CASE TEACHING NOTES for "Dr. Collins and the Case of the Mysterious Infection"

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1 CASE TEACHING NOTES for "Dr. Collins and the Case of the Mysterious Infection" by Paula P. Lemons and Sarah Huber Biology Department Duke University INTRODUCTION / BACKGROUND This case study was designed for the seminar component of Introductory Biology at Duke University. In addition to seminar, students in this course also attend lecture and laboratory. Teaching assistant (TA) mentors lead the 12-student seminars during which students engage in directed inquiry exercises. Students work through the case study in the second week of the course. Topics covered in the lecture and lab components of the course prior to this case include organisms and cells as units of life, proteins and membranes, DNA structure, replication, and repair, and how to ask good questions in biology. Objectives After completing this case, students should be able to: 1. Describe some of the biochemical mechanisms by which antibiotics act against bacteria. 2. Evaluate which antibiotics would be effective against a given type of bacteria. 3. Apply their basic knowledge of DNA, genes, and proteins (and the relationship between them) to a new question. That is, how do changes in DNA observed by pulsed-field gel electorophoresis impact patterns of antibiotic resistance? 4. Synthesize the relationship between antibiotic consumption and antibiotic resistance based on what they have learned about bacteria, antibiotics, and natural selection. 5. More adeptly use data from charts and gels to answer questions. CLASSROOM MANAGEMENT Students are expected to have read the background information about bacteria, antibiotics, and the evolution of antibiotic resistance before coming to class. We assigned this reading in seminar the previous week. The primary aim of the reading is to introduce students to the topics of bacteria, antibiotics and antibiotic resistance. This case is designed as an "interrupted case," that is, students are given one piece of information at a time and asked to do something with that information. After they have completed one part, they receive the next piece of information. There are four parts to the exercise.

2 Given that we use this case study for our 12-student seminar sections, we make the following suggestions to TA mentors about how to teach the case although they are free to choose whichever method they think will work best with their particular section. 1. Small group work. Each group of three or four students receives the first part of the exercise and is given a specified amount of time to work through the questions in that part. At the end of the designated time, the TA mentor surveys most or all of the groups for their answers, then hands out the next part. Again, groups are given a specified amount of time to work through the problem and are surveyed by the TA mentor about their answers. The exercise is written so that the TA mentor needs to provide little information during the polling times; each new part provides "the answer" to the previous part. However, this changes after Part IV when students are asked three questions that challenge them to think about how antibiotics select for resistant strains and the molecular events that lead to the emergence of a new strain of bacteria. Part IV offers a lead-in for TA mentors to end seminar with a class-wide discussion about these more open-ended questions. NOTE: Our TA mentors have found that assigning specific jobs (e.g., group leader; time keeper, and recorder/reporter) to each student in a group helps to keep group work moving at a steady pace. 3. Class-wide discussion. In this format, the TA mentor functions as the group leader and moves students as a class through the exercise. The TA mentor hands out each part, gives students a chance to read the information, then begins discussing the problem. This format allows the TA mentor to exert more control over the classroom, but may prevent some of the quieter students from becoming as involved in the discussion. At the end of both the background reading and the case, a list of references is provided. Students are not required to read these references. Rather, these are provided for students and instructors who may want to do more reading about antibiotic resistance and, more specifically, cases like the one described in this case study. In terms of time requirements, in Introductory Biology, seminar lasts 50 minutes. Based on TA mentors' experiences, we estimate only five to seven minutes for both Parts I and II since they are rather straightforward. Part III is generally the most challenging part, and Part IV generates the most interesting discussion. Because of this, we recommend allowing at least 15 minutes for each of these parts. It might also be possible to expand this case study beyond a 50-minute class session so that there would be time for more discussion among students and between students and the instructor during each part of the exercise. Although the current constraints of Introductory Biology at Duke University prevent us from taking this approach, individuals who use this case study should feel free to make such adaptations. ANSWER KEY Answers to the questions posed in the case study are provided in a separate answer key to the case. Those answers are password-protected. To access the answers for this case, go to the key. You will be prompted for a username and password. For the username and password, contact the National Center for Case Study Teaching in Science administrator at answerkey@sciencecases.org.

3 ASSESSMENT OF LEARNING GOALS As explained in the background reading, we use this case in Introductory Biology as one of three installments in a series on antibiotic resistance. After working through this case, students deal with the topic again in lab by surveying for the prevalence of antibiotic resistant bacteria carried by them and their peers and in nearby water supplies (Lemons et al.). Finally, students work in groups during seminar and outside of seminar to collect their results from lab and analyze these results via a writing assignment. Due to this structure, we do not use the case study class period for assessment. Rather, we assess whether they have achieved the specified learning objectives in three independent ways. First, in the lab that follows this case, students design an experiment in which they select antibiotics that will be effective against members of the genus Staphylococcus and members of the family Enterobacteriaceae. Thus, TA mentors are able to assess how well students are achieving learning objective 2 as they observe the quality of each student's experimental setup. Second, students are assessed by their performance on a group writing assignment dealing with antibiotic resistance. As part of this assignment, students must synthesize what they have learned from lab about the relationship between antibiotic resistance and antibiotic consumption (learning objective 4) to address the concern that Dr. Collins raises in Week 2, Day 10 of the case (i.e., that increased use of antibiotics will lead to the emergence of new strains of resistant bacteria). Finally, on exams, students are responsible for concepts that have been covered not only in lecture but also in lab and seminar. Therefore, we frequently use the curriculum on antibiotic resistance as a source for exam questions. ADDITIONAL COMMENTS AND RECOMMENDATIONS This case study has been run in 26 sections of Introductory Biology. Here are some additional comments and recommendations based upon the experience of our TA mentors:! Many of our TA mentors report that students are highly engaged by and become emotionally invested in the case. For example, they express frustration that Dr. Collins did not prescribe the antibiotic that they recommended and feel that the outcome for Kayla Starnes would have been different if she had.! Some TA mentors caution that due to the large amount of reading involved in the case, managing individual students and groups of students who read at different paces can be challenging. Perhaps one solution is to predetermine groups such that differences in pace among individual students and groups of students will be minimized. Some TA mentors have also found it helpful to have a series of additional questions for each part prepared ahead of time for groups that work more quickly than others.! In response to TA mentors comments, Part III was rearranged to place the description of pulsefield gel electrophoresis into an appendix-like subsection. While some TA mentors felt that it was worthwhile for students to invest time in understanding this technique, others felt that it was not essential. We hope that including it as a subsection to Part III gives you the leeway to either use or ignore this portion of the exercise.

4 FOLLOW-UP WRITING ASSIGNMENT The students are given the following assignment in connection with this case study: For more than a week, you have focused on the problem of antibiotic resistance. You learned about bacteria - "good" ones and "bad" ones - and some of the details about how antibiotics affect them. From that you learned about some of the mechanisms bacteria use to evade antibiotics and understand that changes in the antibiotic resistance patterns of bacteria occur first on the molecular level. You carried out experiments that address the question of whether there is a positive correlation between antibiotic resistance and antibiotic consumption. Finally, you've begun to think about how bacterial transformation and the relative growth rates of different types of bacteria inform our understanding of the antibiotic resistance problem. As a final synopsis of all you have learned in this series of the course, address the following two sets of questions in writing as a group. 1. We have posted the results from our class-wide correlational study (see lab p. 6) to the course web site under "Course Documents." Look at the table showing the mean number of antibiotic courses per student in the following categories: penicillin resistant and penicillin susceptible. Also look at the graph plotting the number of antibiotic courses versus the percent of students with penicillin resistant bacteria. Do our results support or refute Dr. Collins' concern that increased use of antibiotics will lead to emergence of new strains of resistant bacteria (as a reminder about Dr. Collins' concern see Part IV of the case study "Dr. Collins and The Case of the Mysterious Infection")? What additional experimental results would provide more information to support or refute this concern? 2. In the bacterial transformation that you carried out in lab, did the E. coli carrying the ampr gene or those not carrying the ampr gene grow faster? What does this result suggest about what would happen to antibiotic resistance if we curbed our antibiotic consumption? How is the bacterial transformation that you performed in lab limited in answering this question? What type of experiment might provide better information to answer this question? Your paper should address these questions in a concise and coherent way. If you find it helpful to include additional references - journal articles, web pages, literature reviews - please do so, citing them appropriately. This assignment should be a collaborative effort by your group and should be 2 to 3 double-spaced typed pages in length. REFERENCES Lemons, P.P., Corliss, T., and Motten, A.F Bacteria and antibiotics. Written for the laboratory component of Introductory Biology at Duke University, unpublished. Acknowledgements: This case study was developed with support from The Pew Charitable Trusts and the National Science Foundation as part of the Case Studies in Science Workshop held at the State University of New York at Buffalo on May 22-26, Date Posted: 08/01/01 nas

5 BACKGROUND READING for Dr. Collins and the Case of the Mysterious Infection by Paula P. Lemons and Sarah Huber Biology Department Duke University Bacteria, Antibiotics, and the Evolution of Antibiotic-Resistance Although we can't see them, bacteria are everywhere - living on almost every surface, in the soil (one gram of surface soil contains more than 100 million bacteria), and even in some of the harshest environments on earth (e.g., sulfur pools and near-boiling undersea hydrothermal vents). They play a critical role in our ecosystem, carrying out essential processes such as nitrogen fixation and participating in symbiotic relationships with other organisms that cause no harm to the host. Disruption of the delicate interplay between bacteria and their environment is potentially very dangerous to the health and wellbeing of all organisms. Consider the bacteria Staphylococcus aureus. Normally, this species lives in the human oropharynx, nose, large intestine, vagina, and on the skin without causing harm. However, if a breach in the skin or mucosal barrier occurs, S. aureus gains access to nearby tissues or the bloodstream where it can colonize and cause disease. The relationship between S. aureus and its human host, then, is dynamic in nature, capable of quickly shifting from mutualistic or commensualistic to parasitic. The search for ways to eliminate diseases caused by bacteria led to the discovery of antibiotics. These drugs kill or inhibit the growth of susceptible bacteria. When antibiotics became widely available in the 1940s they were hailed as miracle drugs - able to cure diseases and not just reduce their symptoms. However, as early as 1950 strains of bacteria emerged that were resistant to all standard antibiotics. This problem, which has only intensified in severity since the 1950s, demonstrates the biological principle of natural selection. As antibiotic use increases, natural variants of bacteria able to resist antibiotics survive longer than antibiotic susceptible bacteria, their progeny become more numerous, and the pool of antibiotic resistance genes grows. Since bacteria easily exchange genetic information with each other, resistance genes are passed between species, genera, and families of bacteria. Today, several species of bacteria capable of causing life-threatening diseases are able to withstand exposure to every available antibiotic, creating an antibiotic resistance crisis. Antibiotics: Range of Effectiveness and Mechanisms of Action The effectiveness of antibiotics in ameliorating disease depends on two factors: 1. Spectrum. The spectrum of an antibiotic refers to the diversity of bacteria against which an antibiotic acts. Bacteria are typically classified as gram positive (e.g., S. aureus) or gram negative (e.g., Escherichia coli). Gram staining is a procedure in which bacteria are exposed to crystal violet dye, washed, and then exposed to a counter-dye. Gram positive bacteria retain the crystal violet dye while gram negative bacteria do not. Other bacteria do not fit neatly into the gram positive/gram negative classification scheme. These include classes of bacteria like mycobacteria, rickettsia, and chlamydia. Generally, narrow spectrum antibiotics act against one class of bacteria while broad spectrum antibiotics act against more than one class of bacteria. Table 1 shows the spectrum of several antibiotics.

6 2. Selective toxicity. Selective toxicity is a measure of the degree to which an antibiotic is harmful to bacteria but not the bacterial host. Antibiotics with high selective toxicity disrupt enzymes or structures that are unique to bacteria; antibiotics with low selective toxicity inhibit the same process in bacteria as in host cells, or damage host cells in some other way. Table 1 shows the selective toxicity of various antibiotics as well as the cellular processes they disrupt. Table 1. Common antibiotics and some of their characteristics. Antibiotic Spectrum Selective toxicity Mechanism of action Symptoms for which they are the drug of choice Methicillin and penicillin Narrow (gram +) High Inhibit peptidoglycan formation by binding to the enzyme transpeptidase Inflammation of the lungs, strep throat, pathogenic toxins in the blood, skin infections, gonorrhea Cefazolin Broad (gram +, some gram - ) High " Inflammation of the lungs, strep throat, pathogenic toxins in the blood, skin infections, urinary tract infections Tetracyclines Broad (gram +/-, rickettsia and chlamydia) Moderate- High Bind to the small ribosomal subunit and interfere with aminoacyl trna binding Acute diarrhea, vomiting or cramps, inflammation of the lungs, muscular pains combined with skin eruptions; Note not usually prescribed to children due to side effects during formative years Vancomycin Narrow (gram +) Low Inhibits the synthesis of peptidoglycan by binding to the amino acid polymer Inflammation of the lungs or brain meninges, ear infections Trimethoprimsulfamethoxazole (Bactrim ) Broad (gram +,-) High Inhibit folic acid synthesis (bacteria must make, mammals acquire in diet) Urinary tract infections, bronchitis, ear infections Cefazolin is an example of a broad spectrum antibiotic that functions by blocking bacterial cell wall synthesis. Bacterial cell walls, which maintain osmotic balance and provide an added layer of protection against toxic substances, are made up of a typical plasma membrane as well as additional specialized structures like the peptidoglycan (Figure 1). In gram positive bacteria, a series of enzymes that includes transglycosylase and transpeptidase catalyzes cell wall formation; cefazolin binds to and inhibits transpeptidase (Figure 2). Since animal cells do not possess cell walls, cefazolin has a high selective toxicity. Several strains of bacteria have emerged that are resistant to all cefazolin-related antibiotics (including methicillin and penicillin) and are referred to as MRSA (Methicillin resistant S. aureus). Resistance to cefazolin-related antibiotics is usually conferred by the chromosomal gene meca. MecA

7 encodes a protein that binds to and sequesters these antibiotics, preventing them from inhibiting transpeptidase (Figure 2). The most common MRSA strain is hospital-acquired MRSA which is also resistant to many other common antibiotics (e.g. tetracycline, Bactrim). Over the next week and a half you will be examining the problem of antibiotic resistance. In seminar, you will walk in the steps of Dr. Jenna Collins as she attempts to prescribe the appropriate antibiotic to her seriously ill pediatric patient. In lab, you will take an experimental approach to the problem by surveying for the prevalence of antibiotic resistant bacteria carried by you and your peers and residing in nearby water supplies. Finally, you will work with a group of students in your section to summarize the results from your lab and discuss some of the insights you've gained about this global problem.

8 References! Baquero, F. and Blazquez, J Evolution of antibiotic resistance. Tree 12(12): ! Coppoc, G. L. 10/10/99. Chemotherapy: Drug Groups. Clinical Pharmacology Online. Tampa: Gold Standard Multimedia Inc.! Levy, S. B The challenge of antibiotic resistance. Scientific American 278(3): ! Prescott, L. M., Harley, J. P., and Klein, D. A. Microbiology. Dubuque: Wm. C. Brown Publishers, 1990.! Walsh, C. Deconstructing vancomycin Science 284:

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