Scientists have long puzzled about the origins of viruses and how they help explain evolution. Today students look at the current structure of two viral groups to better understand how scientists use the many specific examples to explain the big idea of evolution. Here is an overview of what students will learn today and why I use the methods I do to teach this content.
Start the class by showing a clip from Paul Anderson's podcast on Viruses.
(I start it at 0:14 and play it until 0:50.)
Next, ask student the following questions and have them respond in their lab notebook:
(Possible answer: They both cannot reproduce without a host)
(Possible answer: A computer virus has no form. It is virtual. A real virus has a shape and form.)
(Possible answers: They are different sizes. They have different shapes.)
Explain that today the class will be exploring the differences among two of the major types of viruses by constructing several scale models. (Note: Now that students understand viral size and methods of transmission, it is possible to study how viruses have evolved into the form they have today. The lesson for today and tomorrow has students building models of several examples of the major groups of viruses. Understanding viral structure is the first step to understanding how viruses adapt and take over their host cell.)
Give the students the viral structure handout from the Protein Data Bank. Students should understand that there are currently four classification of viruses: polyhedral, helical, special, and enveloped. Remind students that polyhedral viruses have the typical viral arrangement of capsid made of capsomeres. A nucleic acid is contained inside the capsid.
Show students several electromicrographs of polyhedral viruses.
(Answer: Polyhedral viruses look like a sphere.)
(Answer: The shape of the capsid of polyhedral viruses is actually icosanhedral, a twenty-sided polygon).
Further explain to students that the icosanhedron is made up of equilateral triangles. The triangle are made using three different protein subunits. Then the polygon is made of those equilateral triangles that are fused together. (Note: I show it as a puzzle and then show how the pieces fit together. This has been shown mathematically to be the most optimal way of forming a closed shell.)
(Possible answers include: the poliovirus, rhinovirus, adenovirus, and dengue virus, but there are many more.)
Have students create a concept map with the term virus in the center and the types of virus (polyhedral, helical, special, and enveloped) radiating off of the center. Have students add the structure of the virus and theories of its evolution. (Note: Almost every living thing on the Earth has a type of polyhedral virus that will infect it so these viruses are most definitely polyphyletic.)
Obtain a copy of the Dengue virus from Protein Data Bank. Students should cut the virus capsid out along the solid black lines. Fold the isocosahedron along the dotted lines and tuck the white flaps inside the isocosahedron. Tape all seams, but one closed. Make a model of the RNA genome by measuring 5.4 m of thin yarn. (The yarn can be any color. It is important, however, that you pick one color to represent RNA and one color to represent DNA. Also, make sure you have enough of particular color for all the models you plan to make.) Wrap the yarn in a figure 8 shape and tuck it inside the capsid. Tape the final seam shut.
State to the student that the paper model that they hold in their hand is 1,500,000 times larger than an actual Dengue virus. While students construct their model, I relate the following narrative to them.
(Note: I write these narratives myself and do not read them word for word. Since I typically have small classes, lecture is awkward. I am a very good storyteller so once I write these narratives, I memorize the important points and then just relay them to my students as a story. This makes the exchange more organic. I can move around the room to individual lab group and point out important structures and vital parts of the narrative. This laid back method of delivery allows my students to ask questions as they arise and it allows them to visualize the complexity of these viral structures.)
The Dengue virus causes Dengue hemorrhagic fever which affected an estimated 50 to 100 million people a year. The range of this virus includes Southeast Asia, the Western Pacific Islands, Latin America, and the Caribbean. Since it is primarily a tropical disease, we do not see it in the United States. However, worldwide it is a common disease infecting between 50 and 100 million people a year. Individuals contract the virus through a mosquito bite.
Ask the students what type of transmission this is? (vector-transmission)
Dengue virus antibodies are able to be detected in the blood stream using an diagnostic test known as the DENV Detect IgM Capture ELISA. Once a person has enough has enough antibodies in their bloodstream to detect, the person will also be running a fever. Treatment is important because Dengue is a hemorrhagic disease which can kill the infected person.
On the cellular level, the dengue virus moves through the endoplasmic reticulum and the trans-Golgi network. Before the virus matures, it is not capable of fusing with cell membranes. Therefore, cannot infect their own host cells and must mature. Once mature, however, the virus particle is able to fuse to cell membranes. This ensures it can new host cells. Mature viruses are able to change the protein structure of their capsid. Recently, this cellular model has been tested by scientists in the lab with scientists modeling the trans-Golgi network environment in test tubes. During this study, they were able to determine changes in the virus's structure as the environment became more acidic.
The model you are using today was constructed using electron micrograph images and computer modeling. It is was first published in Science in 2008. Then the National Protein Bank made a copy available for student to use to better understand models.
(Note: It is important that students understand that this model and other virus models are based on scientific studies where scientists have used numerous methods to develop this models. If not brought to their attention, most students will not realize how these models are developed.)
Refer students back to the viral structure handout from the Protein Data Bank. Remind students that helical viruses are made of a long cylindrical capsid.
(Answer: Their capsomeres that are all the same protein arrange in a long strand that wraps around itself like a spring.)
(Answer: The nucleic acid is wound on the inside of the capsid.)
(Possible answers: One of the first virus to be described, Tobacco Mosaic Virus, is a helical virus. Another example of a helical virus is Ebola.)
Have students continue to add to their concept map by adding the structure of the virus and theories of its evolution to the helical virus portion of the concept map. (Note: Almost every living thing on the Earth has a type of helical virus that will infect it so these viruses are most definitely polyphyletic.)
(Note: Because of the expense, we typically make one model of the helical virus. Metal dryer vent tube can be expensive if it is purchased for individual lab groups. I have my students complete the dimensional analysis for the construction of this model. I have shown their calculations below.)
To construct an Ebola virus, use a metal dryer vent tube, yarn, foil, and duct tape. Metal dryer tubes can be found at any heating and air supply company or home remodeling center. 6.9 m of yarn need to be used for each model.
(The length of RNA was determined by assuming a 1 meter long piece of dryer vent tubing. The RNA genome of an Ebola virus is 18,959 to 18,961 nucleotides in length. The length of a single nucleotide is .34 nm. Therefore, the total length of the RNA strand is 6,446.74 nm. A typical Ebola virus capsid is 940 nm. 6446.74 nm/940nm = 6.9 Therefore, multiply the length of your tubing by 6.9 to determine what length of yarn you will need in your model if it is not 1 meter). One roll of duct tape can be used for a class of 20. Each model will need a 5 cm X 5 cm of Al foil to close the open end of the dryer tube.
Wrap the dryer vent tubing into a shepherd crook’s shape and duct tape in place with silver duct tape. The RNA genome should be made from the same color of yarn as the Dengue virus. Measure out 6.9 m of yarn, then wrap it in around the hand in a circular fashion before removing it and placing it in the dryer vent. Seal the dryer vent with aluminum foil and secure with more duct tape.
While students are constructing their model, I relate the following passage to them. (Again, since I am a good storyteller, I do not read this to them. I talk with them about it. I've included some of the primary sources so that they can be compared with the narrative that I wrote. Many times, I have students want to know more. I can refer them to these primary documents.)
Unless you have been living under a rock, you have heard something about the Ebola virus within the last year because of the epidemic that has been raging through West Africa. Ebola is not something to take lightly as it causes severe hemorrhagic fever that has a death rate of 50% to 90% depending on the strain that the patient has contracted. Ebola is spread through direct contract with infected body fluids (saliva, semen, blood, sweat, feces, and vomitus). The incubation period is 2 to 21 days. Early symptoms are flu-like and include fever, chills, and muscle aches. Late symptoms that follow are indicative of multi-organ stress particularly the lungs, liver, and kidneys. First documented in 1976, Ebola has a high occurrence among wildlife in the countries where it is native. There are no vaccines and few treatments available. However, two recently developed vaccines are undergoing clinical trials will be available by the end of 2015. This makes Ebola a significant public health concern.
Take the dryer vent and twist it to form a shepherd’s crook. Use duct tape to hold the end in place. Measure 6.9 m of yarn to represent the single stranded RNA genome found in Ebola. Wrap the yarn around your hand to represent the helical arrangement of the RNA. While you are doing this, I am going to tell you about two studies that helped determine the structure of the virus you are currently constructing.
A team of researchers from Scripps Research Institute has recently determined the capsid structure of Ebola using diffraction techniques. While we will not make any of these surface structures on our model, it is important to understand that they are present. When in a human host, Ebola has a crystalline structure that has glycoproteins on its outside that allows it to binds to host cell and force the host cell to bring the virus inside the cell. The glycoproteins also drive the fusion of viral and host membranes and hide the virus from the host’s immune system. Ebola has very few sites for antibodies to bind so it is difficult for antibodies to neutralize the virus (Lee, et al, 2008).
Another team of researchers also determined that the capsid of Ebola assembled into helical structure that is represented by wire wrapping through the dryer vent. It is important to understand how scientists isolate and study viruses since they are so small and then test them using multiple methods to determine structure. The Ebola virus used in the study that our model is based was isolated from a survivor of a past outbreak. It was cultured in embryonic mammalian kidney cells for three days. Then the cells were broken apart with chemicals, incubated at 4 ºC for 30 minutes, and then put in a high speed centrifuge at 20,000g for 10 minutes to remove insoluble components. The remaining liquid was put into a Cesium chloride gradient and centrifuged at 250000 g at 20 °C for 1 hour. A visible band of virus was collected from the centrifuge tube. It was placed into another centrifuge tube, resuspended, and then subjected to centrifugation at 200000 g at 4 °C for 30 min. The liquid was poured off and the resultant pellet was then resuspended (Noda, et al, 2010).
Once the virus was isolated, they used a variety of techniques to determine the structure of the ebolavirus. The virus was treated with several enzymes (DNase, RNase, and trypsin) to see what was broken down and compared with non treated viruses using a seperation technique called Western blot. Next, the virus’ behavior was analyzed in various environmental salt concentrations and separated with CsCl gradient followed by staining the virus dyed with a protein stain called Coomassie blue. Results of the staining showed one protein component that had the molecular mass of the Ebola virus. Results of the other tests and UV absorption spectrum showed that the nucleic acid was RNA. UV absorption spectrum was used because certain pure biomolecules absorb certain wavelengths of light (Noda, 2010).
As you can see there are various specific test and techniques that must to done to determine the structure of the virus. Now place the wrapped RNA (yarn) the open end of the dryer vent. Seal it with a piece of aluminum foil and secure that end with more silver duct tape.
These references were used in the development of the narrative:
J.E. Lee, M.L. Fusco, W.B. Oswald, A.J. Hessell, D.R. Burton, and E.O. Saphire, "Structure of the Ebola virus glycoprotein bound to an antibody from a human survivor," Nature 454, 177 (2008).
Noda, T., et al. 2010. “Characterization of the Ebola virus nucleoprotein-RNA complex.” Journal of General Virology. 91(6):1478-1483. doi: 10.1099/vir.0.019794-0
(Note: Having a basic understanding some of the methods used by researchers to develop viral models will help students when they take other life science courses. Throughout the life science disciplines, researchers use similar methods.)
In their lab notebooks, students should write down one of the things that was most confusing today. Ask students
When students are done, they should turn in their notebooks for evaluation. Use this information to help prepare for tomorrow's lesson.