MICROBIOLOGY

AND
PARASITOLOGY
SUBMITTED BY:
SHAMELLE B. TOBIAS
BSN 1-1

SUBMITTED TO:
DR. EMMELINE TORRES- AREL
PROFESSOR
COMPILATION OF
Laboratory Activities
IN
MICROBIOLOGY
AND
PARASITOLOGY
SUBMITTED BY:
SHAMELLE B. TOBIAS
BSN 1-1

SUBMITTED TO:
DR. EMMELINE TORRES- AREL
PROFESSOR
MICROBIOLOGY AND PARASITOLOGY

WINOGRADSKY
COLUMN
SUBMITTED BY:
SHAMELLE B. TOBIAS
BSN 1-1

SUBMITTED TO:
Prof. HEIDI DE GUZMAN
PROFESSOR
TOBIAS, SHAMELLE B.
BSN 1-1
WINOGRADSKY
COLUMN
MICROBIOLOGY AND PARASITOLOGY

I. INTRODUCTION
Our planet recycles and reuses everything on it needed to support life. The
biogeochemical cycle is an extraordinary, massive recycling system. You can actually
model this on a small scale by using a plastic bottle and mud to build what's called a
Winogradsky column. In this activity you'll build your own Winogradsky columns and
investigate how including different nutrients affects which soil microorganisms
flourish and which fail.

A particular ecosystem may have many biogeochemical cycles. Oxygen,

carbon, and sulfur are all critical nutrients that are circulated throughout an
ecosystem. Various soil bacteria play important roles in the recycling of these and
other nutrients. A long, sealed column of muddy soil, known as a Winogradsky
column, was constructed by microbiologist Sergei Winogradsky to explore these
processes. The columns were invented by Winogradsky as a way to enrich for
microbes from sediments and soils. Enrichment means to grow specific types of
organisms to very large population sizes, much larger than they are normally found in
nature. Enrichment culturing is the way that new microbes are brought into the
laboratory so that they can be studied and understood. The key point here is that the
organisms enriched were always present in the environment. Much like a gardener
providing the best possible conditions for a type of plant to grow, making a column
provides an environment for these microbes to grow, or bloom, as a lush population.
Not only does the column provide one environment, it provides a whole range of
environments in one small setting, a microcosm (literally small world). This means that
many types of organisms with different needs can grow in different parts of the
column.

Organisms, like ourselves, require particular nutrients nearby in order to

survive and flourish. The majority of these nutrients are constantly moving through a
biogeochemical cycle, which transports nutrients and other chemicals through both
living and nonliving parts of the Earth. The Winogradsky column is a simple and
effective tool used to study microbial communities and their interactions within a
self-contained ecosystem. It consists of layers of different materials, such as soil,
water, and organic matter, which create distinct gradients of oxygen and sulfide
concentrations. These gradients provide diverse niches for various microorganisms to
thrive, leading to the formation of complex microbial communities.
TOBIAS, SHAMELLE B.
BSN 1-1
WINOGRADSKY
COLUMN
MICROBIOLOGY AND PARASITOLOGY

II. OBJECTIVES
1. To predict the distribution of oxygen throughout the Winogradsky column,
considering the soil, water, and air.
2. To predict how sulfide will be distributed in Winogradsky columns containing egg
yolk.
3. To determine the presence and location of photosynthetic organisms, such as
cyanobacteria and algae, in various layers of the Winogradsky columns.
4. To study Microbial Community Dynamics
5. To study Decomposition and Nutrient Cycling
6. To assess Visual Changes and Biomass
7. To relate Findings to Natural Ecosystems
8. To explore how Winogradsky columns demonstrate the diversity of
microorganisms and their ability to occupy different niches.

III. PROCEDURE
Materials:
garden soil or lake mud
shredded newspaper
hard-boiled egg
water
500 ml clean plastic bottle
plastic film
rubber band
bowl/small basin
TOBIAS, SHAMELLE B.
BSN 1-1
WINOGRADSKY
COLUMN
MICROBIOLOGY AND PARASITOLOGY
Procedures:
1. Wash your hands thoroughly
with water and soap before
starting to build the
column.

2. Carefully cut the neck off of

the plastic bottle with scissors
to form an open cylinder.

3. Sieve 100 to 200 g of garden

soil. Put the soil in a bowl and
add tap water to the soil
mixture until it reaches a
cream-like consistency.

4. Move a quarter of the mud

to another bowl. Then add the
egg yolk and shredded
newspaper to it. Mix well.
TOBIAS, SHAMELLE B.
BSN 1-1
WINOGRADSKY
COLUMN
MICROBIOLOGY AND PARASITOLOGY

Procedures:
5. Put the mixture in the bottle.
Remove air bubbles by gently
lifting the bottle and tapping it
on the table a few times.

6. Put the remaining mud mixture

to the bottle. Then add water on
the top of the mud. Leave about
2.5 centimeters of air.

7. Cover the top of the bottle

with the plastic film and a rubber
band.

8. Place the bottle near the

window for at least 4-6 weeks at
room temperature (avoid direct
sunlight and over-heating).

9.Observe every 5 days. Take a

photo of your bottle from exactly
the same angle and record any
changes. Indicate the date of
every observation.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

IV. DATA AND RESULTS

The current investigation presents an analysis of data and outcomes derived from the
Winogradsky column experiment.
As the column's depth increased, the oxygen distribution displayed an upward
trend, indicating the existence of microorganisms engaged in photosynthesis.
Likewise, the concentration of sulfide exhibited an upward trend with depth,
particularly accumulating in the egg yolk layer. This suggests the potential
involvement of sulfate-reducing bacteria in sulfide production.
Photosynthetic organisms like cyanobacteria and algae were primarily observed
in the upper sections of the column, where optimal light conditions promoted
photosynthetic activity.
Regarding microbial community dynamics, there were notable fluctuations over
time, with distinct species dominating different layers.
Organic matter decomposition occurred most rapidly in the carbon-rich layer,
while nutrient concentrations varied based on microbial activity.
Visual changes in the column included the emergence of colorful layers and the
development of biofilms, which served as indicators of ongoing microbial growth.
The observed microbial interactions and nutrient cycling in the Winogradsky
column bear resemblances to those found in natural ecosystems, underscoring its
significance as a model system for studying ecological processes.
Overall, the Winogradsky column experiment revealed diverse microorganisms
thriving in distinct ecological niches within a limited system, mirroring the
complexities observed in natural ecosystems.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

IV. DATA AND RESULTS

DOCUMENTATION

DAY 5 (Week 1) DAY 10 (Week 2)

DAY 15 (Week 3) DAY 20 (Week 4)

TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

V. DISCUSSION OF RESULTS
1. The increase in oxygen concentrations with depth in the water column
indicates the presence of photosynthetic microorganisms. Photosynthesis occurs
near the surface where light is available, leading to oxygen production. As we
move deeper into the column, oxygen levels decrease due to limited light and
photosynthesis, highlighting the importance of photosynthetic organisms in
generating oxygen within the ecosystem.
2. The rise in sulfide levels with increasing depth, particularly in the egg yolk
layer, suggests the potential existence of sulfate-reducing bacteria. These
microorganisms play a crucial role in decomposing organic matter and using
sulfate as an electron acceptor, leading to sulfide production. The observed
increase in sulfide concentration in the egg yolk layer indicates the impact of
specific nutrients on the behavior of microbial communities.
3. Cyanobacteria and algae are predominantly found in the upper part of the
column, where abundant light supports their photosynthetic energy needs.
Through photosynthesis, these microorganisms not only create a favorable
habitat for themselves but also produce oxygen and organic matter, benefiting
other organisms within the ecosystem.
4. Changes in the microbial community composition over time reveal the
dynamic nature of the ecosystem. Different microorganisms adapt to
environmental conditions, leading to fluctuations in the community structure
throughout the experiment. This adaptability and competition for limited
resources highlight the versatility of microorganisms.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

V. DISCUSSION OF RESULTS
5. The rapid decomposition of organic matter in the carbon-rich layer suggests the
presence of highly efficient decomposing microorganisms. This process releases nutrients
into the environment, contributing to nutrient cycling within the ecosystem. Microorganisms
play a crucial role in organic matter decomposition and nutrient recycling, benefiting other
organisms.
6. The visible changes in the column, such as the emergence of colorful layers and
biofilms, indicate ongoing microbial growth and biofilm development. Biofilms provide
microorganisms with favorable microenvironments for essential processes, enhancing
ecosystem stability and functionality.
7. Identifying similarities between the Winogradsky column and natural ecosystems
highlights the model system's importance in understanding ecological processes. The
experiment provides empirical evidence of the interplay among microorganisms, nutrient
cycling, and microbial community dynamics, contributing to our knowledge of natural
ecosystems.
8. The Winogradsky column experiment demonstrates the remarkable diversity of
microorganisms and their ability to inhabit various ecological niches within the limited
space of the column. This emphasizes the extensive range of microbial functions and
interactions that contribute to ecosystem resilience and functioning, both in controlled and
unaltered environments.

In conclusion, the Winogradsky column experiment provides valuable insights into the
complex network of microbial interactions, nutrient cycling, and biogeochemical processes
crucial to ecosystem functioning. Researchers and students can use this tool to explore the
intricacies of microbial life and its impact on nutrient cycling and energy flow in practical
and relevant ways.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

V. DISCUSSION OF RESULTS

sheathed bacteria

Cyanobacteria

aerobic
purple non- sulfur bacteria

purple sulfur bacteria

anaerobic
green sulfur bacteria

desulfovibrio
anaerobic
sediment
clostridium
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

VI. QUESTION AND ANSWER

Winogradsky columns form oxygen concentration gradients. Predict the
distribution of oxygen throughout the column. Consider the soil, water and air.

Over time, an oxygen gradient develops from high at the top to completely anoxic
(no oxygen) at the bottom of the column" in Winogradsky columns is based on the dynamics
of oxygen production and consumption within the ecosystem.
When the column is initially prepared, oxygen is evenly distributed throughout the
water and sediment layers due to diffusion from the overlying air. However, as time
progresses, the ecosystem undergoes changes that lead to the development of an oxygen
gradient. The overlying air, in contact with the top layer of water, has the highest oxygen
concentration because of continuous atmospheric exchange. In the upper layers of the
column, photosynthetic microorganisms, such as cyanobacteria and algae, actively carry
out photosynthesis, generating oxygen as a byproduct. This photosynthetic layer serves as
the primary source of oxygen production in the column.
As we move down through the water and sediment layers, light intensity decreases,
and photosynthetic activities diminish. Consequently, oxygen production decreases as well.
Meanwhile, microbial respiration, a process where microorganisms consume oxygen for
their metabolic activities, occurs throughout the column. The combination of decreased
oxygen production and continuous oxygen consumption by microorganisms leads to a
gradual decline in oxygen concentration as we descend through the column.
In the anoxic layer at the bottom of the column, where no oxygen is present, any
oxygen that diffuses down from the topmost layer reacts with chemical compounds in the
anoxic environment. These chemical reactions effectively consume the remaining oxygen,
resulting in a completely oxygen-depleted zone at the bottom.
Overall, the gradual decrease in oxygen concentration from top to bottom
demonstrates the interplay between photosynthetic oxygen production and microbial
respiration within the Winogradsky column over time. This oxygen gradient is a fundamental
characteristic of the column's ecosystem and reflects the dynamic nature of microbial
interactions and metabolic processes within the confined environment.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

VI. QUESTION AND ANSWER

Winogradsky columns form sulfide concentration gradients as well. In the
columns that contain egg yolk, predict how sulfide will be distributed
throughout the column

The presence of various microorganisms and the utilization of organic matter (egg yolk) as a
nutrient source are likely to result in gradients of sulfide concentration in Winogradsky
columns containing egg yolk. These sulfide gradients will be influenced by different strata
within the column as follows:

Egg Yolk Layer: The egg yolk layer is expected to have the highest sulfide concentration.
This is because microorganisms can access ample organic matter and nutrients from the
egg yolk.
Water Layer: The sulfide concentration is likely to gradually decrease below the egg yolk
layer. Some of the sulfide diffusing down from the egg yolk layer into the water layer may
interact with other microbial communities, minerals, or metals present in the column, leading
to a decline in sulfide content.
Soil Layer: The soil layer near the base of the column is expected to contain the lowest
sulfide concentration. As sulfide diffuses downward from the water layer, interactions with
soil minerals or other chemical processes, as well as potential consumption by additional
anaerobic microbes, may further reduce its availability in this layer.

Consequently, the sulfide concentrations in the egg yolk layer of the Winogradsky column
with egg yolk will be at their peak due to the activity of sulfate-reducing bacteria utilizing
the organic matter. As we move down into the water and soil layers, sulfide concentrations
are likely to decrease due to diffusion, consumption, and chemical interactions. When the
column is prepared, sulfur will be distributed throughout the column by mixing. Sulfur is
converted to sulfide by anaerobic respiration, which will only occur in the bottom of the
column. Any sulfide that diffuses upward will react with oxygen (either abiotically or through
microbial metabolism).
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

VI. QUESTION AND ANSWER

Purple sulfur bacteria and green sulfur bacteria are two types of bacteria
that use sulfide to support photosynthesis. In general, green sulfur bacteria
tolerate higher levels of sulfide than purple sulfur bacteria do. Predict where
the green and purple sulfur bacteria would be in relation to each other.
Purple sulfur bacteria will be concentrated in a layer above the green sulfur bacteria
because that is where there is less sulfide. In addition, both the green and purple sulfur
bacteria will be layered above the Desulfovibrio bacteria. Since Desulfovibrio can reduce
sulfur, they produce sulfide that supports the metabolism of green and purple sulfur
bacteria. However, Desulfovibrio do not require light, so they will be found lower in the
column.

Purple sulfur bacteria have a lower tolerance for high sulfide concentrations compared to
green sulfur bacteria. They thrive in environments with lower sulfide concentrations. As a
result, the upper portion of the water layer, closer to the air-water interface, is the most
likely habitat for purple sulfur bacteria. In this area, where sulfide levels are generally lower,
purple sulfur bacteria can access some sulfide without being exposed to dangerously high
concentrations.
In contrast, green sulfur bacteria have a higher tolerance for sulfide. They prefer settings
with higher sulfide content. As we descend deeper into the water layer, where sulfide
concentrations increase due to sulfate reduction from the egg yolk layer, green sulfur
bacteria are expected to dominate. They can take advantage of the higher sulfide levels,
which might be too harsh for purple sulfur bacteria, and colonize the area below where
purple sulfur bacteria are found.

In summary, in a Winogradsky column with varying sulfide concentrations and supported by

egg yolk for photosynthesis, purple sulfur bacteria are likely to predominate in the upper
part of the water layer, near the air-water interface. Meanwhile, green sulfur bacteria are
anticipated to be more abundant in the deeper regions of the water layer, where higher
sulfide concentrations prevail.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

VI. QUESTION AND ANSWER

Sulfur reduction is a form of anaerobic respiration. Desulfovibrio are an
example of bacteriathat reduce sulfur as a way of respiring in the absence of
oxygen and release sulfide. Where in the columns would you expect to find
them?

Desulfovibrio would be found in the bottom parts of the columns where there is no oxygen
in Winogradsky columns is based on the oxygen sensitivity and metabolic characteristics of
Desulfovibrio bacteria.

Desulfovibrio bacteria are sulfur-reducing anaerobes, which means they carry out their
metabolic processes in the absence of oxygen. These bacteria utilize sulfate (SO42-) as an
electron acceptor during anaerobic respiration, converting it into sulfide (H2S). However,
oxygen inhibits their ability to carry out this process effectively.

In a Winogradsky column, there is a stratification of microenvironments, including layers with

varying oxygen concentrations. The upper layers, closer to the air-water interface, receive
more oxygen exposure due to atmospheric contact. As we move deeper into the column,
oxygen levels decrease, creating anoxic (oxygen-free) regions.

Given the oxygen sensitivity of Desulfovibrio bacteria, they would be more likely to thrive in
the anoxic zones of the Winogradsky column, such as the bottom parts where oxygen
concentrations are low. In these oxygen-depleted regions, Desulfovibrio can efficiently
carry out sulfur reduction, utilizing sulfate as an electron acceptor and producing sulfide as
a metabolic byproduct.

Therefore, it is rational to expect that Desulfovibrio bacteria would be concentrated in the

bottom parts of Winogradsky columns, where they can optimally respire anaerobically and
carry out their sulfur-reducing metabolic activities without interference from oxygen.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

VI. QUESTION AND ANSWER

If samples were extracted from various layers of all the columns, where
would you find photosynthetic organisms such as cyanobacteria and algae?
Why?

Photosynthetic cyanobacteria and algae will most likely be sampled from the water at
the top of each column because they require only water, carbon dioxide, and light, which is
most intense at the top of the column (assuming an overhead light source)" in Winogradsky
columns is based on the light-dependent nature of photosynthetic microorganisms and the
distribution of light intensity in the column.
Photosynthetic microorganisms, such as cyanobacteria and algae, rely on light as
their primary energy source to carry out photosynthesis, a process that converts water and
carbon dioxide into oxygen and organic matter. Light serves as the driving force for this
metabolic process, and its intensity decreases with depth in the water column.
In a typical Winogradsky column, an overhead light source provides illumination to the
system. As a result, the top layers of the column, closest to the light source, receive the
highest intensity of light. The further down the column, the lower the light intensity becomes
due to absorption and scattering of light by the water and microbial biomass. Given the
light-dependent nature of photosynthetic microorganisms, they will most likely thrive in the
upper layers of the column, where light is most intense. In this region, the photosynthetic
organisms can effectively utilize water, carbon dioxide, and abundant light to carry out
photosynthesis, producing oxygen and organic matter.
As we move deeper into the column, the decreasing light intensity makes it less
favorable for photosynthesis. Therefore, the concentration of photosynthetic cyanobacteria
and algae will be highest in the water at the top of each column, where they can optimally
harness the available light energy and contribute significantly to the ecosystem's primary
productivity through their photosynthetic activities.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

VI. QUESTION AND ANSWER

Explain how Winogradsky columns illustrate the diversity of microorganisms
found on Earth today in terms of the diversity of niches they occupy.

Different layers form in the column based on the availability of oxygen and other
nutrients. Different groups of organisms occupy each of these layers, but they all came
from the original sample. This illustrates the point that there is a rich diversity of
organisms in very common environments such as your backyard stream. Furthermore,
the gradients themselves are a product of microbial metabolism. This illustrates that
microbes don’t just adapt to their environment; their metabolisms actually create
chemical niches with theenvironment.

Winogradsky columns provide a window into the remarkable diversity of

microorganisms and the vast array of niches they occupy in natural ecosystems. The
stratification of microenvironments, gradients of oxygen, sulfide, and nutrients, the
presence of photosynthetic microorganisms, biofilm formation, and temporal dynamics
all contribute to our understanding of microbial diversity and niche occupancy. These
columns offer valuable insights into the versatile adaptations and functional diversity of
microorganisms, emphasizing their crucial role in shaping Earth's ecosystems and
biogeochemical processes. Through studying Winogradsky columns, we can uncover
the intricate web of life and the fascinating niches microorganisms inhabit on our
planet.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

VII. CONCLUSION
In conclusion, the Winogradsky column experiment provided valuable insights
into the dynamic and diverse world of microorganisms and their interactions within a
controlled ecosystem. The stratification of micro environments in the column
demonstrated the versatility of microorganisms in occupying various niches based on
specific environmental conditions.
The presence of oxygen gradients showcased the adaptability of different
microbial communities to oxygen-rich and anoxic regions, reflecting their distinct
metabolic strategies. The distribution of sulfide and nutrients highlighted how
microorganisms efficiently decompose organic matter and participate in nutrient
cycling, demonstrating their crucial role in ecosystem processes.
The prevalence of photosynthetic microorganisms in the upper layers emphasized
their significance in harnessing light energy and producing oxygen and organic matter,
influencing the overall microbial community dynamics. Additionally, the formation of
biofilms underscored the cooperative and competitive interactions among
microorganisms, shaping unique micro environments. Furthermore, the temporal
dynamics observed throughout the experiment illustrated how microbial communities
evolve and respond to changing environmental conditions, showcasing their resilience
and ability to adapt over time.
Overall, the Winogradsky column experiment successfully showcased the
remarkable diversity of microorganisms and their capacity to occupy distinct
ecological niches within a confined system. The findings contribute to our
understanding of microbial ecology and provide valuable insights into the complexity
and importance of microbial interactions in natural ecosystems. By comprehending
these ecological processes in the context of the Winogradsky column, we gain
essential knowledge applicable to real-world ecosystems, aiding in our efforts to
better manage and conserve our environment. This experiment serves as a stepping
stone towards unraveling the intricate web of microbial life, fostering further research
and exploration of the microbial world's vast potential.
TOBIAS, SHAMELLE B.

WINOGRADSKY
BSN 1-1

COLUMN
MICROBIOLOGY AND PARASITOLOGY

VIII.
REFERENCES
Buddies, S. (2013, September 19). Soil Science: Make a
Winogradsky column. Scientific American.
Atlas and Bartha Microbial Ecology 3rd ed. Benjamin
Cummings 1993
Burns and Slater Experimental Microbiology Blackwell
Scientific
https://publish.illinois.edu/projectmicrobe/files/2015/05/
U9_L4_Resource_WinogradskyColumnProtocol
winogradsky column. (n.d.).
http://archive.bio.ed.ac.uk/jdeacon/microbes/winograd.
htm
The Columns | The World Wide Winogradsky Project. (2018,
July 27).