Biology Investigatory Project

On
Sporulation of Fungus

by:

Aagnish Banerjee
XII
Roll No. 01
Medical section
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 INDEX

SERIAL NUMBER TOPIC PAGE NO.

1 certificate 03

2 acknowledgement 04

3 Introduction 05

4 Theory 06-16

5 Experiment 17

6 Observation 18-22

7 Conclusion 23

8 bibliography 24

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 CERTIFICATE

This is to certify that Aagnish Banerjee, a student of class XII

has completed research Biology Investigatory Project on the
topic “Sporulation of Fungus” for the session of 2020-2021
under the guidance of miss Ankita Banerjee as per the CBSE
guidelines.

……………………………………. …………………………………….
Signature of Internal Examiner Signature of External Examiner
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 ACKNOWLEDGEMENT

I take this opportunity to express my profound gratitude and

deep regards to my teacher Miss Ankita Banerjee for her
exemplary guidance, monitoring and constant encouragement
throughout the course of this project. The blessing, help and
guidance given by them time to time shall carry me a long way
in the journey of life on which I am about to embark.

I also take this opportunity to express a deep sense of gratitude to

Sir Sujit Kumar Jana, Principal, Senior Secondary School for his
cordial support, valuable information and guidance, which
helped me in completing this task through various stages.

Lastly, I thank almighty, my parents and friends for their

constant encouragement without which this project would not be
possible.

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 INTRODUCTION

Fungus, plural fungi, any of about 144,000 known species of

organisms of the kingdom Fungi, which includes the yeast, rusts,
smuts, mildews, moulds and mushrooms. There are also many
fungus like organisms, including slime
moulds and oomycetes (water moulds) that do not belong to
kingdom Fungi but are often called fungi. Many of these fungus
like organisms are included in the kingdom Chromista. Fungi are
among the most widely distributed organisms on Earth and are of
great environmental and medical importance. Many fungi are
free-living in soil or water; others form parasitic or symbiotic
relationships with plants or animals.
This experiment has been conducted on the fungi which we are
surrounded by and can see in everyday life (namely bread
moulds).

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 THEORY

Fungi: kingdom of saprophytic and parasitic spore-producing

eukaryotic typically filamentous organisms formerly classified as
plants that lack chlorophyll and include moulds, rusts, mildews, smuts,
mushrooms, and yeasts.

Structure of fungi:
The main body of most fungi is made up of fine, branching and usually
colourless threads called hyphae. Each fungus will have vast numbers
of these hyphae, all intertwining to make up a tangled web called
the mycelium.
The mycelium is generally too fine to be seen by the naked eye, except
where the hyphae are very closely packed together. The picture below
was obtained through a microscope. The hyphae are magnified 100
times life size.

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Fungal mycelium is mostly hidden from human view, not only because
of its small size, but also as a result of its location. The tangled mycelial
mass is usually hidden deep within its food sources, such as rotting
matter in the soil, leaf litter, rotting wood, or dead animals. The
mycelium remains undetected until it develops one or more fruiting
bodies, containing the reproductive spores.
They are usually produced at the surface of the food source, rather than
hidden within it, to allow the spores to be shed and carried away by the
wind, or by water, or animals. The fruiting bodies are usually the only
indication we have that a fungus is present. Like icebergs, they
represent a tiny fraction of the whole fungus, with most of it being
hidden from view.

Inside the fungus:

 Fungal cell walls are rigid and contain complex polysaccharides
called chitin (adds structural strength) and glucans.
 Ergosterol is the steroid molecule in the cell membranes that
replaces the cholesterol found in animal cell membranes.
 Fungi can be unicellular, multicellular, or dimorphic, which is
when the fungi is unicellular or multicellular depending on
environmental conditions.
 Fungi in the morphological vegetative stage consist of a tangle of
slender, thread-like hyphae, whereas the reproductive stage is
usually more obvious.
 Fungi like to be in a moist and slightly acidic environment; they
can grow with or without light or oxygen.

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  Fungi are saprophyte heterotrophs in that they use dead or
decomposing organic matter as a source of carbon.
The morphology of the mycelium mode of spore formation and fruiting
bodies form the basis of the kingdom into various classes.

Types of fungi:

1. Phycomycetes: member of phycomycetes are foind in aquatic

habitats and on decaying wood in moist and damp places or as
obligate parasites on plants. The mycelium is aseptate and
coenocytic. Asexual reproduction takes place by zoospores which
is motile or by aplanospores which are non-motile. These spores
are endogenously produced in sporangium. A zygospore is formed
by fusion of 2 gametes. These gametes are similar in morphology
(isogamy) or dissimilar (anisogamous or oogamous).
Example: Rhizopus (bread mould).

2. Ascomycetes: commonly known as sac-fungi these are mostly

multicellular(Penicillium) or rarely unicellular(saccharomyces).
They are saprophytic, decomposers, parasitic or
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 coprophylus(growing on dung). Mycelium is branched and septate.
the asexual spores are conidia produced exogeneouly on the
special mycelium called coidiophores. Conidia on germination
produce mycelium. Sexual spores are called ascospores which are
produced endogeneously in a sac like asci. These asci are arranged
in a different type of fruiting bodies called ascocarps.
Example: asperigillus, claviceps, neurospora.

3. Basidiomycetes: commonly known forms of basidiomycetes are

mushrooms, bracket fungi or puff balls. They grow on soil on logs
and tree stumps and in living plant bodies as parasites. The
mycelium is branched and septate. The asexual spores are
generally not found but vegetative reproduction by fragmentation
is common. The sex organs are absent but plasmogamy is brought
about by fusion of two vegetative or somatic cells of different
strains or genotypes. The resultant structure is dikaryotic which
ultimately gives rise to basidium. Karyogamy or meiosis takes
place in the basidium producing four basidiospores. The
basidiospores are exogeneously produced on the basidium. The
basidia are arranged in fruiting bodies called basidiocarps.
Example: Agaricus, Ustilago, Puccinia.

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4. Deuteromycetes: commonly known as imperfect fungi because
only the asexual or vegetative phases of these fungi are known.
When the sexual forms of these fungi were discovered they were
moved into classes they rightly belong to. It is also possible that
the asexual and vegetative stage have been given one name (and
place under deuteromycetes) and the sexual stage another (and
placed under another class). Later when the linkages were
established, the fungi were correctly identified and moved out of
Once perfect (sexual) stages of members of deuteromycetes were
discovered they were often moved to ascomycetes and
basidiomycetes. The deuteromycetes reproduce only by asexual
spores known as conidia. The mycelium is septate and branched.
Some members are saprophytes or parasites while a large number
of them are decomposers of litter and help in mineral cycling.
Some examples are Alternaria, Colletotrichum and Trichoderma.

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Sporulation of Fungi
Sporulation refers to the formation of spores from vegetative cells
during unfavourable environmental conditions. As such, it may be
described as an adaptive response that allows the organism to survive
given adverse conditions (radiation, extreme heat or cold, lack of
nutrition etc.).

Fungi are some of the most studied spore-producing organisms in the

world. They produce a wide variety of spores that significantly vary in
size, shape and other surface features that suit their environment (for
dispersal etc).
Whereas the spores produced sexually (through meiosis) remain
dormant for survival (e.g. ascospores), those that are produced asexually
(mitospores) are for dispersal.
Produced through mitosis, asexual spores are released in high numbers
and are genetically identical. This allows them to play an important role
in reproduction when they land on the appropriate substrate in the
environment following dispersal.
* Chlamydospore - This is a type of fungal spore that develops from
the hyphal structures during unfavorable conditions. Chlamydospores
are characterized by a thick, melanized wall that protects the contents of
the spore.
Different types of fungi may produce different types of spores.
The following are some of the main groups of fungi and the type of
spores they produce:
· Zygomycota - Members of Zygomycota are known as zygomeycetes.
They produce both sexual (zygospores) and asexual (sporangiospores)
spores.
· Ascomycota - Ascomycetes also produce both sexual (ascospores) and
asexual (conidia) spores.
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· Basidiomycota - Compared to the other groups of
fungi, basidiomyecetes are largely known to produce sexual spores that
are known as basidiospores.

Sporulation Process in Yeast (Saccharomyces cerevisiae)

Saccharomyces cerevisiae is also known as Brewer's yeast and is used

for brewing, baking and winemaking.
As with bacteria (Gram-positive bacteria), fungi do not necessarily start
forming spores when environmental conditions change. Rather, they
attempt to adjust to the new conditions through a number of strategies.
When nitrogen sources diminish, Saccharomyces cerevisiae may
respond by going into a stationary phase or modifying their
morphology. However, in the event that unfavorable conditions persist,
spore formation becomes necessary. This involves a process known as
meiosis.
Early Phase
During unfavorable conditions, the organism responds by entering the
early phase (first phase of sporulation) of sporulation. During this phase,

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cell division processes shift from the mitotic cycle in G1 (An interphase
stage) and enter the S phase in meiosis.
Some of the events that take place during early phase include:
 DNA replication
 Homolog recombination and pairing (prophase)
This phase is therefore largely characterized by changes in the
machinery of the cell cycle as well as altered RNA processing.

Middle Phase
The middle phase is characterized by various cytological events. Here,
the meiotic process results in the production of haploid nuclei (four) that
form four new daughter cells. This involves the modification of the
spindle pole bodies to contribute to the formation
of prospores (membrane compartments).
Some of the other events that take place during this phase include:
 Post-Golgi secretory vesicles are redirected to promote the
regeneration and expansion of the new compartments
 The prospores grow and engulf the nuclei
 Other organelles move to the cytoplasmic space that lies between
the nuclear membrane and the prospores
 The nucleus is completely separated from the cytoplasm of the
mother cell
Late Phase
This is the last phase of sporulation and involves the following events:
 The prospore membrane closes completely
 A thick wall starts forming around the spores- This also contributes
to the maturation of the spores
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  Chromatic compaction in the nucleus of the spores
 Some of the organelles start regenerating
* While given environments may contain various nutrients, the lack of
nitrogen and other appropriate nutrient sources may trigger sporulation
of species like A. nidulans and result in the growth of hyphae and
consequently conidiation.
* Before spores start forming, special reproductive hyphae known as
sporangiophores are produced.
* Compared to sexual reproduction, asexual reproduction is more
common in fungi given that it allows for the rapid spread of the
organisms.
* Some spores are produced in the sporangium (sac) through mitotic
division (e.g. conidia).
* Fungi spore vary in size and structure. The differences allow them to
be effectively dispersed by a variety of agents including wind, animals,
and water.
Uses of fungi:
Fungi are extremely important to the ecosystem because they are one of
the major decomposers of organic material. Decomposing organic
material is how fungi acquire energy. But fungi have other roles in
addition to being decomposers. How do fungi help people? They are
used to help prepare food and beverages, and they have many other
uses.
Importance of Fungi for Human Use

 Yeasts are crucial for the fermentation process that makes beer,
wine, and bread. Fermentation occurs in the absence of oxygen and
allows the first step of cellular respiration, glycolysis, to continue.
 Some fungi are used in the production of soy sauce and tempeh, a
source of protein used in Southeast Asia.

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 Fungi can produce antibiotics, such as penicillin. Antibiotics are
important medicines that kill bacteria, and penicillin was the first
identified cure against many deadly bacterial species. Antibiotics only
treat bacterial diseases; they cannot be used to treat viral or fungal
diseases.
 Mushrooms are fungi that are eaten by people all over the globe.

Harmful effects of fungi:

(1) Plant Diseases:
These are incited by different agents — bacteria, viruses, cell-worms
and fungi; of these the fungi cause the most severe losses. They destroy
field crops, forage crops, fruit and nut crops, vegetable crops, drug plant
crops and ornamental crops.
(2) Human Diseases:
Several fungi are responsible formycoses and tropical diseases of man
and animals.
(3) Destruction of timber and timber products:
Several fungi like Polyporus, Chloros-plenium, Penicilium divaricatum,
Fusarium negundi. Pori microspora, Lentinus lepidens are responsible
for decay of standing trees, decay of felled timber, sap stains and decay
of wood in various uses.
(4) Food Spoilage:
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Besides microorganisms, members of Mucorales (e.g., Rhizopus,
Mucor), yeasts, Moniliales, Aspergillus oryzae. Penicillium digitatum
are chief cause of food spoilage. These are saprophytic fungi which
grow on food articles such as bread, jam, pickles, meat etc.For instance,
food items are spoiled by yeasts, Aspergillus otyzae, Penicillium
digitatum etc.Dairy products are spoiled by Aspergillus repens, Mucor,
Penicillium, Cladosporium, Oiduni lactis, etc.
(5) Mushroom poisoning:
Deadly poisonous toadstools such as Amantia sp and others have
invariably been mistaken for edible mushrooms and have taken a toll of
life. Mushroom poisoning is known as mycetimus.

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 EXPERIMENT

Aim: to study and ascertain the necessary conditions required for the
sporulation of fungus on bread.
Materials required: loafs of bread (4 in number), water (in a sprayer),
plastic bags (4), gloves, rubber bands and marker pen.
Procedure:
 We have taken 4 loafs of bread in a dish and with the help of water
sprayer added a layer of moisture (water) onto the surface of the
bread slices.
 We have used the plastic bags at our disposal and inserted the
bread into the bags.
 We sealed the mouth of those plastic bags with the help of rubber
bands.
 Now we marked each of the prepared bread loaf bags as A B C D
with the help of marker pen.
 A was placed in a positon inside our house with access to direct
sunlight.
 B was placed in a position with access to diffused light.
 C was placed in a position with access to no light at all(complete
darkness)
 D was used as a control setup and placed inside the
refrigerator(cold and dry condition).
 We note that all the places were hot and humid condition with the
exception of D.

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Observing bread samples at the end of 4th day.

A. has clear signs of fungal B. has little to no signs of

growth. fungal growth.

C. shows few signs of fungus D. has no change

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Observation
The setups have been observed after 96 hours (4th day) and
after 168 hours (7th day).

1. DAY 0 (beginning of observation):

Setups look normal, no change.

2. DAY 4 (middle of observation):

A. Setup has clear signs of fungal growth (white

mycelium) with a slight amount of visible spore
formation (dirty green and black colour) albeit the
growth and spread is less in amount.

B. Setup has very less to no signs of fungal growth

with just some strands of visible white mycelium.

C. Setup has some signs of growth of fungus with a

few traces of spore formation.

D. Setup has no change.

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Observing bread samples at the end of 7th day (end).

A. Sporulation increased B. substantial fungal growth and

no increase in mycelium. mycelium covers whole surface.

C. overwhelming growth. D. no change

Spores cover whole surface of
Bread.

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3. DAY 7 (end of observation):

*All setups have clear signs of fungal growth

A. The amount of growth of fungi has not increased.

The sporulation has increased. The amount of
mycelium remains same.

B. The amount of growth of fungi has substantially

increased with the white mycelium now covering
even walls of the dish used. The amount of spores
and mycelium have both increased substantially.

C. Overwhelming growth of spores and mycelium is

observed. Now the spores cover the whole of the
bread surface and even the underside of the bread.
Only green colour is visible in this sample and the
spores leave the surface in the form of a thin mist
when disturbed.

D. No change is observed. (bread became little old)

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 Conclusion: After 7 days, the bread loaf kept in the dark (C) had
the highest amount of growth of fungi and sporulation. The
diffused light bread (B) had moderate growth of fungus and the
direct sunlight one (A) had least amount of growth. It must be
noted that all of the breads had substantial growth of fungus. The
exception is the refrigerated bread (D) which had no change or
growth.

Precautions:
1. Care has to be taken to supply adequate amount of water to
the breads and not overwater the loafs
2. During observation, while handling the loafs we must
always wear gloves.
3. We always wash our hands after handling the breads.

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 CONCLUSION

After completing the given investigatory project and performing the

experiment we now have a brief idea as to what conditions the fungi
requires in order to grow and thrive on common household items like
bread etc. We have concluded from the experiment that the fungi
requires a hot humid and dark environment to thrive properly. We have
also seen that fungi can and will grow in direct sunlight and diffused
light as well though it does not grow in such intensity as when kept in a
dark environment. We also have studied how to prevent fungi from
appearing on food materials by storing them in refrigerator.

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 BIBLIOGRAPHY

The following sources were used in the making of this project:

 NCERT class XI biology text book
 Wikipedia
 Brittanica.com
 Moderns ABC of biology

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