Introduction

Microbiology is a field of science devoted to the study of organisms that are too small to see, and therefore an engaging laboratory experience is often the key to capturing students’ interest. It was with this in mind that the course behind this book was first conceived and developed. The goal was to provide my undergraduate microbiology students with an engaging and meaningful laboratory experience that nurtured a sense of discovery and encouraged greater interest in microbiology as a topic, a field of study, or a career.

This lab manual is suitable for use in a general microbiology or bacteriology course at a two-year or four-year college or university, and works equally well for a course with a Health Science emphasis. The approach, which has been field tested by hundreds of microbiology students over several years, builds skills while reinforcing core microbiology concepts introduced in lecture. The curriculum builds from the ground up. It begins with an introduction to biosafety practices and work with biological hazards, basic but essential microscopy skills, and aseptic technique and culture methods, and builds to include more advanced methods. The progression includes a semester-long investigation of a bacterial isolate and culminates with a practical evaluation of all microbiology skills learned in the course.

Key features and learning objectives of this laboratory manual include:

• Incorporation of the American Society for Microbiology Recommended Curriculum Guidelines for Undergraduate Microbiology.
• A design that permits flexibility in the sequencing of lab activities to correlate with lecture schedule.
• Student training in BSL 1 and 2 containment practices, in compliance with CDC Biosafety in Microbiological and Biomedical Laboratories, 5ed.
• Development of core and foundational laboratory skills required for working safely and effectively with bacterial cultures.
• Exposure to investigational methods used in public health and epidemiology.
• Student-centered investigational projects that encourages students to think and work like a scientist.
• Compatibility in content to most major microbiology textbooks.

Although the sequence in which the labs are done can be varied to suit individual course schedules, some are best ordered to follow a preceding lab. With this in mind, a schedule that works well for a standard semester with 14 weeks of actual class time is provided below.

Suggested Semester Lab Schedule:

Week 1: Biosafety Practices and Procedures for the Microbiology Laboratory

Week 2: The Microscopic World

Week 3: Bacteriological Culture Methods (begin primary culture for HSMP)

Week 4: The Human Skin Microbiome Project (including an introduction to Bergey’s Manual and how to build and use a dichotomous key)

Week 5: Differential Staining Techniques

Week 6: Metabolism, Physiology, and Growth Characteristics of Cocci

Week 7: Lab Practical 1

Week 8: Metabolism, Physiology, and Growth Characteristics of Bacilli

Week 9: Microbiological Food Safety (includes selective/differential media and other physiological tests)

Week 10: Lab time for EI completion OR Microbe Mythbuster presentations

Week 11: Germ Warfare

Week 12: Epidemiology and Public Health

Week 13: Blood: The Good, the Bad, and the Ugly

Week 14: Lab Practical 2

The schedule above includes time for two lab practical exams to evaluate both acquired skills and understanding of concepts. Other assessment options include having students turn in their completed labs, or writing lab summaries (reports) as assignments each week once the lab activities are completed. For the Human Skin Microbiome Project (HSMP), the worksheets may be uploaded to a course management system for students to download and complete before handing in. Similarly, the activities included in other labs can be converted to documents that can be uploaded for student access, and used for assessment opportunities. Lab periods not allocated for lab practical can be used as library time for the Microbe Mythbuster project and student presentations.

Because so many of the techniques used are unique to a microbiology laboratory, demonstrating methods such as aseptic techniques, specific inoculation methods, and appropriate culture disposal methods to students before they perform them has proven to be a very effective way to instill confidence that they are performing the techniques correctly. This also has the additional benefit of improving lab safety and minimizing risks for students as they work with bacterial cultures.

For help with lab preparation and organization, appendices at the end of this book contain information on what supplies and equipment are needed for each lab, along with a list of suggested bacterial cultures. Since many established undergraduate microbiology labs may already have stock cultures of other organisms, the choice of which cultures to use for most labs is left up to the instructor. For some labs in which specific outcomes are necessary to illustrate a particular concept, cultures known to provide the desired outcome are specified. Cultures can be obtained through the American Type Culture Collection with ordering information available at http://www.atcc.org. Cultures are also available through several biological supply companies.

For the bacterial cultures suggested, expected outcomes for the microbiological tests students will be performing are given. These outcomes are based on the information provided in Bergey’s Manual of Determinative Bacteriology, 9th edition, which was originally developed to facilitate identification of bacterial species. Since 1992, Bergey’s Manual of Systematic Bacteriology, which has an expanded scope and encompasses relatedness among species (systematics) in addition to identification, has largely replaced the older text and has become the definitive reference on bacterial taxonomy.

Bergey’s Manual of Determinative Bacteriology (“Bergey’s Manual”) is the primary reference for the phenotypic tests and morphological characteristics of the cultures suggested for use in this book, and the basis for the Human Skin Microbiome Project, in which a bacterial isolate is presumptively identified to genus and species using traditional bacteriological culture methods and phenotypic tests. Although automated and DNA-based tests are replacing this approach in clinical laboratories, for this lab experience the emphasis is on the investigative process and Bergey’s Manual fits the need perfectly. The book is available in paperback form (from Amazon and Carolina), and it is suggested that it be made available as a resource students can use while in the laboratory.

One final consideration is the lab time. Although all of the labs can be completed in three- hour lab periods, many require incubation time to permit bacterial cultures to metabolize, reproduce, and grow colonies or other forms of culture growth. To accommodate this need, we have made available additional time for students to return to lab and complete experiments. “Open Lab” times are supervised by laboratory assistants (most are work-study students who successfully complete the course) who have been trained to provide technical assistance to the course by making culture media, prepping and cleaning up after the lab each week, and serving as peer tutors. Although most students have been able to take advantage of the Open Lab time, for campuses with a large population of students who commute or have families and/or jobs, coming back to lab the next day isn’t always a possibility. We have solved this by giving students the option of incubating cultures at a lower temperature if they know they will be unable to return to lab; students can also arrange to have their cultures transferred to a refrigerator to preserve the results of their tests.