely used than those of the corresponding teleomorphic genera.
The generic name Aspergillus is derived from the Latin aspergillum, a mop for
distributing holy water, and refers to the appearance of the mature conidiophores
which are the means of asexual sporulation in the genus. A conidiophore
(Fig. 2.19D) rises perpendicularly from the substrate hyphae and at its tip
develops a swelling, the vesicle. From the vesicle arise numerous short hyphae.
These may bear conidia, or they may branch to give further short hyphae which
carry the conidia (Fig. 4.1E). The hyphae which actually carry the conidia are
termed phialides, and the hyphae which bear phialides are termed metullae (sing.
metuUus). Some authors term the hyphae that arise directly from the vesicle
primary sterigmata, regardless of whether they bear conidia or a second set of
hyphae, and the second set of hyphae, if present, secondary stigmata. The conidia
that are produced on the phialides are not immediately shed, so the production of
a succession of conidia from a single sterigma results in a chain of conidia. It is the
numerous chains of conidia arising from the vesicle that produce the mop-like
appearance. The conidia are very hydrophobic and thus not readily wetted. On
maturity, however, they separate readily and are dispersed by air currents.
Aspergillus is tolerant of low water activity, being able to grow on substrates of
high osmotic potential and to sporulate in an atmosphere of low relative
humidity.
Aspergillus is divided into subgenera and sections, often informally termed
groups. Each group is named from one of the better known species within it.
Assignment of an isolate to a group is usually easy but the determination of the
precise species more difficult. Some groups are of great practical significance, but
the Aspergillus nidulans group is also of interest because A. nidulans (for
sporulation, see Figs. 2.19 and 4.1D-F) has had an important role in the
development of genetics. A common soil organism, it produces cleistothecia
(often called perithecia by geneticists), on the basis of the morphology of which
the teleomorph is designated Emericella nidulans. It was utilized by Pontecorvo to
establish that genetic analysis could be carried out in self-fertile organisms.
Mutagenic treatment of a single isolate resulted in strains with a range of
nutritional deficiencies, and bringing together such strains on minimal medium
forces heterokaryon formation. Examination of the progeny of such
heterokaryons established that recombination had occurred during meiosis and
that linkage data could be obtained. Following chromosome mapping by meiotic
recombination it was demonstrated in addition that genetic analysis could be
carried out without recourse to the sexual process. In heterokaryons some nuclear
fusion to give diploids occurs, and within these diploids crossing over between
homologous chromosomes sometimes takes place during mitosis. The diploids
may revert to the haploid state giving nuclei which will be genetically different
from those of the parental strains. The frequency of diploidization, mitotic
crossing over and haploidization was low but means were found to increase the
frequency and to detect recombination efficiently. The construction of linkage
maps by mitotic recombination in A. nidulans showed that genetic mapping in
systems lacking a sexual phase, such as mitosporic fungi and mammalian tissue
cultures, was possible. A. nidulans has become genetically and biochemically one
of the most intensively studied of all organisms. Another member of the
A. nidulans group, Aspergillus heterothallicus (teleomorph Emericella
heterothallica) is so far the only known Aspergillus species showing sexual
sporulation in which the cooperation of two mating types is needed.
Aspergillus glaucus (teleomorph Eurotium herbariorum) and other members of
the A. glaucus group are exceptional, even among Aspergillus species, for their
tolerance of low water activities. This renders them important agents in
biodeterioration, attacking foods such as jam that have high sugar contents and
products such as textiles and leather under even moderately damp conditions.
The A. restrictus group also tolerate low water activities, and in the humid tropics
will damage camera and microscope lenses, living on traces of organic materials
and producing acids that etch the glass. Members of the Aspergillus niger group
have long been of importance in the fermentation industry as the principal source
of citric acid for soft drinks (page 509), and more recently for the production of
enzymes (pages 476-481). Aspergillus oryzae and other members of the
important A. flavus-oryzae group have an important role in the production of
many Asian foods and beverages (pages 489, 503). On the other hand, some
strains of A. flavus can attack stored groundnuts and other foods, contaminating
them with aflatoxin, a highly toxic and carcinogenic mycotoxin (pp. 440-441).
Members of the Aspergillus fumigatus group are serious pathogens of birds and
occasionally humans, causing the lung infection aspergillosis. Various other
species can cause infection of individuals with impaired immune responses, and
the inhalation of Aspergillus spores should be avoided - even if infections do not
result, allergic responses (pp. 442-443) may be produced.
The genus Penicillium is of comparable importance to Aspergillus. The name is
derived from the Latin penicillus, an artist's brush, and refers to the branching
conidiophores (Fig. 2.19E) on which chains of conidia are borne. Taxonomically,
it is a very difficult genus, species being distinguished by subtle and frequently
variable differences in the details of asexual sporulation. In some species conidia
are borne on phialides which arise at the apex of the conidiophore. In others the
conidiophore bears metuUae on which the phialides are borne, and in some the
conidiophore itself may branch to varying extents prior to bearing metullae. In
some species branching is symmetrical, in others irregular. The spores as in
Aspergillus are hydrophobic, difficult to wet, but easily dispersed by air currents.
Many members of the genus are important in causing biodeterioration, especially
in conditions of low relative humidity. Many species are involved in food
spoilage. Penicillium digitatum attacks citrus fruits. It produces ethylene which
accelerates ripening and renders the fruit (and adjacent ones in storage) more
susceptible to attack. Finally, the fruit is covered in masses of olive green conidia
and shrivels and dries. Penicillium italicum, on the other hand, causes a nauseous
slimy rot of citrus fruit and produces blue-green conidia. Penicillium expansum
causes a storage rot of apples.
In both P. italicum and P. expansum conidiophores may adhere to each other
to give synnemata (sing. synnema), a tendency reaching a spectacular climax in
Penicillium claviforme, which can produce highly differentiated synnemata
centimetres long. Some members of the genus are useful to humans. Penicillium
camembertii and P. roquefortii, for example, have a role in the ripening of soft
and blue-vein cheeses, respectively (page 505). The most famous contribution of
Penicillium to human welfare, however, was its role as the source of penicillin, the
first clinically useful antibiotic (page 516-517).

The Basidiomycetes

Members of the subdivision Basidiomycota, commonly referred to as

Basidiomycetes, are those fungi in which the sexual process involves the
production of haploid basidiospores borne on a basidium in which a diploid
nucleus undergoes meiosis. They are an important group with about 22 000
species known. Most of the conspicuous fungi of fields and woods are
Basidiomycetes. At the microscopic level there are two features that are
characteristic of Basidiomycetes and very widespread in the group. One such
feature is the presence of 'clamp connections' (page 61) apparently linking
adjacent cells of hyphae. The other is that the basidiospores are usually
ballistospores (page 217)which are actively launched from the basidium. The
classification of Basidiomycetes is controversial. Traditionally, four major groups
have been recognized. Two of these, the Hymenomycetes and Gasteromycetes
are, with a few exceptions, macrofungi with large fruit bodies. The other two, the
smuts and rusts, are microfungi and parasites of plants. The Hymenomycetes
have a hymenium, an extensive area of basidia exposed in such a way that
basidiospores can be launched efficiently into the air by a process of active
discharge. The Gasteromycetes have basidiospores that are not actively launched.
Instead there are a wide variety of fruit body forms and dispersal methods. It
would seem that the only common feature of the Gasteromycetes is that they lack
ballistospores and that the group is not a natural one. A recent view, therefore, is
that they should not be formally treated as a separate group, and that within the
Basidiomycota three classes should be recognized- the Basidiomycetes
(Basidiomycete macrofungi and closely related forms), the Ustomycetes (with a
single order, the Ustilaginales or smuts) and the Teliomycetes (with a single order,
the Uredinales or rusts).

The Basidiomycete Macrofungi- the Class Basidiomycetes

There are many orders within the Basidiomycete macrofungi. Here a few
intensively studied species and the features of some orders will be considered.

The Agaricales: Coprinus and Agaricus

Members of the Agaricales, often referred to as the agarics, include the cultivated
mushroom, Agaricus bisporus, already mentioned and illustrated in Chapter 1.
Other agarics have similar fruiting bodies with stalk, cap and gills, although many
are much smaller and a few larger. They are the fungi commonly referred to as
mushrooms if edible and toadstools if not. Not only A. bisporus but also several
other species are cultivated for food (Chapter 8). Although the cultivated
mushroom is the species best known to the public, it has proved a difficult subject
for physiological and genetical research, and many aspects of its biology are ill
understood. The life cycle of another species, Coprinus cinereus (Fig. 2.20), will
hence first be described. This species, which in nature grows on horse dung, can
be grown easily in the laboratory, readily produces fruiting bodies in pure culture,
and is amenable to genetic analysis. Until recently most publications on this
species purported to be on Coprinus lagopus, which grows on sticks and on soil
in woodlands, leading to some confusion.
A single basidiospore of C. cinereus placed on a suitable substratum will
produce a branched septate mycelium. The septa divide the hyphae into
compartments each of which contains a single nucleus. The septa are of a type
limited to Basidiomycetes and known as dolipore septa (Fig. 3.1C). There is a
pore at the centre of the septum, but the septum is greatly thickened adjacent to
the pore, so that the pore resembles a tube rather than a simple opening. On each
side of the pore modified endoplasmic reticulum forms a septal pore cap

Figure 2.20 The life cycle of the Agaric Coprinus cinereus: Haploid (n) basidiospores
germinate to give primary mycelium which can produce oidia, and fuse with oidia or
mycelium of compatible mating type to give dikaryotic (n + n) secondary mycelium with
clamp connections. Secondary mycelium can produce chlamydospores and give rise to fruit
bodies. Fusion of two haploid nuclei to give diploid (2n) nuclei occurs in basidia. Meiosis
follows and each basidium bears four haploid basidiospores.
(sometimes called a parenthosome) with a number of perforations. This complex
septal apparatus permits cytoplasmic streaming between compartments but not
the passage of nuclei. Asexual sporulation takes the form of the production of
oidia on short hyphal branches to form stalked spore drops. C. cinereus is self-
sterile and the initiation of the sexual process requires an encounter between two
strains that differ in mating type.
The mycelium that arises from a single basidiospore is known as the primary
mycelium. It is a homokaryon, having only one type of nucleus, and a
monokaryon, with only one nucleus per cell. Mating type is determined by two
unlinked genetic factors, A and B. Such a mating system is known as bifactorial.
A survey of natural populations of C. cinereus demonstrated 36 alternative
factors at the A locus and 32 at the B locus, but further sampling would
undoubtedly reveal higher numbers. Even on the basis of 36 A factors and 32 B
factors, 1152 (i.e. 36 • 32) mating types are possible. An encounter between two
strains of identical mating type (e.g. A 1 B 1 and A 1B1, a situation referred to as A=
B-) will lead to no further developments, but if the strains differ with respect to
both mating factors (A ~ B ~, e.g. A~ B~ meets A 2 B2) the sexual process goes to
completion and fruit bodies are formed (Table 2.4). The sexual process can be
initiated as a result of encounters classifiable as A=B~ or A~B= but will
subsequently fail. Full sexual compatibility between two strains requires
differences with respect to both A and B factors, and hence not all possible pairs
of different mating types are fully compatible.
Encounters between two strains can occur as a result of two basidiospores
germinating near each other or of the spreading of two adjacent primary mycelia.
The encounter between the two strains is followed by hyphal anastomosis. Then,
if the two strains are fully compatible (A~B~), nuclei from each strain migrate
rapidly through the mycelium of the other. This migration requires that the
dolipore septa are converted into simple perforate septa, a process that is
controlled by the B factor, so nuclear migration is possible only when the pairs of

Table 2.4 The consequences of encounters between primary mycelia of Coprinus

cinereus of different mating type

Example of mating
Type of encounter type Consequence
A = B = AIB 1 x AIB 1 Dikaryon formation not initiated
A =B ~ AIB 1 • AIB 2 Dolipore septa are converted into simple
perforate septa and nuclear migration
occurs, but no synchronization of nuclear
division or formation of clamp connections
A # B= A I B 1 • A2B 1 Only a few dikaryotic cells and clamp
connections are formed, since nuclear
migration cannot occur and most cells
remain monokaryons
A ~ B# A1B 1 • AzB 2 Nuclear migration and dikaryon formation
occur, the secondary mycelium develops
clamp connections, and fruiting bodies are
formed
strains differ with respect to the B factor (i.e. A~B~ or A=B~). Nuclear migration
in C. cinereus occurs at a rate of up to 1 mm h -1. The result of migration is that
each compartment in both mycelia contains a nucleus of each mating type. The
mycelia have hence become dikaryons, and the migration process is termed
dikaryotization or, in some of the older literature, diploidization. Migration is
accompanied by nuclear division, so as to permit the doubling of the number of
nuclei in both mycelia. Dikaryotization can occur by the deposition of an oidium
from one strain close to the mycelium of another. A hypha from the mycelium
shows chemotropic curvature towards the oidium and fuses with it and, if the
strains are compatible, dikaryotization follows. Dikaryotization can also occur if
a monokaryon encounters a dikaryon carrying nuclei of a compatible mating
type. Under these circumstances nuclear migration will be unilateral instead of
reciprocal.
The processes of nuclear migration and dikaryon development are controlled
by proteins encoded by the A and B factors. The B factors encode pheromones
and their receptors which control interactions between the nuclei of the two

A o 9 )

B 9 o . )

C o o 9 )

0 o io 9 )

F o o I -'.". . . . )

Figure 2.21 Diagram illustrating how the dikaryotic state is maintained by means of
clamp connections. A, A dikaryotic hyphal tip, with two nuclei of different mating types.
B, The development of a backwardly growing side branch, into which one of the nuclei has
moved. C, Synchronous division of the two nuclei has occurred, and one of the daughter
nuclei from the side branch nucleus has moved back into the main hypha. D, A septum has
developed between the daughter nuclei derived from the nucleus that remained in the main
hypha, and a second septum cutting off the side branch. Septal pores are not indicated. E,
The tip of the side branch has fused with the main hypha, giving the clamp connection
typical of the Basidiomycete dikaryon, and the side branch nucleus has moved into the
main hypha. A new compartment with two nuclei of differing mating type has thus been
formed, with the apical compartment remaining dikaryotic. F, An illustration of the
probable consequence of nuclear division is a dikaryotic hyphal tip as illustrated in A, but
without clamp connection development, resulting in hyphal compartments reverting to the
monokaryotic state.
parent strains in the mycelium. The A factors encode transcription factors which
regulate the development of the dikaryotic mycelium.
The dikaryotic mycelium produced by the fusion of two fully compatible
mycelia will continue to grow, but the new growth will differ in form and
constitute a secondary mycelium. The most striking feature of the secondary
mycelium is the presence of clamp connections (Fig. 2.21), devices for
maintaining the dikaryotic condition as growth continues. Consider a dikaryotic
apical cell with two nuclei of differing mating type. If these undergo nuclear
division the most likely consequence is two nuclei of one mating type near the
apex and two nuclei of the other mating type towards the rear of the cell. Cross-
wall formation which restores the binucleate state would then result in two
homokaryotic cells. What in fact happens is that the apical cell produces a short
backward pointing side branch and one nucleus moves into it, the other nucleus
remaining in the hypha but close to the branch. Simultaneous nuclear division
then occurs, and a nucleus from within the side branch and one from within the
hypha move forwards towards the hyphal apex. The formation of a cross-wall at
the base of the side branch and across the main hypha close to it results in a new
apical cell containing two nuclei, one of each mating type. The side branch curves
in towards the hypha and fuses with the sub-apical cell which already contains
one nucleus. The nucleus in the side branch, which is of the complementary
mating type, migrates into the sub-apical cell. Thus one cell has given rise to two,
still dikaryotic, cells.
The proper development of secondary mycelium requires the participation of
nuclei that differ with respect to both the A and B factors. The cooperation of
complementary A factors is needed for the properly synchronized division of the
two nuclei in apical cells, and for the production of clamp connections, and the
cooperation of complementary B factors for the fusion of the clamp cell with
the sub-apical cell. The secondary mycelium (dikaryon) differs from the primary
mycelium (monokaryon) not only in having binucleate cells and clamp
connections but in other ways. The hyphae are wider (about 7 lam instead of
4 lam), advance faster, giving a less dense growth, and have branches at a more
acute angle with respect to the parent hypha. Oidia are not formed; instead thick
walled chlamydospores may be produced.
Fruiting bodies (Fig. 2.22, for a related species) are formed only on Ar162
secondary mycelia. The production of primordia can be initiated by light, even a
short exposure at low intensity being effective. The primordia develop in about 5
days from a tiny tangle of mycelium into a 'button' about 1.5 cm high in which
the various parts of the mature fruiting body are present in miniature. Then, over
a period of about 6-9 hours, elongation of the stipe to a length of about 10 cm
occurs. The stipe at first grows towards the light (positive phototropism) and then
upwards (negative geotropism). This sequence of sensory responses is probably
the most effective for placing the gills, which meanwhile are exposed by the
expansion of the cap, in a position optimal for the discharge of basidiospores.
Stipe extension and cap expansion is due almost entirely to cell elongation and
enlargement, there being very little cell division at this stage. Glycogen
accumulated in the primordium disappears, and a high chitin synthase activity is
observed. It is likely, therefore, that the glycogen is the source, via glucose, of the
N-acetylglucosamine from which the chitin of new wall material is synthesized.
Figure 2.22 The fruit bodies (basidiocarps) of Basidiomycetes. A, A young fruit body of the ink cap
Coprinus comatus, with older fruit bodies above and to the side. These are undergoing autodigestion, which
follows a wave of basidiospore production moving upwards along the gills, which are concealed from view.
B, The stipe and under surface of the fruit body of Boletus tomentosus, showing the openings of the pores
which are lined with basidia. C, View from below of the fruit body of Hericium crinaceus, showing spines
which are covered with basidia. (A-C, John and Irene Palmer.) See also Fig. 4.10. Further illustrations of fruit
bodies are available at http://www.wisc.edu/botany/fungi/html

The surface of a gill is covered by basidia (Fig. 2.23) and by paraphyses, cells
which act as spacers, keeping the basidia separate. Nuclear fusion and meiosis
take place during the final stages of button development but basidiospore
production only occurs after the gills are exposed. Each basidium develops four
projections, sterigmata, the tip of each of which swells to become a basidiospore.
The four nuclei resulting from meiosis move into the four developing
basidiospores. The basidiospores are discharged violently from sterigmata in a
manner which is characteristic of all actively discharged basidiospores. A droplet
Figure 2.23 Diagram of basidiospore development in Agaricus bisporus. A, A basidium
with two haploid nuclei of compatible mating types. B, A diploid nucleus has resulted from
nuclear fusion. C, Meiosis has occurred, and since A. bisporus is bipolar, two nuclei are of
one parental mating type and two of the other. Sterigmata have been formed and the
basidiospore initials have begun to swell. D, Two nuclei have moved into one spore and
two into the other. The basidia of other Agarics (e.g. Coprinus cinereus) are very similar,
but bear four spores each of which receives a single nucleus.

appears at the base of the basidiospore and in the course of a few minutes
expands. The spore is then shot off, by a 'surface-tension catapult' mechanism,
described in detail later (pages 217-219). Although the initial launching speed is
high, basidiospores are small so the momentum is such that they are discharged
only a short distance. This is essential, otherwise they would be deposited on the
opposite gill. Instead, gravity results in the basidiospores falling clear of the gills
to be dispersed by air currents. Actively discharged basidiospores, which occur
throughout the Hymenomycetes, are termed ballistospores (page 217).
Basidium maturation commences at the base of the gills and moves upwards.
Following basidiospore discharge, the exhausted area of the gill is lysed by
chitinases. Its removal eliminates an obstacle to falling basidiospores, enabling
gills to be close to each other and not absolutely vertical in orientation. Gill
autolysis produces black droplets, hence the popular name for members of the
genus Coprinus, ink caps. A large ink cap, Coprinus comatus, is estimated to
produce a total of about nine thousand million basidiospores in its active period
of 2-3 days, or about 30 000 s-1.
Since fruiting bodies of Coprinus cinereus are produced only on A~B~
mycelium, the diploid basidium nucleus is heterozygotic for both mating type
factors, having a genetic constitution with respect to mating type of, for example,
AIA2BIB2. This means that a single basidium can produce four basidiospores each
of which has a different mating type specificity, e.g. AIB1, AIB2, A2B1, A2B2. Such
a bifactorial mating system is hence also termed tetrapolar. The way that it
promotes outbreeding is discussed in Chapter 5 (Table 5.2).
The large ink cap Coprinus comatus also has many mating types, although they
result from the occurrence of many alleles of one instead of two mating type
genes. This means that a basidium nucleus with, for example, the mating
genotype A1A2, can give rise to only two basidiospore mating types, A 1 and A 2.
Such mating type determination is termed unifactorial or bipolar. Its effectiveness
in promoting outbreeding is discussed in Chapter 5 (page 250, Table 5.1).
Another ink cap, Coprinus sterquilinus, is self-fertile. A single basidiospore gives
a mycelium that after a short period of growth produces cells with two nuclei and
clamp connections and is able to give rise to fruiting bodies.
The genus Agaricus is a large one, with many edible species. Agaricus bisporus,
relatively uncommon in nature, is well known as the cultivated mushroom.
Basidiospores of the cultivated mushroom are difficult to germinate, but some
germination can be obtained by placing spores on an agar surface in the same
vessel as growing mycelium of the fungus, which produces a volatile factor
stimulating germination of spores of the same species. A single spore that
germinates will usually yield a mycelium capable of producing fruiting bodies.
The mycelium has many nuclei per cell, and nuclear migration and clamp
connections do not occur. The existence of a mating factor that occurs in two
forms, A 1 and A 2 has been established. In a basidium an A 1 and an A 2 nucleus fuse
to give a diploid nucleus that undergoes meiosis to yield two A 1 and two A 2
nuclei. The two basidiospores commonly receive a nucleus of each type and hence
a single spore can usually produce a self-fertile mycelium. A. bisporus thus seems
to have evolved to a self-fertile state from a self-sterile ancestor with a bipolar
mating system. Bipolar Basidiomycetes commonly have a large number of A
factors, and the occurrence of only two A factors in cultivated strains of
A. bisporus suggests that the cultivated mushroom arose from a single wild source
rather than having been repeatedly introduced into cultivation. The origin of the
cultivated mushroom and its cultivation are considered further in Chapter 8.
The gills of Agaricus are wedge shaped, broad at the base and tapering
downwards, and are precisely oriented in the vertical plane by geotropism. This
permits discharged spores to fall clear of the gills. Such an arrangement is usual in
the agarics, the wave of spore maturation and gill autolysis in Coprinus being
exceptional.

The Boletales
Fruit bodies in this order have a central stalk like those in the Agaricales, but the
hymenium lines vertically oriented pores instead of gills (Fig. 2.22B). Some
genera, such as Rhizopogon and Suillus, are important as mycorrhizal fungi
(Figs. 7.13, 7.14).

The Poriales
Members of this order are important agents of timber decay (Chapter 6). Some,
such as Coriolus versicolor (Fig. 6.19A) and Fomes fomentarius (Fig. 6.19C),
have leathery or woody fruit bodies with the hymenium lining vertical pores.
Others, such as the oyster fungus, Pleurotus ostreatus (Fig. 6.19A) and shiitake,
Lentinus edodes (Fig. 8.11C), have gills, and an attractive texture and flavour and
are grown commercially for food.

The Schizophyllales
This order includes an intensively studied species, Schizophyllum commune,
which is common on fallen branches. It produces basidia on gill-like lamellae on
the undersurface of small fruit bodies. Under dry conditions the fruit bodies curl
up, but even after a couple of years in the dry state will uncurl in humid
conditions and start discharging basidiospores within a few hours. Schizophyllum
is easily grown in pure culture on defined media and readily produces fruit bodies
under such conditions. It is self-fertile with dikaryon formation and fruiting
controlled, as in Coprinus cinereus, by two genetic factors, A and B. A study of
natural populations identified 96 A and 56 B factors. Both the A and B factors
have been shown to consist of two closely linked genes, A0~ and AI3, and B0~ and
BI3. Many alleles have been found at each locus, and recombination between the
0~and [3 loci results in new specificities. Nine A0~ and 32 A~ alleles are known, and
these recombined in different ways would give 9 x 32 or 288 different A factors.
Similarly, the 9 B0~ and 9 B~ alleles known would allow 81 B factors. With many
more 0~ and [3 alleles likely in nature, the probable number of A and B factors
existing is likely to be very large. In Coprinus cinereus too both the A and B
factors are made up of closely linked genes. Both S. commune and C. cinereus
have been used in studies on the mode of action of the A and B factors.

The Lycoperdales (Puff-balls), Nidulariales (Bird's Nest Fungi) and Phallales

(Stinkhorns)
Members of these orders have basidiospores that are not ballistospores, and
which detach from the basidia in a non-violent manner. In puff-balls
(Fig. 4.10A, B) they are released into a cavity in the fruit body to form a powdery
mass. Drops of rain striking the fruit body, or other pressures on it, can then eject
puffs of spores through an aperture. Bird's nest fungi (Fig. 4.10C) have splash
cups from which rain drops fling spore packages. In the stinkhorns (Fig. 4.10D)
the spores form a sticky layer which attracts flies and other insects which effect
dispersal. All these and other orders which lack ballistospores were formerly
included in the Gasteromycetes.

The Auriculariales and Tremellales

Members of these orders have phragmobasidia, in which septa develop in the
basidium after meiosis, rendering it multicellular. This contrasts with other
orders, members of which have holobasidia, in which the basidium does not
undergo septation, and is a single club-shaped cell which when mature carries the
basidiospores - almost always four - at one end. A member of the Auriculariales,
Auricularia polytricha, and a member of the Tremellales, Tremella fusiformis, are
cultivated on a large scale for food in the Far East. Other species of Tremella have
been used for studies on sex hormones (page 205) and mating systems (page 208).
Many members of the Tremellales readily form a yeast phase, and some
Basidiomycete yeasts (page 75) have been assigned to the Tremellales.

The Class Ustomycetes and Order Ustilaginales (smuts)

The largest order in the class Ustomycetes is the Ustilaginales or smuts, important
plant pathogens. The smuts, of which about 1000 species are known, are
biotrophic (page 391) pathogens of flowering plants, and cause economically
important diseases of cereals. Masses of black sooty spores are formed, hence the
popular name, smuts. The part of the plant which is commonly the most
susceptible to infection and damage is the flower, so smuts can cause loss of seed,
such as cereal grain. The smut of maize, Ustilago maydis, results in galls
(hypertrophied host tissue containing fungus mycelium) that replace kernels in
the maize cob. These galls are a Mexican delicacy, huitlacoche. The life cycle
(Fig. 2.24) of this well studied species will be described.
The galls produced by Ustilago maydis on maize kernels break open to expose
the sooty spores characteristic of smuts. The spores when mature are uninucleate
and diploid and are known as teliospores, teleutospores, chlamydospores or
sometimes brandspores. These spores, produced on the maturing maize cobs in
the autumn, are both a means of dispersal and the way in which the fungus

Figure 2.24 The life cycle of Ustilago maydis. Diploid (2n) teliospores undergo meiosis
when they germinate to give haploid (n) promycelia. The promycelium bears sporidia,
equivalent to basidiospores. Sporidia can bud like yeasts, and probably multiply in this
way on plant debris and other organic matter at the soil surface. If two sporidia of
compatible mating type germinate in proximity on the surface of a maize plant, they mate
to give a dikaryon (n + n) able to initiate infection. Galls composed of mycelium and host
tissue arise on stems, leaves and corn cobs. Mycelial cells finally round off to give
teliospores, during the maturation of which nuclear fusion gives the diploid (2n) state.
survives the winter. Since mature teliospores are diploid they constitute a stage
comparable to the basidia of other Basidiomycetes. Meiosis occurs in the
germinating spore and a promycelium with four cells each containing a haploid
nucleus is produced. Each cell produces a bud, the sporidium (pl. sporidia) which
is regarded as equivalent to a basidiospore. The cells of the promycelium,
however, are capable of repeated sporidium production, and the sporidia are able
to multiply by budding. The haploid sporidial phase grows readily in pure culture
on nutritionally simple media, forming yeast-like cells and compact colonies on
agar. Mating is controlled by two genetic factors, a and b.
Only two alleles occur at the a locus, a I and a 2. Fusion between haploid cells
only occurs if they differ at the a locus. The a alleles each have two regions. One
region specifies a diffusible sex hormone that attracts the conjugation tube that
emerges from a nearby cell of the opposite mating type. The other region specifies
the receptor for the sex attractant produced by the opposite mating type. Hence,
with two compatible cells close to each other, the conjugation tubes home in on
each other and fusion results. There are at least 30 alleles at the b locus.
U. maydis, with two alleles at one mating type locus and many at a second, thus
displays what has been termed modified tetrapolar incompatibility (page 251).
The development of a dikaryotic mycelium and growth within the host plant
takes place only if fusion was between haploid cells that carried different b
factors. Mating can be carried out on agar media but attempts to grow the
dikaryotic mycelial phase in pure culture have had very limited success. The
dikaryotic mycelium that grows within the plant may send hyphae to the surface
where dikaryotic conidia are produced and dispersed to initiate further infections.
Nuclear fusion occurs in the developing brandspore to give the diploid condition.
Since mating is controlled by two genetic factors, sporidia of four different mating
types can be produced by meiosis during the germination of a single brandspore.
Another smut that has received considerable study is Ustilago violacea, the anther
smut of the Caryophyllaceae (carnations and campions). Infection results in the
production of brandspores instead of pollen in the anthers. Two hosts, the campions
Silene dioica and Silene alba, have separate male and female plants. Infection of
female plants results in partial suppression of the female parts of the flower and
the production of anthers within which brandspores develop. Ustilago violacea
has a unifactorial mating system with two alleles, a I and a 2. This is more common
in the Ustilaginales than is the bifactorial system of Ustilago maydis. Mating in
U. violacea has received considerable study. Contact is first established between
compatible cells by means of fimbriae, long proteinaceous hairs produced by both
mating types and visible only by electron microscopy. A conjugation tube grows
from the a 2 cell and fuses with a peg it induces in the a I cell.
Another order, the Sporidiales, includes some important yeasts, such as
Rhodosporidium (page 75).

The Class Teliomycetes and Order Uredinales (rusts)

The larger of the two orders in the class Teliomycetes is the Uredinales or rusts,
so called because of the rust-coloured masses of spores produced by many species
on the plants that they infect. About 7000 species are known. All are biotrophic
parasites, attacking ferns, conifers and especially flowering plants. Some, for
example Puccinia graminis (Fig. 2.25), which attacks cereals and grasses, are
economically important. The variety of P. graminis that attacks wheat has been
intensively studied. The diploid stage, the teliospore or teleutospore, develops in
the autumn on wheat plants and survives the winter in stubble. Germination and
meiosis occur in the spring and four basidiospores are produced on sterigmata.
These are of two mating types, designated plus (§ and minus (-). The actively
discharged basidiospores cannot infect wheat plants but will infect a second host,
the barberry, in which a haploid monokaryotic mycelium is formed. Pustules

Figure 2.25 The life cycle of the rust Puccinia graminis. Teliospores (teleutospores)
develop on wheat plants during the autumn, with nuclear fusion occurring during their
formation to give the diploid (2n) state. The teliospores survive the winter in stubble.
Meiosis to give the haploid (n) state occurs during germination, and a promycelium which
bears four basidiospores emerges from each of the two compartments of the teliospore.
The basidiospores infect barberry plants, and pycnia develop on the upper surface of
leaves. These contain pycniospores (spermatia), which ooze from the pycnia, and receptive
hyphae. Insects can carry pycniospores to the receptive hyphae of pycnia of the opposite
mating type, resulting in the establishment of dikaryotic (n § n) mycelia. Dikaryotic
aeciospores formed in aecia on the lower surface of barberry leaves are dispersed, and may
infect wheat plants. Uredospores spread the infection among wheat plants, but in early
autumn their production is gradually replaced by that of teliospores.
develop at the leaf surface. These have pycniospores (spermatia) and receptive
hyphae and produce a sweet-smelling sugary solution that attracts insects. The
insects may carry pycniospores between pustules formed by strains of different
mating type. If a pycniospore of one mating type reaches a receptive hypha of the
other, cell fusion occurs and a dikaryotic mycelium is established. The dikaryotic
mycelium produces binucleate, dikaryotic aeciospores that can infect wheat but
not barberry. In the wheat plant binucleate dikaryotic uredospores are produced,
which spread the infection among wheat plants, and finally teliospores in the
production of which nuclear fusion occurs to give the diploid condition once more.
Some rusts have a less complex life cycle and only one host. About 30 species have
been grown in pure culture, although growth under such conditions is very slow.

The Mitosporic F u n g i

Mitosporic fungi were at one time referred to as Fungi Imperfecti or imperfect

fungi, and later as Deuteromycetes. The 'imperfection' that leads to classification
as a mitosporic fungus is the absence of any sexual stage in the life cycle. Not all
fungi, however, that are permanently anamorphic are assigned to the mitosporic
fungi. For example, anamorphic fungi that have sporangia resembling those of
Zygomycetes are assigned to Zygomycete genera, and those that are clearly
anamorphic rusts, to rust genera. Most fungi classified as mitosporic fungi are
likely to have arisen from Ascomycetes through loss of the sexual phase, although
some are anamorphic Basidiomycetes. About 15 000 mitosporic fungi are known.
Mitosporic fungi can be divided informally into three classes, the Hyphomycetes,
the Agonomycetes and the Coelomycetes. Mitosporic fungi that usually multiply
by budding can be included in the Hyphomycetes but are here dealt with as
anamorphic yeasts (page 75).

The Hyphomycetes
Hyphomycetes are abundant in the soil, and many are of importance as plant
pathogens, as agents of biodeterioration, and in fermentation technology. Their
accurate identification is hence of importance, and there is an extensive literature
on the taxonomy of Aspergillus, Penicillium and other common genera. Most
Hyphomycetes bear conidia on separate conidiophores, but in some the
conidiophores adhere to give synnemata. Spore form and the way in which the
spores are arranged on the conidiophores has been extensively used in
classification, but increasing use is being made of the details of the way in which
the conidia are produced.

The Agonomycetes
Agonomycetes are 'sterile' in the sense of lacking spores and so are sometimes
termed Mycelia Sterilia. Some species produce sclerotia (page 171), and some,
predators on eelworms (page 429), the traps that entangle their prey, but many
have few features that aid identification.

The Coelomycetes

The Coelomycetes, many of which infect plants, are distinguished from the
Hyphomycetes by the production of conidiophores and conidia within
conidiomata (sing. conidioma), structures which with plant pathogenic species
may consist of plant as well as fungus tissue. A common type of conidioma is the
pycnidium (pl. pycnidia), which is a flask-shaped structure resembling the
perithecium of Ascomycetes. Classification of Coelomycetes is based mainly on
conidium and conidioma form, but, as with the Hyphomycetes, increasing use is
being made of the details of conidiophore and conidium development.

T h e Yeasts

Yeasts are fungi which occur predominantly or exclusively in a unicellular state.

Many are found on the surfaces of plants where they are able to exploit naturally
exuded nutrients and more copious exudates that may follow injury or senescence
of plant parts. They are abundant on leaves and fruits and in the nectaries of
flowers. Other yeasts are found on the surface and in the gut of animals, especially
insects, and a few are pathogens of humans and warm-blooded animals (pages
433-438). The surfaces of plants are, however, the most common habitat. Here
yeasts have to compete with fast-growing bacteria in the exploitation of the
nutrients present. Droplets or films of liquid on the surface of plants and animals
will be liable to evaporation, leading to very high solute concentrations and even
to desiccation. Alternatively, rain or dew may result in dilution. Hence
microorganisms on plant and animal surfaces are liable to experience violent
fluctuations in ambient water potential. As will be discussed later (page 163), an
approximately spherical form, and perhaps especially the cell multiplication by
budding characteristic of yeasts, seems to be more appropriate than the hypha
and its apical growth for coping with such fluctuations in water potential.
The production of a yeast form is very widespread in fungi. Relatively few
species, however, have given up the advantages of mycelial growth that permit
success in many environments. There are only a few hundred species that, on the
basis of an exclusively or predominantly unicellular form, are regarded as yeasts.
Their growth in the natural environment seems to be restricted to the limited class
of habitats already discussed, with a frequent occurrence in soil and water
reflecting a capacity for prolonged survival during the course of dispersal. A few
are pathogens of plants or animals, but the great significance of yeasts for humans
lies in their ability to grow readily and with high metabolic rates in nutrient rich
solutions and especially in sugary liquids. This has resulted in their importance in
the production or spoilage of a variety of foods and beverages.
Some yeasts produce ascospores, some basidiospores and some are
anamorphic, lacking a sexual phase.
Ascosporogenous or Ascomycete Yeasts

The ascospore-producing (ascosporogenous) yeasts are by definition

Ascomycetes. They do not, however, form ascocarps, and so were formerly
included in the Hemiascomycetes (page 46). Almost all are assigned to the order
Saccharomycetales. Although most members of the order can bud and are
regarded as yeasts, there are a few, such as Dipodascus candidum, which cannot
do so and are hence not yeasts. A few species constitute a second order, the
Schizosaccharomycetales. These increase in numbers by not by budding but by

Mating
(cell fusion
and nuclear
fusion)
I
I
'

SHMOOS ' MATEDSHMOOS

,. CO.T C ODD,.G

UDD,.Q ?
CELL (~ CELL CE L (~~

ASCUS WITH ASCUS

ASCOSPORES y ( ~

0
l
l
I

Sporulation and
meiosis

Figure 2.26 The life cycle of the Ascomycete yeast Saccharomyces cerevisiae. Diploid
(2n) vegetative cells multiply by budding, but on nitrogen starvation may give rise to asci
in which meiosis occurs to give four haploid (n) ascospores, two of each mating type.
Ascospores germinate to give haploid vegetative cells which are slightly smaller than
diploid cells. Although they can multiply by budding, proximity of cells of the opposite
mating type leads to shmoo formation, mating and return to the diploid phase. In some
yeasts ('haploid yeasts') budding occurs only in the haploid phase, and mating is followed
by meiosis and ascospore formation. In others ('diploid yeasts'), ascospore germination is
immediately followed by fusion between cells of opposite mating type and return to the
diploid state. S. cerevisiae is commonly regarded as a diploid yeast, since mating usually
soon follows ascospore germination; single cells can, however, be used to establish
permanently haploid cultures.
the formation of a cross-wall followed by fission, so are informally known as the
fission yeasts.

Saccharomyces cerevisiae
The best known of all yeasts is Saccharomyces cerevisiae (Figs. 2.26, 8.7); its
natural habitat is the surface of fruit, but it has been used by man for thousands
of years to produce alcoholic beverages (page 481) and bread (page 500).
Biochemically and genetically it is one of the most intensively studied of all
organisms, and is widely used in genetic manipulation as a host for the genes of
other organisms, both for basic research and in biotechnology (page 533).
Saccharomyces - the name means 'sugar fungus' - metabolizes glucose via the
glycolytic (Embden-Meyerhof) pathway to pyruvate. If oxygen is present the
pyruvate can be oxidized via the tricarboxylic acid cycle to carbon dioxide and
water. In the absence of oxygen, or at high sugar concentrations (see below),
alcoholic fermentation occurs, with ethanol and carbon dioxide being produced.
Alcoholic fermentation by Saccharomyces is responsible not only for the
production of beer and wine but also, through carbon dioxide formation, for the
raising of the dough in bread making.
The cells of Saccharomyces cerevisiae are normally oblate spheroids - they have
two axes of equal length and the third longer. Sometimes the third axis is only a
little longer and hence the cell is nearly spherical, and sometimes it is much longer
giving an elliptical outline. Cells usually have a prominent central vacuole. The
nucleus is small with a haploid DNA content about four times that of Escherichia
coli. Since the haploid chromosome number is 16, on average the chromosomes
are smaller than the chromosome of E. coli. In 1996 S. cerevisiae became the first
eukaryote to have its DNA sequence fully determined. About 6000 genes are
present. Mitochondria vary in number and shape, but several are normally
present with one larger and more branched than the rest. Cells increase in number
by budding (Figs. 3.6 and 8.7). A bud is at first small but rapidly swells to reach
the same size as the mother cell. Meanwhile nuclear division has occurred and one
nucleus has passed into the daughter cell. Finally the two cells separate. The
process of budding leaves a birth scar on the daughter cell and a bud scar on the
mother cell. Birth and bud scars can be distinguished by scanning electron
microscopy. A mother cell will carry a single birth scar and bud scars
corresponding in number to the daughter cells that it has produced. A non-
growing population of yeast consists of single cells, but if rapid growth is
occurring many buds and pairs of cells will be seen. Sometimes separation of
daughter cells occurs more slowly than their production and clusters result,
mother cells being attached to several daughter cells, some of which may
themselves have become mother cells and carry buds or daughter cells.
S. cerevisiae is able to grow rapidly in rich media under anaerobic conditions,
a cell population doubling in about 90 minutes. However, since the energy yield
from fermentation is low, about 98% of the glucose present is metabolized to
provide energy and only 2% is incorporated into cell materials. Cell yield per
gram of sugar metabolized is hence low. A high level of glucose in the cell will
suppress aerobic metabolism even if oxygen is present, a phenomenon known as
catabolite repression or the Crabtree effect. Hence on a rich medium with
abundant sugar, fermentation of glucose to ethanol and carbon dioxide occurs
even in aerobic conditions. However, when the glucose supply is exhausted,
ethanol is converted into pyruvate. Some is then metabolized to carbon dioxide
and water via the tricarboxylic acid cycle to yield energy. In addition some is
converted back into the sugars needed for wall synthesis by a reversal of the
glycolytic pathway, a process known as gluconeogenesis (gluco-neo-genesis).
Since aerobic metabolism yields much more energy than fermentation,
approximately 10% instead of 2% of the glucose supplied is converted into cell
material. This increased cell yield under aerobic conditions per gram of sugar
supplied is known as the Pasteur effect.
The life cycle of S. cerevisiae is illustrated in Fig. 2.26. The strains employed in
baking and brewing are commonly diploid. Sporulation can usually be induced in
such diploid strains. Aerobic conditions are essential, and a suitable medium will
lack assimilable nitrogen and sugars and contain acetate or glycoxylate as the
carbon source. Under these circumstances vegetative growth cannot occur and the
operation of the glyoxylate cycle is favoured. A proportion of the vegetative cells
will then act as asci. Within an ascus meiosis occurs and four ascospores are
formed. The ascus wall is not readily lysed, except by treatment with the enzyme
glucuronidase, but presumably in nature breakdown ultimately occurs to release
ascospores. The ascospores are resting cells, more resistant to adverse conditions
than vegetative cells. An ascospore will germinate on a suitable medium
containing assimilable sugar to yield a haploid vegetative cell. Such a cell is
smaller than a diploid cell but has a similar metabolism and can bud and multiply
in the same way. The isolation and germination of single ascospores will give
haploid strains that can be propagated indefinitely in the vegetative state. Return
to the diploid condition requires mating between two haploid cells differing in
mating type. Two mating types occur and are designated a and ~. Cells of mating
type ~ produce ~-factor, a peptide sexual hormone (Fig. 4.7). This acts on cells of
mating type a causing their cell cycle to be arrested in phase G1 (page 106) and
their surface properties to change so that they readily adhere to cells of mating
type ~. Cells of mating type a produce a-factor, another peptide sexual hormone
(Fig. 4.7). This acts in a similar way to m-factor but on cells of mating type ~. The
hormones also cause the yeast cells to change in shape, becoming 'shmoos' (Fig.
4.8). These have projections which fuse with corresponding projections from cells
of the opposite mating type. Cell fusion is followed by nuclear fusion, initiating
the diploid phase (Plate 7). The control of the mating process in S. cerevisiae is
considered further in Chapter 4 (page 207) and the basis for self-fertility in some
strains in Chapter 5 (page 252).

Other Ascomycete yeasts

The grouping of Ascomycete yeasts into genera is based largely on ascus and
ascospore morphology, but nutritional and biochemical tests are important in
delimiting species. Some yeasts differ from Saccharomyces in being exclusively
oxidative in their metabolism, and others are only weakly fermentative.
Vegetative form is also of importance; some yeasts are able to produce mycelium
and some are not. Schizosaccharomyces (Fig. 2.27C, D), which like
Saccharomyces has been intensively studied, is known as the fission yeast because
Figure 2.27 Diversity of yeast form. A, Saccharomycodes ludwigii. Budding is from a
broad base and from the ends of cells. The two mother cells have not yet separated. B, S.
ludwigii. Ascospores germinate and mate within the ascus to restore the diploid state. C,
Schizosaccharomyces pombe. Cell division is by binary fission. D, S. pombe. Ascospore
production immediately follows mating to restore the haploid state. E, Sporobolomyces sp.
Production of a ballistospore, later forcibly discharged, on a sterigma. F, Candidaalbicans.
Production of budding yeast cells, from a hypha.

its rod-shaped cells increase in number by fission, instead of budding. After

nuclear division a cross-wall is formed halfway along the cell; then the two
daughter cells so formed separate.
The life cycles of Ascomycete yeasts appear varied and complicated. They are,
however, basically similar to that of Saccharomyces, except that in some, diploid
vegetative multiplication is lacking and in some, haploid multiplication. In
Schizosaccharomyces, for example, mating is immediately followed by meiosis. It
is hence regarded as a haploid yeast. In Saccharomycodes ludwigii (Fig. 2.27A, B),
however, ascospores of different mating type fuse within the ascus to restore the
diploid state. It is hence a diploid yeast. Although Saccharomyces is capable of
both haploid and diploid budding, it is commonly regarded as a diploid yeast.
This is because, unless cultures have been directly or indirectly derived from a
single ascospore, encounters between cells of different mating type and the
restoration of the diploid state will soon occur, and because the strains used in
baking and brewing are diploid.
Basidiosporogenous or Basidiomycete Yeasts

There are many Basidiomycetes that have been shown to produce a yeast phase in
which cells multiply by budding. This does not usually result in such fungi being
referred to as yeasts. This term is usually restricted to fungi long recognized as
yeasts and subsequently found to produce a sexual phase with Basidiomycete
features. Some Basidiomycete yeasts have been assigned to the Tremellales (class
Basidiomycetes) and others to the Sporidiales (class Ustomycetes).
A well-studied Basidiomycete member of the Sporidiales is Rhodosporidium
toruloides, long known in its haploid anamorphic state as Rhodotorula glutinis.
This species has two mating types, and following conjugation between two cells
differing in mating type, a dikaryotic mycelium with clamp connections is
formed. The mycelium bears teliospores, which contain single diploid nuclei
formed by nuclear fusion during their development. When these thick-walled
resting spores germinate, meiosis occurs, and the short promycelium, the
equivalent of a basidium, carries sporidia (basidiospores). These, on dispersal,
bud to give the yeast phase so completing the life cycle. An example of a yeast
assigned to the Tremellales is Filobasidiella neoformans, the teleomorph of
Cryptococcus neoformans, a pathogen of humans responsible for cryptococcosis
(page 436). Basidiomycete yeasts, like other Basidiomycetes, have a variety of
mating systems. F. neoformans, like R. toruloides, is bipolar with two mating
types. There are also species that have many mating types, some being bipolar and
some tetrapolar, and species that are self-fertile.

Asporogenous or Anamorphic Yeasts

'Asporogenous' means 'not forming spores' and asporogenous yeasts are those
which do not undergo sexual sporulation. Many, however, do produce spores
asexually, so the term 'anamorphic yeasts', referring to the absence of a sexual
cycle, is perhaps preferable.
An example of an anamorphic yeast that produces spores is Sporobolomyces
(Fig. 2.27E), the anamorph of Sporidiobolus (class Sporidiales). It is a pink yeast
that is very common on the surface of leaves. Cells of Sporobolomyces, as well as
multiplying by budding, can produce a single ballistospore on a sterigma.
Inversion of a Petri dish in which cells have been streaked on an agar medium
results in a faint reproduction of the pattern of streaking on the Petri dish lid. This
'shadow' or 'reflection' on the lid is the result of ballistospore discharge and fall
on to the lid. The ballistospore-producing yeasts have hence been termed the
shadow or mirror yeasts.
Most anamorphic yeasts do not form ballistospores. Many species are of
importance in food spoilage, as pathogens of humans or animals or in the
fermentation industry. The identification of species is mainly on the basis of
nutritional and biochemical tests, and species are grouped into genera using such
features as the pattern of budding, whether the cells are pigmented, whether
mycelium is formed and the kinds of spores produced, if any. The genera are often
artificial, in the sense of bringing together species that are only distantly related.
Teleomorphs have not yet been found in some genera, but increasingly molecular
and ultrastructural detail will make it possible to judge whether purely
anamorphic species are related to Ascomycetes or to Basidiomycetes.

The Lichens

Lichens (Figs. 2.28, 2.29, Plate 1) can be loosely grouped into crustose, foliose
and fruticose forms. Crustose species form a crust so firmly attached to rock
or bark as to make removal difficult. Foliose species (folium, Latin, a leaf) are
leaf-like and more loosely attached to the substratum. Fruticose species
(fruticosus, Latin, like a bush) are varied in form and are free from the
substratum except at the point of attachment; some occur on the ground and
others festoon trees. About 14 000 species are known. They are world wide in
distribution but are most striking, both in abundance and variety, in humid
areas free from human disturbance, particularly atmospheric pollution. Moist
conditions are needed for growth but many species can survive prolonged
drought. Lichens are found in both hot and cold deserts and dominate about
8 % of the earth's surface.
A lichen species is an intimate symbiotic association of a fungus- nearly always
an Ascomycete but sometimes a Basidiomycete- and a photosynthetic
microorganism. The fungal component of a lichen (the mycobiont) is unique to
that species and otherwise unknown in nature. The photosynthetic component
(the photobiont) is an alga or a cyanobacterium. Common photobionts are the
green alga Trebouxia, which is uncommon except as a lichen component, and the
green alga Trentepohlia and the cyanobacterium Nostoc, which are widespread in
the free-living state. The cells of the photobiont are in close contact with those of
the mycobiont. They may be limited to a distinct layer or, more rarely, distributed
throughout the lichen. The photobiont provides the association with the products
of photosynthesis, and, if a cyanobacterium, also those of nitrogen fixation. The
benefits it receives are less obvious, but may include protection from desiccation,
screening from excessive light and the provision of mineral nutrients extracted
from the substratum by the fungus. Lichens grow at a very slow rate - often only
a few millimetres per y e a r - and may be very long-lived. The situations in which
they are found are often too poor in nutrients or too exposed to be colonized by
other organisms. Some even occur within the translucent outer layers of porous
stones in the cold deserts of Antarctica. These slow-growing cryptoendoliths
(Greek, hidden in stones) may be the longest-lived organisms on earth. Lichens
have provided chemists with an immense variety of novel substances, including
one of the most useful of pH indicators, litmus.
Lichens may be dispersed in a variety of ways. Fragments of dried lichen may
get scattered by wind. Some species release soredia, small clumps of algal cells and
fungal mycelia, whereas others produce isidia, in effect miniature lichens, easily
detached and dispersed. Most species have perithecia or more commonly
apothecia containing asci from which ascospores are discharged, usually forcibly.
These fruiting bodies are long-lived and may continue to discharge ascospores for
Figure 2.28 Diversity of form in lichens. A, Rhizocarpon concentricum, a crustose (encrusting) lichen, with
concentric rings of black apothecia. It is widespread on slightly basic, siliceous rocks. B, Peltigera
membranacea, a foliose (leaf-like) lichen, here on bark, but common also in grass in damp meadows and even
by roadsides. C, Cladonia arbuscula, a fruticose (bush-like) lichen, among sedges and mosses, it is common
on acid heathlands and peat moors in upland areas. D, Xanthoria parietina, a placodioid lichen-crustose at
the centre but lobed and detachable at the margin. There are numerous nearly circular apothecia. This lichen
is unusual in favouring nutrient-rich sites, such as roofs below TV aerials on which birds perch, and in being
resistant to all but the most extreme air pollution. (A-D reproduced with permission from Dobson, F. S.
(1992). The Lichens: an Illustrated Guide to British and Irish Species, 3rd edn. Richmond Publishing
Company, Slough.)
Figure 2.29 Scanning electron micrographs of sections across lichen thalli. A, Xanthoria sp. At the top there
is a dense cortex of fungal cells, then a zone in which, in addition to fungal hyphae, there are spherical cells
(diam. ca 7 lam) of Trebouxia, a green alga uncommon except in lichens. Fungal hyphae cross the medulla,
the central area with air spaces, to reach the lower cortex. B, Peltigera canina. At the top are fine hairs (fungal
hyphae), then the palisade-like cortex, also composed of fungal hyphae, and at the bottom the algal zone. This
contains spherical cells (diam. ca 4 lam) of Nostoc, a nitrogen-fixing member of the cyanobacteria, a group
formerly known as the blue-green algae. (A, B.E. Juniper. B, Bronwen Griffiths.)

years, unlike the short-lived fruiting bodies of non-lichenized Ascomycetes.

Presumably if an ascospore germinates close to an appropriate alga the lichen is
re-established. In a few species such proximity is guaranteed since the ascospores
carry cells of the photobiont on their surface when they are discharged.
Many lichens are very sensitive to atmospheric pollutants, especially to sulphur
dioxide. This has led to the disappearance of many species from industrialized
countries over the last century, although some species are moderately and a few
very tolerant of pollutants. These differences in sensitivity enable the severity of
pollution in different areas to be evaluated by observing the presence or absence
of lichen species with known degrees of tolerance.
Careful attention to environmental conditions is needed for lichens to be
maintained in the laboratory, otherwise they will die or dissociate into
photobiont and mycobiont. The germination of ascospores and isolation of single
algal cells has permitted a number of photobionts and mycobionts to be
established in pure culture. The growth of the mycobiont is commonly extremely
slow, the production of a medium-sized colony taking many months instead of
the few days characteristic of most free-living fungi. The early stages in the
establishment of a lichen have been studied by bringing together an appropriate
mycobiont and photobiont. Sparse nutrition is required, in order to prevent
excessive growth of one of the partners, and the simulation of natural conditions,
especially alternate wetting and drying. Careful attention to such factors has led
to the establishment of several species in pure culture throughout their life cycles.

Further Reading and Reference

General Works on Fungal Diversity

Alexopoulos, C. J., Mims, C. W. & Blackwell, M. (1996). Introductory Mycology, 4th.
edn. Wiley, New York.
Hawksworth, D. L. (1991). The fungal dimension of biodiversity: magnitude, significance
and conservation. Mycological Research 95,641-655.
Hawksworth, D. L., Kirk, P. M., Pegler, D. N. & Sutton, B. C. (1995). Ainsworth &
Bisby's Dictionary of the Fungi, 8th edn. CAB International, Wallingford.
Isaac, S., Frankland, J. C., Watling, R. & Whalley, A. J. S, ed. (1993). Aspects of Tropical
Mycology. Nineteenth Symposium of the British Mycological Society. Cambridge
University Press, Cambridge.
McLaughlin, D. J., McLaughlin, E. G. & Lemke, P. A. (2000). Systematics and Evolution.
Vol 7B, The Mycota, K. Essen & P. A. Lemke, eds. Springer-Verlag, Berlin.
Onions, A. H. S., Alsopp, D. & Eggins, H. O. W. (1981). Smith's Introduction to
Industrial Mycology, 7th edn. Arnold, London.
Reynolds, D. R. & Taylor, J. W., eds. (1993). The Fungal Holomorph" Mitotic, Meiotic
and Pleomorphic Speciation in Fungal Systematics. CAB International, Wallingford.
Taylor, J. W., Bowman, B., Berbee, M. L. & White, T. J. (1993). Fungal Model Organisms:
Phylogenies of Saccharomyces, Aspergillus and Neurospora. Systematic Biology 42,
440-457.
Webster, J. (1980). Introduction to Fungi, 2nd edn. Cambridge University Press,
Cambridge.
Wessels, J. G. H. (1999). Fungi in their own right. Fungal Genetics and Biology 27,
134-145.

Cellular Slime Moulds

Ashworth, J. M. & Dee, J. (1975). The Biology of Slime Moulds. Arnold, London.
Aubry, L. & Firtel, R. (1999). Integration of signalling networks that regulate
Dictyostelium differentiation. Annual Review of Cell and Developmental Biology 15,
469-517.
Kessin, R. H., Gundersen, G. G., Zaydfudim, V., Grimson, M. & Blanton, R. L. (1996).
How cellular slime moulds evade nematodes. Proceedings of the National Academy of
Sciences of the United States of America 93, 4857-4861.
Loomis, W. F. (1975). Dictyostelium discoideum- a Developmental System. Academic
Press, New York.
Newell, P. C. (1981). Chemotaxis in the cellular slime moulds. In Lackie, J. M. &
Wilkinson, P. C., eds., Biology of the Chemotactic Response, pp. 89-114. Cambridge
University Press, Cambridge.
Olive, U S. (1975). The Mycetozoans. University of Iowa Press, Iowa.
Raper, K. B. (1984). The Dictyostelids. Princeton University Press, Princeton, New Jersey.

Plasmodial Slime Moulds (Myxomycetes)

Aldrich, H. C. & Daniel, J. W., eds. (1982). Cell Biology of Physarum and Didymium, 2
vols. Academic Press, New York.
Bailey, J. (1995). Plasmodium development in the myxomycete Physarum polycephalum:
genetic control and cellular events. Microbiology 141, 2355-2365.
Dove, W. F., Dee, J., Hatano, S., Haugli, F. B. & Wohlfarth-Bottermann, K. E., eds.
(1986). The Molecular Biology of Physarum polycephalum. Plenum Press, New
York.
Ing, B. (1999). The Myxomycetes of Britain and Ireland. Richmond Publishing Company,
Slough.
Martin, G. W. & Alexopoulos, C. J. (1969). The Myxomycetes. University of Iowa Press,
Iowa.
Sauer, H. (1982). Developmental Biology of Physarum. Cambridge University Press,
Cambridge.
Stephenson, S. L. & Stempen, H. (1994). Myxomycetes" A Handbook of Slime Moulds.
Timber Press, Portland, Oregon.

Oomycetes, Chytridiomycetes and Zygomycetes

Buczaki, S. T., ed. (1983). Zoosporic Plant Pathogens: a Modern Perspective. Academic
Press, London.
Cerd~-Olmedo, E. & Lipson, E. D., eds. (1987). Phycomyces. Cold Spring Harbor
Laboratory, New York.
Couch, J. N. & Bland, C. E., eds. (1985). The Genus Coelomomyces. Academic Press,
Orlando, Florida.
Erwin, D. C., Bartnicki-Garcia, S. & Tsao, P. H., eds. (1983). Phytophthora" its Biology,
Taxonomy, Ecology and Pathology. American Phytopathological Society, St. Paul,
Minnesota.
Fuller, M. S. & Jaworski, A., eds. (1987). Zoosporic Fungi in Teaching and Research.
Southeastern Publishing Corporation, Athens, Georgia.
Ingram, D. S. & Williams, P. H., eds. (1991). Phytophthora infestans, the cause of late
blight of potato. Advances in Plant Pathology, volume 7, Academic Press, London.
Lucas, J. A., Shattock, R. C., Shaw, D. S. & Cooke, L. R., eds. (1991). Phytophthora.
Seventeenth Symposium of the British Mycological Society. Cambridge University
Press, Cambridge.
Mountford, D. O. & Orpin, C. G., eds. (1994). Anaerobic Fungi: Biology, Ecology and
Function. Marcel Dekker, New York.
Spencer, D. M., ed. (1981). The Downy Mildews. Academic Press, London.
Trinci, A. P. J., Davies, D. R., Gull, K., Lawrence, M. I., Nielsen, B. B., Rickers, A. &
Theodorou, M. K. (1994). Anaerobic fungi in herbivorous animals. Mycological
Research 98, 129-152.
Ascomycetes, Basidiomycetes and Mitosporic Fungi

Banuet, F. (1992). Ustilago maydis, the delightful blight. Trends in Genetics 8, 174-180.
Barnett, H. L. & Hunter, B. B. (1998). Illustrated Genera of Imperfect Fungi, 4th edn. APS
Press, St. Paul, Minnesota.
Braun, U. (1995). The Powdery Mildews (Erysiphales) of Europe. Gustav Fischer Verlag,
Stuttgart.
Breitenbach, J. & Kranzlin, F. (1984-1996). Fungi of Switzerland. 4 vols. Verlag
Mykologia, Lucerne.
Carmichael, J. W., Kendrick, B. W., Connors, I. L. & Sigler, L. (1980). Genera of
Hyphomycetes, University of Alberta Press, Edmonton, Alberta.
Casselton, L. A. (1997). Molecular recognition in fungal mating. Endeavour 21,159-163.
Cole, G. T. & Kendrick, B., eds. (1981). Biology of Conidial Fungi, 2 vols. Academic Press,
New York.
Cole, G. T. & Samson, R. A. (1979). Patterns of Development in Conidial Fungi. Pitman,
London.
Coley-Smith, J. R., Verhoef, K. & Jarvis, W. R., eds. (1980). The Biology of Botrytis.
Academic Press, London.
Cummins, G. B. & Hiratsuka, Y. (1983). Illustrated Genera of Rust Fungi, revised edn.
American Phytopathological Society, St Paul, Minnesota.
Ellis, M. B. & Ellis, J. P. (1990). Fungi Without Gills (Hymenomycetes and
Gasteromycetes). An Identification Handbook. Chapman & Hall, London.
Ellis, M. B. & Ellis, J. P. (1997). Microfungi on Land Plants: An Identification Handbook.
2nd edn, Richmond Publishing Co., Slough.
Ellis, M. B. & Ellis, J. P. (1998). Microfungi on Miscellaneous Substrates" An Identification
Handbook. Richmond Publishing Co., Slough.
Ginns, J. & Lefebvre, M. N. L. (1993). Lignicolous Corticioid Fungi (Basidiomycota)
of North America: Systematics, Distribution and Ecology. APS Press, St Paul,
Minnesota.
Glass, N. L. & Kuldau, G. A. (1992). Mating type and vegetative incompatibility in
filamentous Ascomycetes. Annual Review of Phytopathology 30, 201-224.
Hanlin, R. T. (1990). Illustrated Genera of Ascomycetes. The American Phytopathological
Society, St Paul, Minnesota.
Hawksworth, D. L. (1994). Ascomycete Systematics. Problems and Perspectives in the
Nineties. NATO ASI Series A, Life Sciences, Vol. 269. Plenum, New York.
Hibbett, D. S., Pine, E. M., Langer, E., Langer, G. & Donoghue, M. (1997). Evolution of
gilled mushrooms and puffballs inferred from ribosomal DNA sequences. Proceedings
of the National Academy of Sciences, USA, 94, 12002-12006.
Hyde, K., ed. (1997). Biodiversity of Tropical Microfungi. University of Hong Kong Press,
Hong Kong.
Kinghorn, J. & Martinelli, S., eds. (1994). Physiology and Genetics of Aspergillus
nidulans. Chapman & Hall, London.
Kubicek, C. P. & Harman, G. E. (1998). Trichoderma and Gliocladium. Vol I: Basic
Biology, Taxonomy and Genetics; Vol II: Enzymes, Biological Control, and
Commercial Applications. Taylor & Francis, London.
Minter, D. W. (1984). New concepts in the interpretation of conidiogenesis in
Deuteromycetes. Microbiological Sciences 1, 86-89.
Moore, D., Casselton, L. A., Wood, D. A. & Frankland, J. C., eds. (1985). Developmental
Biology of Higher Fungi. Tenth Symposium of the British Mycological Society.
Cambridge University Press, Cambridge.
Moss, M. O. & Smith, J. E., eds. (1984). The Applied Biology of Fusarium. Seventh
Symposium of the British Mycological Society. Cambridge University Press,
Cambridge.
Nelson, P. E., Tousson, T. A. & Cook, R. J., eds. (1981). Fusarium" Diseases, Biology and
Taxonomy. Pennsylvania State University, University Park and London.
Peberdy, J. F., ed. (1987). Penicillium and Acremonium. Plenum Press, New York.
Pegler, D. N. (1998). Field Guide to the Mushrooms and Toadstools of Britain and
Europe. Larousse, London.
Phillips, R. (1981). Mushrooms and other Fungi of Great Britain and Europe. Pan Books,
London.
Pitt, J. I. (1988). A Laboratory Guide to Common Penicillium Species. CSIRO Food
Research Laboratory, North Ryde, New South Wales.
Powell, K. A., Renwick, A. & Peberdy, J. F., eds. (1994). The Genus Aspergillus" From
Taxonomy and Genetics to Industrial Applications. Plenum Press, New York.
Ryvarden, L. & Gilbertson, R. L. (1993-94). European Polypores. 2 vols. Fungiflora, Oslo.
Samson, R. A. & Pitt, J. I., eds. (1990). Modern Concepts in Penicillium and Aspergillus
Classification. Plenum Press, New York.
Scott, K. J. & Chakravorty, A. K., eds. (1982). The Rust Fungi. Academic Press, London.
Spencer, D. M., ed. (1978). The Powdery Mildews. Academic Press, London.
Subramanian, C. V. (1983). Hyphomycetes" Taxonomy and Biology. Academic Press,
London.

Yeasts

Barnett, J. A., Payne, R. W., Yarrow, D. & Barnett L. (2000). Yeasts: Characteristics and
Identification, 3rd edn. Cambridge University Press, Cambridge.
Campbell, I. & Duffus, J. H., eds. (1988). Yeasts: a Practical Approach. IRL Press, Oxford.
De Hoog, G. S., Smith, M. Th. & Weijman, A. C. M., eds. (1987). The Expanding Realm
of the Yeast-like Fungi. Elsevier, Amsterdam.
Feldmann, H. (1999). The yeast Saccharomyces cerevisiae: insights from the first complete
eukaryotic gene sequence. In R. P. Oliver & M. Schweizer, eds., Molecular Fungal
Biology, pp. 78-134. Cambridge University Press, Cambridge.
Forsberg, S. L. & Nurse, P. (1991). Cell cycle regulation in the yeasts Saccharomyces
cerevisiae and $chizosaccharomyces pombe. Annual Review of Cell Biology 7, 227-256.
Herskowitz, I. (1988). Life cycle of the budding yeast Saccharomyces cerevisiae.
Microbiological Reviews 52, 536-553.
Kurtzman, C. P. & Fell, J.W. (1996). The Yeasts, a Taxonomic Study, 4th edn. North-
Holland, Amsterdam.
Nasim, A., Young, P. & Johnston, B. F., eds. (1989). Molecular Biology of the Fission
Yeast. Academic Press, New York.
Pringle, J. R., Broach, J. R. & Jones, E. W., eds. (1997). The Molecular and Cellular Biology
of the Yeast Saccharomyces. Vol. 3. Cell Cycle and Cell Biology.
Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.
Rose, A. H. & Harrison, J. S., eds. (1987-1993). The Yeasts, 2nd edn, 5 vols. Academic
Press, London.
Russell, P. & Nurse, P. (1986). Schizosaccharomyces pombe and Saccharomyces
cerevisiae: a look at yeasts divided. Cell 45, 781-782.
Skinner, F. A., Passmore, S. M. & Davenport, R. D., eds. (1980). Biology and Activities of
Yeasts. Academic Press, London.
Tuite, M. F. & Oliver, S. G., eds. (1991). Saccharomyces. Plenum Press, New York.
Walker, G. M. (1998). Yeast Physiology and Biotechnology. Wiley, Chichester.

Lichens

Bates, J. W. & Farmer, A. M., eds. (1992). Bryophytes and Lichens in a Changing
Environment. Oxford University Press, Oxford.
Dalby, D. H., Hawksworth, D. L. & Jury, S. L., eds. (1988). Horizons in Lichenology.
Academic Press, London.
Dobson, F. S. (2000). Lichens: an Illustrated Guide to British and Irish Species, 4th edn.
The Richmond Publishing Company, Slough.
Gargas, A., DePriest, P. T., Grube, M. & Tehler, A. (1995). Multiple origins of lichen
symbioses in fungi suggested by SSU rDNA phylogeny. Science 268, 1492-1495.
Gilbert, O. (2000). Lichens. Harper Collins, London.
Hale, M. E. (1983). The Biology of Lichens, 3rd edn. Arnold, London.
Hawksworth, D. L. & Hill, D. J. (1984). The Lichen-forming Fungi. Blackie, Glasgow.
Honegger, R. (1991). Functional aspects of the lichen symbiosis. Annual Review of Plant
Physiology and Plant Molecular Biology 42, 553-558.
Kershaw, K. A. (1988). The Physiological Ecology of Lichens. Cambridge University Press,
Cambridge.
Lawrey, J. W. (1984). The Biology of Lichenized Fungi. Praeger, New York.
Nash, T. H., ed. (1996). Lichen Biology. Cambridge University Press, Cambridge.
Price, R. C. (1992). The Methuselah Factor: age in cryptoendolithic communities. Trends
in Ecology and Evolution 7, 21.
Purvis, O. W., Coppins, B. J., Hawksworth, D. L., James, P. W. & Moore, D. M. (1992).
The Lichen Flora of Great Britain and Ireland. Natural History Museum, London.
Richardson, D. H. S. (1975). The Vanishing Lichens. David & Charles, Newton Abbot.
Richardson, D. H. S. (1992). Pollution Monitoring with Lichens. The Richmond
Publishing Company, Slough.
Richardson, D. H. S. (1999). War in the world of lichens: parasitism and symbiosis as
exemplified by lichens and lichenicolous fungi. Mycological Research 103, 641-650.
Seaward, M. R. D. (1988). Contributions of lichens to ecosystems. In: M. Galun, ed., CRC
Handbook of Lichenology, vol 2, pp. 107-129. CRC Press, Boca Raton, Florida.

Questions
With each question one statement is correct. Answers on page 561.
2.1 Organisms which belong to the Chromista, not true Fungi, but are important
members of the fungi in the broad sense and include many important plant
pathogens:
a) Chytridiomycetes
b) Zygomycetes
c) Oomycetes
d) Basidiomycetes

2.2 The number of fungal species so far described is about:

a) 700
b) 7000
c) 70 000
d) 70O 000

2.3 Fungi which form extensive mycelial systems consisting typically of hyphae with two
different nuclear genotypes are found in the:
a) Ascomycetes
b) Basidiomycetes
c) Oomycetes
d) Myxomycetes

2.4 Many plants have mutualistic symbioses with fungi in the genus:
a) Glomus
b) Mucor
c) Erysiphe
d) Phytophthora
2.5 Lichens are formed by the association of:
a) Ascomycetes or Basidiomycetes with algae or bacteria
b) Ascomycetes only, with algae or bacteria
c) Basidiomycetes only, with algae or bacteria
e) Ascomycetes only, with algae only