Table Of Content2
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COMPLETE VERSION
Benson’s
Microbiological Applications
Laboratory Manual in General Microbiology
Fourteenth Edition
Alfred E. Brown
Emeritus Professor, Auburn University
Heidi R. Smith
Front Range Community College
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BENSON’S MICROBIOLOGICAL APPLICATIONS: LABORATORY MANUAL IN GENERAL MICROBIOLOGY, COMPLETE
VERSION, FOURTEENTH EDITION
Published by McGraw-Hill Education, 2 Penn Plaza, New York, NY 10121. Copyright © 2017 by McGraw-Hill Education. All rights reserved.
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About the Authors
Alfred Brown
Emeritus Professor of Microbiology
Auburn University
B.S. Microbiology, California
State College, Long Beach
Ph.D. Microbiology, UCLA
Teaching Dr. Brown has been a member of the American Society for Microbiology for 50 years, and he has
taught various courses in microbiology over a teaching career that spans more than 30 years. Courses have
included general microbiology, medical microbiology, microbial physiology, applied and environmental
microbiology, photosynthesis, microbiological methods, and graduate courses such as biomembranes. In 2008,
Dr. Brown retired from the Auburn University faculty as an emeritus professor of microbiology.
Administration During his tenure at Auburn University, Dr. Brown served as the director of the University
Electron Microscope Facility. He also served as the chair of the Department of Botany and Microbiology and the
chair of the Department of Biological Sciences.
Research My research has focused on the physiology of the purple nonsulfur bacteria. This has involved how
bacteriochlorophyll and photosynthetic membrane synthesis are coordinated. Herbicides, such as atrazine, have
been used to determine the binding site for ubiquinone in photosynthetic electron transport. Binding occurs on
the L-subunit, a protein in the photosynthetic reaction center. Resistance to atrazine involves a single amino acid
change in the L-subunit that prevents the herbicide from binding to the protein and inhibiting electron transport.
This is comparable to how atrazine inhibits electron transport in plants and how resistance to these herbicides
develops. My laboratory also investigated how the sulfonylurea herbicides inhibit acetolactate synthase, a crucial
enzyme in the pathway for branched-chain amino acids. Recently, I consulted for a company that manufactures
roofing shingles. Because of the presence of calcium carbonate in shingles, cyanobacteria can easily grow on their
surface, causing problems of contamination. My laboratory isolated various species of cyanobacteria involved in
the problem and taxonomically characterized them. We also tested growth inhibitors that might be used in their
control.
Dr. Brown and his wife have traveled extensively in Europe since his retirement. His three children have
followed him in science, having earned doctorates in physics, chemistry and anatomy.
Heidi Smith
Front Range Community College
B.A. Biology/Pre-Medicine, Taylor University, IN
M.S. Biology, Bowling Green State University, OH
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Heidi Smith is the lead faculty for microbiology at Front Range Community College in Fort Collins, CO, and
teaches a variety of biology courses each semester including microbiology, anatomy/physiology, and
biotechnology. Heidi also serves as the principal investigator on a federal grant program designed to increase
student success in transfer and completion of STEM degrees at the local university. In that role, Heidi works
directly with students to train them for and support them through summer undergraduate research experiences.
Student success is a strategic priority at FRCC and a personal passion of Heidi’s. She continually works to
develop professionally in ways that help her do a better job of reaching this important goal. During the past 7
years, Heidi has had the opportunity to collaborate with faculty throughout the United States in developing digital
tools, such as SmartBook, LearnSmart Labs, and Connect, that measure and improve student learning outcomes.
This collaborative experience with these tools has revolutionized her approach to teaching in both face-to-face and
hybrid courses. The use of digital technology has given Heidi the ability to teach courses driven by real-time
student data and with a focus on active learning and critical thinking activities.
Heidi is an active member of the American Society for Microbiology. She has presented instructional
technology and best online and face-to-face teaching practices on numerous occasions at the annual conference for
undergraduate educators. She also served as a member of the ASM Task Force on Curriculum Guidelines for
Undergraduate Microbiology Education, assisting in the identification of core microbiology concepts as a guide to
undergraduate instruction.
Off campus, Heidi spends as much time as she can enjoying the beautiful Colorado outdoors with her husband
and three young children.
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Contents
Preface
Digital Tools for Your Success
Basic Microbiology Laboratory Safety
Biosafety Levels for Selected Infectious Agents
Microorganisms Used or Isolated in the Lab Exercises in This Manual
PART 1 Microscopy
1 Brightfield Microscopy
2 Darkfield Microscopy
3 Phase-Contrast Microscopy
4 Fluorescence Microscopy
5 Microscopic Measurements
PART 2 Survey of Microorganisms
6 Microbiology of Pond Water—Protists, Algae, and Cyanobacteria
7 Ubiquity of Bacteria
8 The Fungi: Molds and Yeasts
PART 3 Manipulation of Microorganisms
9 Aseptic Technique
10 Pure Culture Techniques
PART 4 Staining and Observation of Microorganisms
11 Smear Preparation
12 Simple Staining
13 Negative Staining
14 Capsular Staining
15 Gram Staining
16 Spore Staining: Two Methods
17 Acid-Fast Staining: Kinyoun Method
18 Motility Determination
PART 5 Culture Methods
19 Culture Media Preparation
20 Preparation of Stock Cultures
21 Enumeration of Bacteria: The Standard Plate Count
22 Slide Culture: Fungi
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PART 6 Bacterial Viruses
23 Determination of a Bacteriophage Titer
24 A One-Step Bacteriophage Growth Curve
25 Isolation of Phages from Flies
26 Phage Typing
PART 7 Environmental Influences and Control of Microbial Growth
27 Effects of Oxygen on Growth
28 Temperature: Effects on Growth
29 pH and Microbial Growth
30 Water Activity and Osmotic Pressure
31 Ultraviolet Light: Lethal Effects
32 The Effects of Lysozyme on Bacterial Cells
33 Evaluation of Alcohol: Its Effectiveness as an Antiseptic
34 Antimicrobic Sensitivity Testing: The Kirby-Bauer Method
35 Evaluation of Antiseptics and Disinfectants: The Filter Paper Disk Method
36 Effectiveness of Hand Scrubbing
PART 8 Identification of Unknown Bacteria
37 Morphological Study of an Unknown Bacterium
38 Cultural Characteristics
39 Physiological Characteristics: Oxidation and Fermentation Tests
40 Physiological Characteristics: Hydrolytic and Degradative Reactions
41 Physiological Characteristics: Multiple Test Media
42 Use of Bergey’s Manual
PART 9 Miniaturized Multitest Systems
43 Enterobacteriaceae Identification: The API® 20E System
44 Enterobacteriaceae Identification: The EnteroPluri-Test System
45 Staphylococcus Identification: The API® Staph System
PART 10 Diversity and Environmental Microbiology
46 Isolation of an Antibiotic Producer: The Streptomyces
47 Nitrogen Cycle: Ammonification
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48 Symbiotic Nitrogen Fixation: Rhizobium
49 Free-Living Nitrogen Fixation: Azotobacter
50 Denitrification: Paracoccus denitrificans
51 The Winogradsky Column
52 Purple Nonsulfur Photosynthetic Bacteria
53 Sulfate-Reducing Bacteria: Desulfovibrio
54 Bacterial Commensalism
55 Bacterial Synergism
56 Microbial Antagonism
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PART 11 Applied Microbiology
57 Bacterial Counts of Foods
58 Bacteriological Examination of Water: Most Probable Number Determination
59 Bacteriological Examination of Water: The Membrane Filter Method
60 Reductase Test
61 Temperature: Lethal Effects
62 Microbial Spoilage of Canned Food
63 Microbiology of Alcohol Fermentation
PART 12 Bacterial Genetics and Biotechnology
64 Understanding Antibiotic Resistance: Replica Plating
65 Polymerase Chain Reaction for Amplifying DNA
66 Bacterial Transformation
PART 13 Medical Microbiology
67 The Staphylococci: Isolation and Identification
68 The Streptococci and Enterococci: Isolation and Identification
69 Gram-Negative Intestinal Pathogens
70 A Synthetic Epidemic
PART 14 Immunology and Serology
71 Slide Agglutination Test: Serological Typing
72 Slide Agglutination Test for S. aureus
73 Slide Agglutination Test for Streptococcus
74 Enzyme-Linked Immunosorbent Assay (ELISA)
75 Blood Grouping
Appendix A Tables
Appendix B Indicators, Stains, Reagents
Appendix C Media
Appendix D Identification Charts
Appendix E Metric and English Measurement Equivalents
Index
Indicates a LearnSmart Lab® activity is available for all or part of this exercise. For more
information, visit http://www.mhhe.com/LearnSmartLabsBiology/
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