1 Title: Teaching Crafty Microbiology: Safely Teaching Hands-on Microbiology Skills at Home 2 3 Authors: Anne M. Estes 1*^ , Anna K.S. Jozwick 2*, Jennifer E. Kerr 3* 4 5 Affiliations:1 Towson University, Department of Biological Sciences; 2 Goucher College, Center for Natural 6 Sciences; 3 Notre Dame of Maryland University, Department of Biology 7 8 * Authors contributed equally 9 10 ^Corresponding author. Mailing address: Towson University 8000 York Road, Towson, MD 21204. Phone: 410- 11 11 704-2126. E-mail: [email protected] 12 13 Running head or foot line of approximately 40 characters: Teaching Crafty Microbiology 14 15 Number of figures (1), tables (2), and supplemental materials (4). 16 17 The authors declare no conflicts of interest. 18 Using non-toxic craft items and disposable lab consumables, we have developed nine modules to teach fundamental,

19 hands-on microbiology lab skills safely at home. These ‘Crafty’ teaching modules can be paired with virtual

20 instruction and/or data collected by an instructor to replicate traditional unknown microbiology lab exercises.

21 Materials and procedures used were carefully chosen to best mimic the texture of ‘media,’ represent microbial

22 ‘diversity,’ assess aseptic technique, and produce analyzable data from results. Some protocols build upon and

23 extend previously unpublished ideas, while others provide novel methods. The lab skills include proper PPE usage

24 and basic biosafety, aseptic technique, microscopy and staining, streaking for isolation, spread plating, serial

25 dilutions, filtering, disk diffusion method, and modeling an epidemic. Each protocol includes a student handout with

26 background, links to videos of the methods performed with microbes, a rationale for the pairing of craft and

27 consumable lab supplies along with technique used, a video/image demonstration of the ‘Crafty’ technique when

28 needed, post-lab questions, and an instructor guide. This resource was developed for an undergraduate microbiology

29 course and each lab is aligned with learning outcomes within American Society for Microbiology’s undergraduate

30 curriculum guidelines. This work would also be useful for outreach and K-12 educators. The development of

31 microbiology lab skills by all students, regardless of economic or health status, will lead to a more scientifically

32 minded society.

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34 Keywords: 3-10

35 Aseptic techniques, biosafety, microbiology teaching, hands-on lab skills

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42 Introduction

43 The COVID-19 pandemic forced microbiology teaching labs out of a directly supervised, controlled

44 laboratory setting and into student homes. Current solutions have been to either send home pre-assembled kits with

45 viable microbes or rely on virtual labs (1). Pre-assembled kits, whether commercially available or assembled by the 46 instructor, typically contain live Risk Group Level 1 microbes that are safe to work with in a supervised lab, yet

47 could present a health risk in home environments with immunocompromised people, young children, and/or pets (2).

48 Virtual labs do not rely on live microbes; however, they fail to provide hands-on skills practice and depend on

49 internet access and stability. Additionally, both pre-assembled kits and virtual labs are costly. Our goal was to teach

50 essential microbiology skills at home using inexpensive craft supplies and disposable microbiology tools (Appendix

51 1).

52 With ‘Crafty Microbiology’ kits (Figure 1), students develop authentic motor skills for microbiology in a

53 safe and economical fashion in their own home. While some of the methods have been used previously (3-5), we

54 present improvements, extensions, or quantitative assessments. The PPE removal exercise uses colored paint, glitter,

55 and glow-in-the-dark paint. These different materials add multiple layers for students and professors reviewing

56 student videos, to assess glove-removal issues. In isolating single colonies, students use three types of model media

57 prior to using gelatin or agar-based media. Each model media helps students assess appropriate streak pattern, sterile

58 technique, loop angle, and pressure. Self-assessment guides, including assessing aseptic technique via the ‘bench

59 space’, and quantification further deepen the ‘Crafty’ methods. Modifications have been made to previously

60 unpublished exercises, including 1) the pairing of serial dilutions with mock CFU calculations for mixed culture

61 samples, 2) filtering demonstrations using different weaves of fabrics and a heterogenous sized glitter mixture, 3)

62 microscopy exercises that incorporate artwork (modeling and collage) as a means to demonstrate outcomes of

63 various staining techniques, and 4) spread plating inoculation to provide immediate feedback on technique with

64 biosafety incorporation. Some novel methods, that attempt to replicate the actual microbial technique as closely as

65 possible, include a mock Kirby Bauer experience that uses disc diffusion based color changes via soluble marker ink

66 or pH differences and epidemic modeling exercise, which pairs computer modeling with a hands-on disease-

67 transmission demonstration. Throughout the techniques special attention was paid to the texture of the ‘media’,

68 representing microbial ‘diversity’ and numbers, ability to assess aseptic technique, and the results produced by each

69 procedure. 70

71 Procedure

72 Student Buy-in

73 To alleviate any concerns students might have in time or money being wasted on a box of craft supplies,

74 teaching transparency is essential (7). Thus, each protocol includes a rationale on how the craft supply choice

75 mimics the laboratory supplies. Exercises are paired with learning objectives (Table 1), video or images

76 demonstrating proper technique using the ‘Crafty supplies’ (Appendix 1), a link to demonstration/images using live

77 microbes, and a standard microbiology lab textbook explanation. Comparing the crafty versus standard methods side

78 by side allows students to understand the fundamental learning objective, motor and quantitative microbiology lab

79 skills they gain.

80 81 Activities

82 Nine crafty labs (Table 1) were created with full student handouts, instructor guides, suggested supply list

83 (Appendix 2 and 3, and Table 2, respectively) and videos (Mostly Microbes’ “Crafty Microbiology” YouTube

84 Playlist: https://www.youtube.com/playlist?list=PL-pe-Oi8vx5PskybWBnpC7hTEZ5d5RETa ) of certain

85 techniques. For each exercise, students create their ‘bench space’ using packing paper. They assemble the materials

86 needed for that exercise, watch the live microbe and ‘Crafty’ video/image demonstrations, and attempt the crafty

87 procedure. When they feel confident about their technique, they make a video of themselves performing the

88 technique, upload the video to their electronic notebook, evaluate their technique, and then clean up any supplies for

89 subsequent use. To address aseptic technique, students check their ‘bench space’ (packing paper), for glitter or paint

90 contamination after each procedure.

91 An example of the importance of material choice can be seen when conducting a quadrant streak to isolate

92 individual colony forming units. We needed a proper ‘media’ mimic and ‘microbial colony.’ Students often have

93 trouble with holding the loop at the correct angle that prevents gouging the media, but allows for adequately

94 spreading the cultures. The media needed to be slightly slippery, reducing friction between the media surface and the

95 inoculation loop. After much experimentation with different paper types, we determined that our ideal ‘media’

96 solution was to stretch parafilm over the edges of the bottom dish of a petri dish. This allowed for a stable, but

97 delicate surface that the loop could glide over dispersing the microbial ‘culture.’ The microbial ‘culture’ sample was

98 composed of two different commercially available glitter paints mixed together. These silver and gold paints (10) 99 (Table 2) have slightly different sized glitter that can be distinguished from each other. Finally, to assess aseptic

100 technique, additional glow-in-the-dark paint can be added. To make the microbial ‘sample’ more diverse, several

101 colors of super-fine glitter can be added to the mix. The glitter size is very important in this case. The glitter needs to

102 be small enough so that it can be easily picked up and transferred by a loop tool.

103 These labs are designed to be modular, allowing instructors to include as many exercises as needed or to

104 change the order to fit their course design. Each module is individually assessed using 1) post lab questions included

105 in each student handout (Appendix 2) and 2) an image and/or video of the students’ technique(s) using the 3-point

106 (Poor, Adequate, Proficient) rubrics as a guide for grading (Appendix 4). Post lab questions are intended to highlight

107 the biological context of the lab technique covered in the module, connect a learning objective to the technique

108 and/or provide exploration of the differences in the crafty technique and the standard technique. Videos and images

109 allow for personalized feedback not always available during in the in-person lab setting when one instructor may

110 have many students in one lab section to monitor for technique.

111 Extension 112 While these techniques were developed in response to the COVID-19 pandemic, they have numerous

113 benefits and applications. During in-person teaching labs, students are often highly anxious about safely working

114 with live microbes, their abilities, and the 24 hours or more lag before results are known. In particular, properly

115 isolating single colonies is extremely problematic and may keep a student stuck at that skill set for weeks. When

116 working with microscopic organisms, many students use too much inoculum since they are unsure that anything is

117 being manipulated. Providing students with materials to work with in the comfort of their own home that closely

118 mimic their in-lab resources gives them an opportunity to practice prior to coming into the lab and/or to supplement

119 standard procedures. This should increase skill success and decrease waste of laboratory materials. An unintentional

120 side benefit to conducting these skills at home is that others in the student’s household may be inspired to try to

121 learn more about microbiology. Indeed, students reported that parents and siblings were intrigued about the ‘crafty’

122 exercises.

123 ‘Crafty Microbiology’ can easily be adopted for use with outreach activities and K-12 educators. Thus, the

124 public can better understand for example, the essential techniques that clinical microbiologists use to isolate

125 microbes from clinical samples or how antibiotic resistance is determined.

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127 Safety Issues

128 There are no safety issues as no biological materials are used. Craft materials are standard, consumer art

129 supplies and the consumable lab materials are plastic. The supplies suggested were chosen since they are non-toxic

130 and washable.

131

132 Conclusion

133 Initial responses have shown these ‘Crafty’ methods do provide real and immediate hands-on skills

134 feedback. Assessment rubrics were created for the nine crafty labs (Appendix 4). Post-lab questions are included

135 with each student handout to reinforce the learning objectives and assess students’ mastery of the material. The

136 efficacy of the ‘Crafty Microbiology’ exercises will be assessed in both intro and upper level microbiology classes

137 Spring 2021 (Goucher College IRB #20141673). We anticipate that these unique, crafty methods provide a way to

138 successfully engage students with hand-on skills without the need for live microorganisms, increase accessibility to 139 microbiology protocols, and promote a critical thinking environment as students master learning objectives and

140 essential motor skills safely at home.

141 142 REFERENCES

143 1. Mawn M, Herzog J, Horak R. Remote Teaching for ASM Microbiology Laboratory Skills. 2020.

144 Available from:

145 https://docs.google.com/document/d/1NJNwjdId3Kv0gRyxtVKQ47HBn4VlLJNEp1tPtaXcrL4/edit

146

147 2. Herzog J & Mawn M. 2020. Teaching Lab-Based Courses Online & Remote: From “Are You Kidding

148 Me?” to “This Is Effective!” Available from: https://asm.org/Articles/2020/May/Teaching-Lab-Based-

149 Courses-Online-Remote-From-Are

150 151 3. Byrd J, Emmert E, Maxwell R, & Townsend H. 2019. ASM Guidelines for Biosafety in Teaching

152 Laboratories: Biosafety Considerations Regarding at-home or DIY Microbiology Kits, on American

153 Society of Microbiology. [Cited 2020 Aug 21]. Available: https://asm.org/ASM/media/Education/ASM-

154 Biosafety-Considerations-with-At-Home-Microbiology-Kits-1.pdf

155

156 4. Hartman D. 2011. Perfecting Your Spread Plate Technique. J. Microbiol. Biol. Educ., 12:2, 204–205.

157 doi:10.1128/jmbe.v12i2.324

158 5. Axler-DiPerte GL. 2017. Modeling and Visualizing Bacterial Colony Purification Without the Use of

159 Bacteria or Laboratory Equipment. J. Microbiol. Biol. Educ. 18:18.2.41. doi: 10.1128/jmbe.v18i2.1308

160 6. Tomas ME., Kundrapu S., Thota P., Sunkesula VC., Cadnum JL., Mana TS., Jencson A., O'Donnell M.,

161 Zabarsky TF., Hecker MT., Ray AJ., Wilson BM., & Donskey CJ. 2015. Contamination of Health Care

162 Personnel During Removal of Personal Protective Equipment. JAMA internal medicine, 175(12), 1904–

163 1910. https://doi.org/10.1001/jamainternmed.2015.4535

164 7. Winkelmes MA. 2013.Transparency in Learning and Teaching: Faculty and Students Benefit Directly

165 From A Shared Focus On Learning and Teaching Processes. NEA Higher Education Advocate 6 -

166 9.

167 168 8. Merkel S & The ASM Task Force on Curriculum Guidelines for Undergraduate Microbiology. 2012.

169 The Development of Curricular Guidelines for Introductory Microbiology that Focus on Understanding. J.

170 Microbiol. Biol. Educ. 13:1, 32-38. doi:10.1128/jmbe.v13i1.363

171

172 9. Norman-McKay L & The ASM MINAH Undergraduate Curriculum Guidelines Committee. 2018.

173 Microbiology in Nursing and Allied Health (minah) Undergraduate Curriculum Guidelines: A Call to

174 Retain Microbiology Lecture and Laboratory Courses in Nursing and Allied Health Programs. J. Microbiol.

175 Biol. Educ. 19(1). doi:10.1128/jmbe.v19i1.1524

176

177 10. Sargent Art ®. Washable Glitter Gold (ITEM: #17-3781) and Washable Glitter Silver (ITEM: #17-

178 3782). Art-Time Washable Tempera Paint. [Cited 2020 Aug 21] Available from:

179 http://sargentart.com/sargent-art-art-time-washable-tempera-paint/

180

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182 183 Appendix 1: Master Supplies List and Recipes

184 Appendix 2: Student Handouts

185 Appendix 3: Instructor Guides

186 Appendix 4: Assessment Rubric

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191 192 193 194

195 Figure 1. What’s in the ‘Crafty’ Micro Box? Lab supplies: Disposable loops/needles, petri dishes, bulb pipettes,

196 parafilm, conical tubes, gloves, gelatin, L-shaped plastic spreader, filter paper, permanent marker, cotton swabs,

197 paper discs, agar only slants, ‘mock Bunsen burner tube’, microcentrifuge tubes, ruler; Craft supplies: Packing

198 paper, high density Glitter & Glow paint, wax paper, aluminum foil, glow-in-the-dark paint, different sized fabrics,

199 colored paper, pipe cleaners, washable marker, rubber band, red cup, dark tempra paint, glitter filtration mixture.

200 The cost per student is approximately $6.82.

201 202 Table 1. Microbiology lab technique learning objects and curriculum alignment. *ASM undergraduate 203 curriculum guidelines (8) and microbiology in nursing and allied health (MINAH) undergraduate curriculum 204 guidelines in parentheses (9). 205 Learning Objectives Curriculum Microbiology Lab Guideline Number* Technique

PPE and Aseptic -Demonstrate and assess the proper use of PPE 36,37 Technique -Demonstrate the steps to follow for putting on and safely removing (22,24) -Glove removal PPE -Recognize the importance of hand hygiene in aseptic technique -'Biohazard -Recognize how to properly dispose of materials in the appropriate Container' use receptacles (general/universal trash, 'biohazard') Microscopy and -Demonstrate understanding of light microscope total magnification 6,32 Staining relative to the microscope field of view and specimen type (22, 23) -Recognize the procedures and resulting microscopic images for simple, Gram, endospore, acid-fast, and negative capsule staining

Streak for -Display proper aseptic technique in sample transfer or inoculation 33,36 Isolation and -Demonstrate proper streaking for isolation using the quadrant (23) method. -Demonstrate skill in isolation of items from a mixed sample using different types of solid media. -Learn the procedures used in preparing media needed for culturing microorganisms. Spread Plating -Demonstrate proper aseptic technique for transfer of samples to a 33,34,36 plate (23) -Learn a technique used to characterize and/or count the number of microorganisms in a sample

Inoculating Liquid -Demonstrate proper aseptic technique for transfer of samples to a 33,34,36 and Slant Media broth and slant (23)

Serial Dilutions, -Practice and explain the importance of sterile technique 29,33,35,36 Plating, and -Calculate dilution factors and perform serial dilutions effectively (23) Counting

Filtering -Understand the physical method of controlling microbial growth via 14,31,33,36 filtration (15) -Practice a technique to enrich for and isolate microorganisms.

Disk Diffusion -Demonstrate the disk diffusion method of antimicrobial susceptibility 29, 36 Method (Kirby testing (25) Bauer) -Interpret zones of inhibition qualitatively as resistant, intermediate, or susceptible. -Determine which agent(s) is best for 'treatment' when multiple zones of inhibition are present. Epidemic -Understand that diseases spread more slowly when the rate of 23, 28, 31 Modeling contact, and thus transmission, is lower. (12,13)

206 207 208 Table 2. Crafty microbiology student supplies and approximate costs.

209