Microbial Discovery Activity
“I Can’t Live Without You!”
A Close-up Examination of
Microorganisms Involved in
Mutually Beneficial Symbiotic Relationships
Author
Robin Patterson
Butler County Community College
Butler, PA 16001
robin.patterson@bc3.edu
robinp@connecttime.net
Contributors
Michelle S. Thomas
Campbell University
Department of Biological Sciences
Buies Creek, NC 27506
suhan@campbell.edu
Susan Deines
Colorado State University
Department of Microbiology, Immunology and Pathology
Ft. Collins, CO 80523
susan.deines@colostate.edu
Intended Audience
K-4
5-8
9-12
X
X
Activity Characteristics
Classroom setting
Uses hands-on manipulatives
Requires special equipment
Can be performed individually
Appropriate for students with special needs
Requires more than one class period
American Society for Microbiology
Education Department
1752 N Street, NW
Washington, DC 20036
EducationResources@asmusa.org
X
X
X
X
X
X
Introduction
Description
In this activity, students learn about mutually beneficial symbiotic relationships that exist between
microbes and other microbes, plants, and animals, prepare samples for microscopic observation from a
termite hindgut, a lichen, and a legume root nodule, and use a microscope to observe and identify the
microbial symbionts in each of the samples.
Abstract
By visiting three stations, each equipped with microscopes, slides, live materials and various supplies
arranged by the teacher, the students will observe three symbiotic relationships involving microbes.
The students prepare wet mounts to observe the microbial symbionts found in the termite gut, lichen,
and Rhizobium root nodules.
Core Themes Addressed
General Microscopy Concepts
Microbial Cell Biology
Microbial Genetics
Microorganisms and Humans
Microorganisms and the Environment
Microbial Evolution and Diversity
Other -Common properties of life;
Cellular components
X
X
Keywords
Ecology; Microbial Ecology, Endosymbiosis, Evolution, Microbial Interactions
Learning Objectives
At completion of this activity, the learner will be able to:
• describe different types of symbiotic relationships.
• identify three mutually symbiotic relationships involving a microbe and another microbe, an
animal or a plant.
• describe the microbial symbionts in the termite gut, a lichen, and a legume.
• identify how these symbiotic relationships benefit both partners.
• transfer and apply the principles presented in this activity to other examples of symbiosis.
• prepare a wet mount slide of termite hindgut to observe endosymbionts.
• Use the squash technique to prepare lichen and root nodules.
ASM Microbial Discovery Activity
Symbiotic Relationships – Page 2
National Science Education Standards Addressed
All Levels of Learning:
1. Unifying concepts and Processes:
Systems, order and organization
• Systems – The organisms involved in mutually beneficial symbiotic relationships function
together as a whole.
• Organization – Mutually beneficial symbiotic relationships are living systems in which
the various partners interact with one another in a type of community.
Evidence, models and explanations
• Evidence and explanations – This activity requires the learner to make microscopic
observations of various organisms involved in mutually beneficial symbiotic relationships
and then develop a scientific explanation for their roles in these relationships.
Change, consistency and measurement
• Change – In many cases of mutually beneficial symbiosis, the microbial symbiont
generates energy through the conversion of a substrate to another form, and shares the
energy or the resulting product with the other partner in the relationship.
Form and function
The various partners involved in mutually beneficial symbiotic relationships have specific
roles. Form may be influenced by the specific requirements of these organisms, such as the
formation of root nodules by legumes to provide an anoxic environment for anaerobic
nitrogen-fixing bacteria.
Science Content Standards 5- 8:
1. Life Science: Content Standard C
• Regulation and behavior – Demonstrates how organisms obtain and use resources
symbiotic relationships
through
•
Populations and ecosystems – Demonstrates how populations of different organisms live
together and interact with one another
•
Diversity and adaptations of organisms - Demonstrates how endosymbiotic relationships are
biological adaptations that enhance survival of the associated partners in a particular
environment
Science Content Standards 9-12:
1. Life Science: Content Standard C
• Biological Evolution Demonstrates how symbiotic relationships have evolved over time and
are maintained because they provide a biological advantage to both partners
•
The interdependence of organisms – Demonstrates how microbes recycle atoms and molecules,
how organisms cooperate in an ecosystem, and how microbes form interdependent
relationships with other organisms.
ASM Microbial Discovery Activity
Symbiotic Relationships – Page 3
Teacher Handout
I Can’t Live Without You!”
A Close-up Examination of Microorganisms
Involved in Mutually Beneficial Symbiotic
Relationships
Student Prior Knowledge
The students should be familiar with the classification of different types of microbes (specifically,
bacteria, algae, and protozoa) and the definition of symbiosis and the different types of symbiotic
relationships, and the terms used to describe various oxygen requirements (specifically strict aerobe,
facultative anaerobe, aerotolerant anaerobe, and strict anaerobe). The students should have a
fundamental understanding of the basic nutritional requirements for living organisms, the function of
enzymes, and the process used in biological nitrogen fixation. Additionally, students should have basic
microscopy skills.
Teacher Background Information
Symbiosis describes the relationship between two or more organisms. This relationship can occur
between two different types of microbes, between a microbe and a plant, or between a microbe and an
animal. When both partners depend on each other and benefit from one another the relationship is
termed mutualism. Should one partner gain an advantage, and the other partner is neither harmed
nor benefits from the presence of the partner, commensalism is used to describe the relationship. In
parasitism, only one of the symbiotic partners benefits; the other partner suffers. Sometimes the
symbiotic partners are physically associated with one another. If one of the partners lives inside the
cell of the other partner, the relationship is called endosymbiosis. In ectosymbiosis, the partners
may not be physically associated or one partner may exist in or on the body of the other partner, but
not within its cells.
Endosymbiosis examples:
- microbe/microbe: Paramecium bursaria/photosynthetic green algae
- plant/microbe: legume(clover)/nitrogen-fixing Rhizobium bacteria
Ectosymbiosis examples:
- animal/microbe: ruminants (cows, sheep, buffaloes, etc.) /cellulase producing
anaerobic microbial community (protozoa and bacteria)
- microbe/microbe: lichen (fungus/ photosynthetic bacterium or alga)
- microbe/microbe: sulfur cycling by bacteria
- insect/microbe: leaf-cutting ants/fungi
The three examples of symbiosis explored in this exercise are observed using preparations made from
termite hindguts, lichens, and legume root nodules.
Termites: Termite guts are the world's smallest bioreactors. The hind-gut of wood-feeding termites
harbor mainly aerotolerant lactic acid bacteria, facultatively anaerobic enterobacteria, and even a
significant fraction of strictly aerobic bacteria, in addition to the larger anaerobic protozoa, such as
Trichonympha spp. These indwelling microbes degrade the wood cellulose into sugar, and the sugar is
then consumed by the termite. Interestingly, some of the protozoa living within the termite have
ASM Microbial Discovery Activity
Symbiotic Relationships – Page 4
bacterial endosymbionts of their own! For example, some of the protozoa have no mitochondria; they
rely on their bacterial endosymbionts to generate the energy (ATP) they need.
Lichens: Lichens are a successful alliance between a fungus and a photosynthetic alga or bacterium.
The fungus partner provides a home for the alga or bacterium; the alga or bacterium produces food
(organic compounds) for the fungus through photosynthesis. Only certain algae and certain fungi, or
certain bacteria and certain fungi, can get together to form a lichen. This means that each union
creates a unique type of lichen body (thallus). Scientists identify and assign names to lichen based on
their observations of the thallus. Several features are used to characterize lichens, including body type,
life form, lobules, and surface texture. (Lobules are small outgrowths, usually from the edge or margin
of the lichen. They are important for reproduction.) Lichens occur in one of four basic growth forms:
1.
2.
3.
4.
crustose - crustlike, growing tight against the substrate
squamulose - tightly clustered and slightly flattened pebble-like units
foliose – leaf-like, with flat sheets of tissue not tightly bound
fruticose - free-standing branching tubes
The true identity of lichens as symbiotic associations of two different organisms was first proposed by
Beatrix Potter, who is best remembered for her children's books about Peter Rabbit. In addition to her
books, she spent time studying and drawing lichens. Her illustrations are still appreciated for their
detailed and accurate portrayal of the delicate beauty of these bizarre organisms.
Legumes: A well known symbiotic relationship is that between legumes, such as peas, beans,
soybeans, clover, alfalfa, and peanuts, and soil bacteria in the genus Rhizobium. The rhizobia, which
exist in nodules on the roots of the legume, can fix nitrogen gas when the symbiosis is achieved. They
can't fix nitrogen in the free-living state, however. (Legume/Rhizobium nodules are red. This is
due to the production of leghemoglobin which sequesters oxygen. This helps to create an anoxic
[oxygen-free] environment for the strictly anaerobic Rhizobium bacteria.)
Fixed nitrogen, in the form of ammonia, is used as a nitrogen source by the legume and the
Rhizobium gets a nice place to live with all of the amenities, including photosynthate, water and
minerals! This is an extremely important relationship because nitrogen is usually the most limiting
element in terrestrial ecosystems. Furthermore, many legumes, like soybean, form the basis for
agriculture on a world-scale. Some legume seeds, like soybeans, contain high levels of protein. These
are the most important agricultural sources of protein in the world.
Class Time
•
•
•
Approximately two 45-minute class periods are required to make and observe all three
preparations.
The amount of waiting time will depend on the number of microscopes available for use.
Depending on the amount of time available, students may visit the stations in teams or
individually.
The instructor may wish to follow-up in a subsequent period with a discussion of these
activities.
Teacher Preparation Time
Teacher preparation time will vary depending on approach. Generally, one hour of preparation time is
required in order to acquire the various specimens and to set up all three stations.
Materials That Need to Be Prepared in Advance
• Termites can be ordered from a biological supply house (check regulations regarding shipment
to your state or school) or can be captured if you live in a suitable environment (one that is not
too dry).
Ordering Termites
Ward’s Natural Science - Zootermopsis angusticollis 30 for $21.99
http://www.wardsci.com/product.asp_Q_pn_E_IG0013506_A_Termites+Living+Specimen
ASM Microbial Discovery Activity
Symbiotic Relationships – Page 5
1-800-962-2660
Capturing termites
Cut 4" x 4" squares of corrugated cardboard and stack. Secure with 2 rubber bands and place outside
under a rotting log for a few weeks. Alternatively, examine logs for termites by peeling off the outer
bark. Often, termites can be found beneath the bark and easily collected. Otherwise, you may need to
pry the wood apart to expose the termites for collection. Termites can be maintained in the classroom
by keeping in a small, loosely capped jar with a source of cellulose (wood shavings).
•
Lichen can be gently scraped from a rock or tree. If you are lucky to live in an area where many
types of lichen grow, collect a variety of specimens.
•
Clover or other legumes can be pulled from the ground.
Materials and Equipment
Station 1: Termite Hindgut
Termites
Microscope slides
Dissecting needles or probes
Forceps
Plastic pipette
Coverslips
Bottle of immersion oil with dropper (for observation at 100x magnification)
Beaker of 1:10 bleach solution for disposal of slides and dead termites
Microscope
Lens tissues
Station 2: Lichen
Fresh lichen samples
Microscope slides
Plastic pipette
Forceps
Coverslips
Dropper bottle of water
Microscope
Lens tissues
Beaker for disposal of slides
Station 3: Root Nodules
Legume (e.g. clover) root nodules
Scalpel or knife
Microscope slides
Coverslips
Dropper bottle of water
Microscope
Bottle of immersion oil with dropper (for observation at 100x magnification)
Lens tissues
Beaker for disposal of slides
Safety Precautions
The microorganisms used in this activity are naturally found in the environment. It is always
important to observe caution when using environmental samples. Exercise care when preparing
squash preparations and when using probes.
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Methods
Procedure for preparing the specimens: The preparation of the specimens should be demonstrated
to the students. The students may then prepare their own specimens as they move through the
different symbiosis stations.
1. Demonstrate the technique needed to remove the hindgut of the termite.
a. Obtain a microscope slide, coverslip, compound microscope and two needles or sharp probes
from the supply area.
b. Termite dissection
Method I:
1. Use the forceps to obtain a termite and hold it on the microscope slide.
2. Using the two needles, place one on the front end of the termite and one on the
abdomen. When these needles are pulled in opposite directions, the alimentary canal
will open and reveal the milky-colored gut fluid, which contains a wealth of microbes
within.
3. Use the pipette to place a drop of distilled water on the slide near the alimentary canal.
This will allow the protozoa to swim from the alimentary canal into the water and
allow the students much easier viewing.
4. Quickly cover the drop of water/gut fluid with a coverslip to preserve an anoxic
environment for the anaerobic microbes living in the gut.
Method II:
1. Use the pipette to place a drop of water on the slide.
2. Use the forceps to grasp the front end of the termite and use a second pair of forceps to
gently milk the hindgut, allowing the hindgut contents to drop on to the slide into the
drop of water.
3. Quickly cover the drop of water/gut fluid with a coverslip to preserve an anoxic
environment for the anaerobic microbes living in the gut.
c. Observe the preparation under 10X and 40X magnification. If the compound microscopes are
equipped to operate using the darkfield technique, Trichonympha spp. can be seen with
great ease.
d. To observe the motion of the protozoal flagella and bacterial endosymbionts, place a drop of
immersion oil on the coverslip and observe under 100X magnification.
2. Demonstrate the squash technique for observing lichen:
a. Use the pipette to place a drop of water on the slide.
b. Use the forceps to pick up a small piece of lichen and place it in the drop of water.
c. Use the short end of a microscope slide to grind up the material in the water.
d. Place a coverslip over the drop of water/lichen material.
e. Observe under 10X and 40X magnification. Look for oval or round green algal cells and
filamentous fungal cells.
3. Demonstrate the squash technique for observing the root nodules:
a. Use a scalpel or knife to cut the nodule in two and observe the color.
b. Use the pipette to place a drop of water on the slide.
ASM Microbial Discovery Activity
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c. Use the short end of a microscope slide to grind up the nodule in the drop of water.
d. Place a coverslip over the drop of water/root nodule material.
e. Observe under 10X and 40X magnification. Place a drop of immersion oil on the coverslip
and observe under 100X magnification for the presence of rod-shaped bacteria.
4. Divide students into groups and ask them to proceed through the stations, following the
directions in the notes. Ideally 2-4 participants work well together, and each group should
have its own supplies.
Delivery
Suggestions for the introduction of this activity in class:
The rumen of a cow is an excellent example for the introduction of symbiotic relationships to the class.
Query the students about the diet of cows. What types of food do cows consume? Ask them to take a
moment and consider their previous observations of cows. Where were the cows and what were they
doing? Most of us have observed cows in the fields on a farm and they are generally eating grass. How
is it that a cow survives by eating only hay and grass, and we cannot?
Cows are ruminants. Ruminants are mammals containing a specialized and relatively complex
stomach/digestive system, which includes an organ called the rumen. A cow would not be able to use
plants as a food source if it were not for the presence of microorganisms in the rumen. Ruminants
have developed a symbiotic relationship with these microorganisms and it is this relationship that
allows them to utilize food sources such as grass, hay, and straw, which are composed largely of
cellulose. (Grass is composed of about 50% cellulose and 50% xylan and fructosan.) No vertebrate
produces the enzyme cellulase, which is required for the digestion of cellulose, but the rumen microbes
do!
The size of a typical adult cow rumen is 120-150 liters (25-40 gallons), while a sheep rumen is about 6
liters in size. The rumen operates like an anaerobic (oxygen free) bacterial chemostat (an apparatus
that supplies a constant supply of food and removal of waste) that breaks down complex plant
materials to molecules that can be used by the animal. The contents of the rumen turn over every 1012 hours. Query the students as to what the cow could receive from the relationship and also on what
the microorganisms receive from the relationship.
The cow provides:
- constant temperature of 37-39 °C
- well buffered (bicarbonate, phosphate pH 6.5) mineral solution (100-200 liters saliva/day)
- periodic supply of nutrients and output of rumen fluid (chemostat like growth)
- mixing of rumen contents
- urea as source of ammonium (nitrogen)
- strict anaerobic environment
- chewing of cud (i.e. plant materials) to aid bacterial attack of polymers
The microorganisms provide:
- Exoenzymes, such as cellulase, that digest plant polymers
- Fermentation of simple sugars to provide fatty acids,
- Fatty acids (e.g. acetate, propionate, butyrate) and amino acids are absorbed by the cow via the
rumen and intestines.
- Microorganisms that migrate into the digestive tract are lysed by the enzyme lysozyme
(produced in epithelium of the rumen), and are further broken down and absorbed. They
provide a source of amino acids and vitamins for the cow.
Three groups of microorganisms are required for degradation of complex carbohydrates:
1. The cellulolytic and hydrolytic fermentative protozoa and bacteria – break down complex
carbohydrates, such as cellulose, into simple sugars
2. The acetogenic bacteria - ferment the sugars to acetate
3. The methogenic bacteria – reduce acetate to methane
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Microorganisms
The microorganisms used in this activity may all be obtained from the environment or ordered
from biological supply houses.
• Termite microflora
• Lichen
• Rhizobia bacteria in a legume root nodule
•
Assessment and Evaluation of Activity
The following are possible assessment techniques. Use a rubric to assess any of these suggestions (See
Appendix for several sites on rubrics).
Have the students:
• compose an independent formal lab write up.
• write quiz questions pertaining to the lab and quiz other members of their class. (The teacher
may also use the student-generated questions to create a test or quiz.)
• keep a journal.
• make a presentation (verbal or computer–based).
• create a concept map that describes the lab and its results.
• participate in group discussion.
Supplementary Materials
Links to obtain addition information:
Termites
•
Lots of information about termites!
University of Toronto
http://www.utoronto.ca/forest/termite/termite.htm Including: Termites 101:
http://www.utoronto.ca/forest/termite/trmts101.htm
•
More information about termite guts!
BioMEDIA Associates
http://www.ebiomedia.com/gall/guts/guts1.html
•
Termite Movies:
University of California Berkeley
http://nature.berkeley.edu/lewis/index2.html
Spartanburg Community College
Video of termite protozoa
http://www.stcsc.edu/ecology/TermSymb.htm
Lichens
•
Welcome to lichen-land!
http://ocid.nacse.org/lichenland/
•
Lichens of North America
http://www.lichen.com/
ASM Microbial Discovery Activity
Symbiotic Relationships – Page 9
•
The British Lichen Society
http://www.thebls.org.uk/
Legume Root Nodules
•
The Nitrogen Cycle:
Windows to the Universe (Beginning, Intermediate & Advanced Levels)
http://www.windows.ucar.edu/tour/link=/earth/Life/nitrogen_cycle.html&edu=high
Rhizobium:
University of Hawaii at Manoa
http://www.botany.hawaii.edu/faculty/webb/BOT410/Roots/RootSymbioses.htm
•
John Innes Centre, Norwich Research Center
http://www.jic.bbsrc.ac.uk/SCIENCE/molmicro/Rhizo.html
USDA Agricultural Research Center: Rhizobium to the rescue—Cleaning up the
environment with bacteria:
http://www.ars.usda.gov/is/kids/environment/story3/rhizobium.htm
Endosymbiosis
•
University of California Berkley – Understanding Evolution
Endosymbiosis: Lynn Margulis
http://evolution.berkeley.edu/evolibrary/article/0_0_0/history_24
Evidence for Endosymbiosis
http://evolution.berkeley.edu/evolibrary/article/0_0_0/endosymbiosis_04
•
Discovery Biology - Endosymbiosis Animation: The Evolution of Organelles
http://www.sumanasinc.com/webcontent/anisamples/nonmajorsbiology/organelles.html
Appendix
Links for Information on Developing Rubrics for Learning Assessment
•
Discovery Education
http://school.discovery.com/schrockguide/assess.html
•
Kennesaw State University
http://edtech.kennesaw.edu/intech/rubrics.htm
•
Science Learning Network
http://www.sln.org/guide/dukerich/assessment.html
ASM Microbial Discovery Activity
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Answer Key for Questions
1. What organisms are involved in each relationship and describe the benefits each might obtain
from this relationship?
Answers may vary. Students may choose to apply those relationships discusses in the
exercise. These would include the following:
-
Termites: Termites carry microbes including various bacteria and the protozoan
trychonympha, in their gut. The termite provides a home and food while the
microorganisms assist with breaking down cellulose.
Lichens: Lichen is a symbiotic relationship between a fungus and photosynthetic
alga. The fungus provides a home and the alga provides food to the fungus.
Legumes: Legume plants carry root nodules that house the symbiotic bacterium,
Rhizobium. The rhizobium will convert nitrogen to ammonia which can be used
by the plant. The plants provides a home and food for the rhizobium.
2. You observed many different types of microbes at each station. What types of nutrients might
these microorganism be consuming for energy?
Answers will vary. Some may include: cellulose (used by microbes in the termite gut)
and glucose or sugars (produced by the alga during photosynthesis and the legume).
3. Many of the organisms observed today are able to move or are motile. How do organisms that
are single celled move?
Many of the microbes carry a flagellum.
4. How could these relationships (those observed today) have been established? How might these
relationships change over time?
Answers will vary. These relationships may have begun initially as simple
association and over time both organisms evolved to become dependent upon one
another.
5. Do you think that microorganisms might reside in or on you? What type of role might they play
in your health?
Answers may vary. Microorganisms do reside on humans including for example, skin
and gut. The human intestine carries numerous bacteria including E. coli. The human
gut provides a home and food for the bacteria while the bacteria can assist with
digestion of food and produce vitamins that could be used by the human. The human
normal flora may keep you healthy by competing with pathogens and blocking
pathogen attachment and growth.
ASM Microbial Discovery Activity
Symbiotic Relationships – Page 11
Student Handout
A Close-up Examination of Microorganisms
Involved in Mutually Beneficial Symbiotic
Relationships
Introduction
You will work in groups. Each group will visit three observation stations equipped with microscopes
and various supplies, where you will observe three symbiotic relationships involving microbes. You
will prepare wet mounts of termite gut microflora, lichen, and Rhizobium root nodules for microscopic
examination.
Student Background Knowledge
You should read and be familiar with the following information about symbiosis, as well as the
information that can be found at the Internet websites listed as references and the vocabulary terms
listed below. Also, read over the laboratory procedure.
Symbiotic Relationships
Symbiosis describes the relationship between two or more organisms. This relationship can occur
between two different types of microbes, between a microbe and a plant, or between a microbe and an
animal. When both partners depend on each other and benefit from one another the relationship is
termed mutualism. Should one partner gain an advantage, and the other partner is neither harmed
nor benefits from the presence of the partner, commensalism is used to describe the relationship. In
parasitism, only one of the symbiotic partners benefits; the other partner suffers. Sometimes the
symbiotic partners are physically associated with one another. If one of the partners lives inside the
cell of the other partner, the relationship is called endosymbiosis. In ectosymbiosis, the partners
may not be physically associated or one partner may exist in or on the body of the other partner, but
not within its cells.
Endosymbiosis examples:
- microbe/microbe: Paramecium bursaria/photosynthetic green algae
- plant/microbe: legume(clover)/nitrogen-fixing Rhizobium bacteria
Ectosymbiosis examples:
- animal/microbe: ruminants (cows, sheep, buffaloes, etc.) /cellulase producing
anaerobic microbial community (protozoa and bacteria)
- microbe/microbe: lichen (fungus/ photosynthetic bacterium or alga)
- microbe/microbe: sulfur cycling by bacteria
- insect/microbe: leaf-cutting ants/fungi
The three examples of symbiosis explored in this exercise are observed using preparations
made from termite hindguts, lichens, and legume root nodules.
Termites: Termite guts are the world's smallest bioreactors. The hind-gut of wood-feeding
termites harbor mainly aerotolerant lactic acid bacteria, facultatively anaerobic enterobacteria,
and even a significant fraction of strictly aerobic bacteria, in addition to the larger anaerobic
protozoa, such as Trichonympha spp. These indwelling microbes degrade the wood cellulose
into sugar, and the sugar is then consumed by the termite. Interestingly, some of the protozoa
ASM Microbial Discovery Activity
Symbiotic Relationships – Page 12
living in the termite have bacterial endosymbionts of their own! For example, some of these
protozoa have no mitochondria; they rely on their bacterial endosymbionts to generate the
energy (ATP) they need.
Lichens: Lichens are a successful alliance between a fungus and a photosynthetic alga or
bacterium. The fungus partner provides a home for the alga or bacterium; the alga or
bacterium produces food (organic compounds) for the fungus through photosynthesis. Only
certain algae and certain fungi, or certain bacteria and certain fungi, can get together to form a
lichen. This means that each union creates a unique type of lichen body (thallus). Scientists
identify and assign names to lichen based on their observations of the thallus. Several features
are used to characterize lichens, including body type, life form, lobules, and surface texture.
(Lobules are small outgrowths, usually from the edge or margin of the lichen. They are
important for reproduction.) Lichens occur in one of four basic growth forms:
1.
2.
3.
4.
crustose - crustlike, growing tight against the substrate
squamulose - tightly clustered and slightly flattened pebble-like units
foliose - leaflike, with flat sheets of tissue not tightly bound
fruticose - free-standing branching tubes
The true identity of lichens as symbiotic associations of two different organisms was first
proposed by Beatrix Potter, who is best remembered for her children's books about Peter
Rabbit. In addition to her books, she spent time studying and drawing lichens. Her
illustrations are still appreciated for their detailed and accurate portrayal of the delicate
beauty of these bizarre organisms.
Legumes: A well known symbiotic relationship is that between legumes, such as peas, beans,
soybeans, clover, alfalfa, and peanuts, and soil bacteria in the genus Rhizobium. The rhizobia,
which exist in nodules on the roots of the legume, can fix nitrogen gas when the symbiosis is
achieved. They can't fix nitrogen in the free-living state, however. (Legume/Rhizobium
nodules are red. This is due to the production of leghemoglobin which sequesters oxygen. This
helps to create an anoxic [oxygen-free] environment for the strictly anaerobic Rhizobium
bacteria.)
Fixed nitrogen, in the form of ammonia, is used as a nitrogen source by the legume and the
Rhizobium gets a nice place to live with all of the amenities, including photosynthate, water
and minerals! This is an extremely important relationship because nitrogen is usually the most
limiting element in terrestrial ecosystems. Furthermore, many legumes, like soybean, form the
basis for agriculture on a world-scale. Some legume seeds, like soybeans, contain high levels of
protein. These are the most important agricultural sources of protein in the world.
Internet Resources:
Termites
•
Lots of information about termites!
University of Toronto
http://www.utoronto.ca/forest/termite/termite.htm Including: Termites 101:
http://www.utoronto.ca/forest/termite/trmts101.htm
•
More information about termite guts!
BioMEDIA Associates
http://www.ebiomedia.com/gall/guts/guts1.html
Termite Movies:
University of California Berkeley
http://nature.berkeley.edu/lewis/index2.html
•
Spartanburg Community College
ASM Microbial Discovery Activity
Symbiotic Relationships – Page 13
Video of termite protozoa
http://www.stcsc.edu/ecology/TermSymb.htm
Lichens
•
Welcome to lichen-land!
http://ocid.nacse.org/lichenland/
•
Lichens of North America
http://www.lichen.com/
•
The British Lichen Society
http://www.thebls.org.uk/
Legume Root Nodules
•
The Nitrogen Cycle:
Windows to the Universe (Beginning, Intermediate & Advanced Levels)
http://www.windows.ucar.edu/tour/link=/earth/Life/nitrogen_cycle.html&edu=high
•
Rhizobium:
University of Hawaii at Manoa
http://www.botany.hawaii.edu/faculty/webb/BOT410/Roots/RootSymbioses.htm
John Innes Centre, Norwich Research Center
http://www.jic.bbsrc.ac.uk/SCIENCE/molmicro/Rhizo.html
USDA Agricultural Research Center: Rhizobium to the rescue—Cleaning up the
environment with bacteria:
http://www.ars.usda.gov/is/kids/environment/story3/rhizobium.htm
Endosymbiosis
•
University of California Berkley – Understanding Evolution
Endosymbiosis: Lynn Margulis
http://evolution.berkeley.edu/evolibrary/article/0_0_0/history_24
Evidence for Endosymbiosis
http://evolution.berkeley.edu/evolibrary/article/0_0_0/endosymbiosis_04
• Discovery Biology - Endosymbiosis Animation: The Evolution of Organelles
http://www.sumanasinc.com/webcontent/anisamples/nonmajorsbiology/organelles.html
Vocabulary
Symbiosis- a relationship between two or more organisms.
Endosymbiont- the member of the symbiotic relationship that resides inside the cell of the
other member of the relationship.
Ectosymbiont- The member of a symbiotic relationship that may not be physically associated
with the other partner, or may reside in or on the other partner, but not within its cells.
Mutualism- a symbiotic relationship where both organisms involved benefit from the
relationship.
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Commensalism- a symbiotic relationship where one organism benefits from the relationship
while the other organism is neither harmed nor benefits from the relationship.
Parasitism- a symbiotic relationship where one member benefits but the other member is
harmed by the relationship.
Strict Aerobe- an organism that requires oxygen to live.
Facultative Anaerobe – an organism that prefers to live in the presence of oxygen, but can
survive without it.
Aerotolerant Anaerobe – an organism that prefers to live in the absence of oxygen, but can
tolerate low levels.
Strict anaerobe – an organism that cannot survive in the presence of oxygen.
Biological Nitrogen-Fixation – The process in which a microorganism uses nitrogen gas
(N2) for a nitrogen source and “fixes” it to ammonia (NH3).
Materials
The necessary materials are already located at each activity station.
Station 1: Termite Hindgut
Termites
Microscope slides
Dissecting needles or probes
Forceps
Plastic pipette
Coverslips
Bottle of immersion oil with dropper (for observation at 100x magnification)
Beaker of 1:10 bleach solution for disposal of slides and dead termites
Microscope
Lens tissues
Station 2: Lichen
Fresh lichen samples
Microscope slides
Plastic pipette
Forceps
Coverslips
Dropper bottle of water
Microscope
Lens tissues
Beaker for disposal of slides
Station 3: Root Nodules
Legume (e.g. clover) root nodules
Scalpel or knife
Microscope slides
Coverslips
Dropper bottle of water
Microscope
Bottle of immersion oil with dropper (for observation at 100x magnification)
Lens tissues
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Beaker for disposal of slides
Procedure
Pay attention as the instructor demonstrates how to perform the slide preparation for each activity.
Following the demonstrations, each group will prepare the samples and observe them under the
microscope.
1. Technique needed to remove the hindgut of the termite.
a. Obtain a microscope slide, coverslip, compound microscope and two needles or sharp probes
from the supply area.
b. Termite dissection
Method I:
1. Use the forceps to obtain a termite and hold it on the microscope slide.
2. Using the two needles, place one on the front end of the termite and one on the
abdomen. When these needles are pulled in opposite directions, the alimentary canal
will open and reveal the milky-colored gut fluid, which contains a wealth of microbes
within.
3. Use the pipette to place a drop of distilled water on the slide near the alimentary
canal.. This will allow the protozoa to swim from the alimentary canal into the water
and allow the students much easier viewing.
4. Quickly cover the drop of water/gut fluid with a coverslip to preserve an anoxic
environment for the anaerobic microbes living in the gut.
Method II:
1. Use the pipette to place a drop of water on the slide.
2. Use the forceps to grasp the front end of the termite and use a second pair of forceps to
gently milk the hindgut, allowing hindgut contents to drop on to the slide into the drop
of water.
3. Quickly cover the drop of water/gut fluid with a coverslip to preserve an anoxic
environment for the anaerobic microbes living in the gut.
c. Observe the preparation under 10X and 40X magnification. If the compound microscopes
are equipped to operate using the darkfield technique, Trichonympha spp. can be seen with
great ease.
d. To observe the motion of the protozoal flagella and bacterial endosymbionts, place a drop of
immersion oil on the coverslip and observe under 100X magnification.
2. The squash technique for observing lichen
a. Use the pipette to place a drop of water on the slide.
b. Use the forceps to pick up a small piece of lichen and place it in the drop of water.
c. Use the short end of a microscope slide to grind up the material in the water.
d. Place a coverslip over the drop of water/lichen material.
e. Observe under 10X and 40X magnification. Look for oval or round green algal cells and
filamentous fungal cells.
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3. The squash technique for observing the legume root nodules
a. Use a scalpel or knife to cut the nodule in two and observe the color.
b. Use the pipette to place a drop of water on the slide.
c. Use the short end of a microscope slide to grind up the nodule in the drop of water.
d. Place a coverslip over the drop of water/root nodule material.
e. Observe under 10X and 40X magnification. Place a drop of immersion oil on the cover slip
and observe under 100x magnification for the presence of rod-shaped bacteria.
Safety Precautions
Today you will be working with microorganisms that occur in the environment. It is always important
to observe caution when using environmental samples. Exercise care when preparing squash
preparations and when using sharp instruments (probes, needles, scalpels, knives, etc.).
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SYMBIOSIS OBSERVATION STATIONS WORKSHEET
Name: __________________________________
I. For each station, please draw your observations below and describe what you see
(provide the color of the organism (if any) and the shape and describe if the organisms are
motile).
Station 1: Termites
Station 2: Lichens
Station 3: Root Nodules
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Please answer the following questions:
1. What organisms are involved in each relationship and describe the benefits each might obtain
from this relationship?
2. You observed many different types of microbes at each station. What types of nutrients might
these microorganism be consuming for energy?
3. Many of the organisms observed today are able to move or are motile. How do organisms that
are single celled move?
4. How could these relationships (those observed today) have been established? How might these
relationships change over time?
5. Do you think that microorganisms might reside in or on you? What type of role might they play
in your health?
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