Intended Audience. Keywords Lichen, Termites, Fungus, Algae, Nitrogen Fixation, Legumes, Root Nodules, Symbiotic Relationships, Rhizobium 5-8 X 9-12 X

Transcription

1 Microbial Discovery Activity Symbiosis Stations Abstract By visiting stations equipped with microscopes and various supplies, students will observe three symbiotic relationships involving microbes. Three stations equipped with microscopes, slides, and live material are arranged by the teacher. Students visit the stations and prepare wet mounts to observe termite gut microflora, lichen, and Rhizobium root nodules. Intended Audience 5-8 X 9-12 X Learning Objectives By the completion of this activity, the students will be able to: Prepare a wet mount slide of termite hindgut and observe endosymbionts. Prepare a squash preparation of lichen and identify the fungal and algal mutualists. Prepare and observe a squash preparation of a root nodule. Describe the components and types of symbiotic relationships. Explain the importance of symbiosis. Necessary Student Background Students will need a general understanding of the symbiosis and the symbiotic relationship of mutualism. Suggestions on how to lead a discussion on symbiotic relationships may be found in the background and supplementary information in the Teacher Handout. Keywords Lichen, Termites, Fungus, Algae, Nitrogen Fixation, Legumes, Root Nodules, Symbiotic Relationships, Rhizobium American Society for Microbiology Education Department 1752 N Street, NW Washington, DC Education Resources@asmusa.org

2 General Informatiion Author Robin Patterson Butler County Community College Butler, PA Contributor Michelle S. Thomas Campbell University Department of Biological Sciences Buies Creek, NC National Science Education Standards Addressed All Levels of Learning: 1. Unifying concepts: Systems, order and organization Evidence, models and explanations Change, consistency and measurement Form and function Science Content Standards 5-8: 1. Life Science Regulation and behavior Populations and ecosystems Diversity and adaptations of organisms 2. Science in Personal and Social Perspectives Populations, resources and environments Science Content Standards 9-12: 1. Life Science: The interdependence of organisms Biological evolution Behavior of organisms Activity Specifications Classroom setting Requires special equipment Uses hands-on manipulatives Requires mathematical skills Can be performed individually Requires group work Requires more than one (45 min) class period Appropriate for special needs student X X X X X Symbiosis Stations Page 1

3 Teacher Handout Symbiosis Stations Class Time Approximately one hour of class time is required. 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 the stations. Background The students should be familiar with the different types of symbiotic relationships and the definition of symbiosis. 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 cell. 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 does it benefit 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 of the other partner s cell the relationship is called endosymbiosis. In ectosymbiosis, both partners exist outside one another. Endosymbiosis examples: - microbe/microbe: paramecium cell/algae - plant/microbe: clover/rhizobium Ectosymbiosis examples: - animal/microbe: cow rumen/anaerobic microbial community (protozoa) - microbe/microbe: lichen (fungi/ photosynthetic bacteria) - microbe/microbe: sulfur cycling by bacteria - insect/microbe: leaf-cutting ants/fungi The three examples of symbiosis explored in this exercise include: termite hindguts, lichens, and legumes. Termites: Termite guts are the world's smallest bioreactors. 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 protozoa (Trichonympha) that live within the termite. These indwelling protozoa degrade the wood cellulose into sugar, and the sugar in turn is consumed by the termite. Lichen: Lichens are a successful alliance between a fungus and an alga. Only certain algae and certain fungi can get together to form a lichen. Since only certain algae and fungi can get together, this means that each union creates a unique type of lichen body (thallus). This thallus enables scientists to make identifications and assign names. Several features are used to identify lichens, including body type, life form, lobules, and surface texture. Lobules are small outgrowths, usually Symbiosis Stations Page 2

4 from the edge or margin of the lichen. They are important for reproduction. Lichens occur in one of four basic growth forms: 1. crustose - crustlike, growing tight against the substrate 2. squamulose - tightly clustered and slightly flattened pebble-like units 3. foliose - leaflike, with flat sheets of tissue not tightly bound 4. 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 and soil bacteria in the genus Rhizobium. The rhizobia can fix nitrogen gas when the symbiosis is achieved. They can't fix nitrogen in the free-living state, however. Fixed nitrogen is obtained by the legume and the Rhizobium gets a nice place to live with all of the amenities! This includes 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. 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. Materials and Equipment Station 1: Termite Hindgut Termites they can be ordered or captured (see teacher notes) Slides Probes Forceps Coverslips Beaker of 1:10 bleach solution for disposal of slides and dead termites Microscope Lens tissues Station 2: Lichen Fresh lichen samples Slides Coverslips Dropper bottle of water Microscope Lens tissues Beaker for disposal of slides Station 3: Root Nodules Legume or clover root nodules Slides Coverslips Dropper bottle of water Microscope Lens tissues Beaker for disposal of slides Symbiosis Stations Page 3

5 Methods Instructor Preparation for the Activity: 1. Organize the stations so that they will be accessible by the groups of students. Each station will consist of the materials listed above. 2. Depending on the amount of time available, students may visit the stations in teams or individually. 3. 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). To capture 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). 4. 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. 5. Clover or legumes can be pulled from the ground or can be ordered from a biological supply house. 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. Obtain a termite and hold it on the microscope slide. c. 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 wealth of protozoans within. d. A drop of distilled water should be added to the slide and a coverslip place in a normal wet mount fashion. This will allow the protozoan to swim from the alimentary canal and allow the students much easier viewing. e. If the compound microscopes are equipped to operate using the darkfield technique, the trichonympha can be seen with great ease. f. Alternately, termites may be grasped with forceps and a second pair of forceps used to gently milk the hindgut, allowing hindgut contents to drop on to the slide into a drop of water. 2. Demonstrate the squash technique for observing lichen: a. Scrape some lichen from a tree or rock into a small container. b. Pick up a small piece and macerate it into a drop of water, using the short end of a microscope slide. c. Apply a coverslip and observe. You will see oval or round green algal cells and filamentous fungi. Symbiosis Stations Page 4

6 3. Demonstrate the squash technique for observing the root nodules: a. Cut the nodule in two and observe the color. b. Macerate the nodule in a drop of water on a slide. c. Apply a coverslip and observe for 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 their own supplies. Microorganisms Microorganisms are present in the natural living specimens that are being examined. Safety Precautions Exercise care when preparing squash preparations and when using probes. Assessment Below are possible assessment techniques. Students can compose an independent formal lab write up. Have students 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. Journal. Presentation (verbal or computer based). Create a concept map that describes the lab and its results. Group discussion. Use a rubric to assess any of the suggestions above (See Appendix at the end of the Teacher Handout for several sites on rubrics). Symbiosis Stations Page 5

7 Supplementary Information Suggestions for the introduction of this activity in class: To introduce symbiotic relationships to the class, the rumen of a cow is an excellent example of symbiotic relationships. Query the students about the diet of cows. What types of food do cows consume? Take a moment and consider your previous observations of cows. Where were they 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 eats only hay and grass, yet provides us with meat and milk? Cows are ruminants. Ruminants are mammals containing a specialized and relatively complex stomach/digestive system, called a rumen (a stomach-like organ) that allows the animal to live on plant materials. 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 microorganisms and it is this relationship that allows them to utilize grass, hay, and straw as a food source. Grass is composed of about 50 % cellulose and 50 % xylan and fructosan. The site of the animal/microbe symbiosis is the cow s rumen. The size of a typical adult cow rumen is liters (25-40 gallons), while a sheep rumen is about 6 liters. 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 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 C - well buffered (bicarbonate, phosphate ph 6.5) mineral solution ( l saliva/day) - periodic supply of nutrients and output of rumen fluid (chemostat like growth) - mixing of rumen contents - urea as source of ammonium - strict anaerobic environment - chewing of cud (i.e. plant materials) to aid bacterial attack of polymers - The microorganisms provide: - Exoenzymes 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 bacteria and fungi. 2. The acetogenic bacteria. 3. The methane producing bacteria In this activity, the students will be observing symbiosis in three systems termites, lichen, and legumes. Ask the students to think about and discuss the nutrients these organisms use to survive and grow. How might a symbiont assist them in doing this? Symbiosis Stations Page 6

8 Appendix Links to obtain addition information: Welcome to lichen-land! Lichens of North America: The British Lichen Society: Lots of information about termites! Including: Termites 101: Termite Movies: More information about termite guts! The Nitrogen Cycle: Video of termite protozoa: Rhizobium: - m Rhizobium to the rescue Cleaning up the environment with bacteria: Symbiosis Stations Page 7

9 Student Handout Symbiosis Stations Student Name: Introduction Today you will be exploring three different examples of symbiotic relationships that are established between living organisms. 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 cell. 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 does it benefit 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 of the other partner s cell the relationship is called endosymbiosis. In ectosymbiosis, both partners exist outside one another. Endosymbiosis examples: - microbe/microbe: paramecium cell/algae - plant/microbe: clover/rhizobium Ectosymbiosis examples: - animal/microbe: cow rumen/anaerobic microbial community (protozoa) - microbe/microbe: lichen (fungi/ photosynthetic bacteria) - microbe/microbe: sulfur cycling by bacteria - insect/microbe: leaf-cutting ants/fungi The three examples of symbiosis explored in this exercise includes: termite hindguts, lichens, and legumes. Termites: Termite guts are the world's smallest bioreactors. Wood-feeding termites harbor different types of microorganisms including bacteria and protozoans. Interestingly, these microbes can be quite different from one another. For example, one resident includes the lactic acid bacteria. The lactic acid bacteria do not need oxygen to survive. They can, however, grow in an environment where oxygen is present but do not use oxygen. A different type of symbiont inside the termite gut are the enterobacteria. Enterobacteria prefer to use oxygen but they can also grow in absence of oxygen. In other words, they do not have to have oxygen to grow but do prefer to have it. Lastly, there are microorganisms present that are called strict aerobes. These aerobes, such as the aerobic bacteria and the larger protozoa such as Trichonympha require oxygen for growth. Aerobic organisms cannot survive in the absence of oxygen. All of these microorganisms can be found inside the termite gut. These indwelling protozoa and bacteria degrade the wood cellulose into sugar, and the sugar in turn is consumed by the termite. Lichen: Lichens are a successful alliance between a fungus and an alga. Only certain algae and certain fungi can get together to form a lichen. Since only certain algae and fungi can get together, this means that each union creates a unique type of lichen body (thallus). This thallus enables scientists to make identifications and assign names. Several features are used to identify lichens, including body type, life form, lobules, and surface texture. Lobules are small outgrowths, usually Symbiosis Stations Page 8

10 from the edge or margin of the lichen. They are important for reproduction. Lichens occur in one of four basic growth forms: 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 and soil bacteria in the genus Rhizobium. The rhizobia can fix nitrogen gas when the symbiosis is achieved. They can't fix nitrogen in the free-living state, however. Fixed nitrogen is obtained by the legume and the Rhizobium gets a nice place to live with all of the amenities! This includes 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 soybeans, 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. 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. Terms Symbiosis- a relationship between two or more organisms. Mutualism- a symbiotic relationship where both organisms involved benefit from the relationship. 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. Aerobe- an organism that requires oxygen to live. Endosymbiont- the member of the symbiotic relationship that resides inside the body of the other member of the relationship. Ectosymbiont- Both partners of the symbiotic relationship reside on the outside of each other. Symbiosis Stations Page 9

11 Activity Safety Considerations 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 probes. Materials Station 1: Termite Hindgut Termites Slides Probes Forceps Coverslips Beaker of 1:10 bleach solution for disposal of slides and dead termites Microscope Lens tissues Station 2: Lichen Fresh lichen samples Slides Coverslips Dropper bottle of water Microscope Lens tissues Beaker for disposal of slides Station 3: Root Nodules Legume or clover root nodules Slides Coverslips Dropper bottle of water Microscope Lens tissues Beaker for disposal of slides Procedure Today you will examine three different specimens that are set up at three different stations. Please listen to the instructions provided by your instructor. The directions for preparing the specimens are described below and will be explained by your teacher. 1. Termite Station- Technique 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. Obtain a termite and hold it on the microscope slide. Symbiosis Stations Page 10

12 c. 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 wealth of protozoans within. d. A drop of distilled water should be added to the slide and a coverslip placed in a normal wet mount fashion. This will allow the protozoan to swim from the alimentary canal and allow the students much easier viewing. e. If the compound microscopes are equipped to operate using the darkfield technique, the Trichonympha can be seen with great ease. f. Alternately, termites may be grasped with forceps and a second pair of forceps used to gently milk the hindgut, allowing hindgut contents to drop on to the slide into a drop of water. 2. Lichen Station- The squash technique for observing lichen: a. Scrape some lichen from a tree or rock into a small container. b. Pick up a small piece and macerate it into a drop of water, using the short end of a microscope slide. c. Apply a coverslip and observe. You will see oval or round green algal cells and filamentous fungi. 3. Root Nodule Station: The squash technique for observing the root nodules: a. Cut the nodule in two and observe the color. b. Macerate the nodule in a drop of water on a slide. c. Apply a coverslip and observe for rod-shaped bacteria. Symbiosis Stations Page 11

13 Student Results 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, describe if the organisms are motile). Station 1: Termites Station 2: Lichens Station 3: Root Nodules Symbiosis Stations Page 12

14 II. 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? Symbiosis Stations Page 13