This laboratory manual and workbook,
now in its seventh edition, maintains its original
emphasis on the basic principles of diagnostic microbiology for
students preparing to enter the allied health professions.
It remains oriented primarily toward meeting the interests
and needs of those who will be directly involved in
patient care and who wish to learn how microbiological
principles should be applied in the practice of their professions.
These include nursing students, dental hygienists,
dietitians, hospital sanitarians, inhalation therapists, operating room
or cardiopulmonary technicians, optometric technicians,
physical therapists, and physicians’ assistants. For such
students, the clinical and epidemiological applications of
microbiology often seem more relevant than its technical
details. Thus, the challenge for authors of textbooks and
laboratory manuals, and for instructors, is to project microbiology
into the clinical setting and relate its principles to
patient care.
The authors of this manual have emphasized the purposes and functions of the clinical
microbiology laboratory in the diagnosis of infectious
diseases. The exercises illustrate as simply as possible the
nature of laboratory procedures used for isolation and
identification of infectious agents, as well as the principles of
asepsis, disinfection, and sterilization. The role of the health
professional is projected through stress on the importance of
the clinical specimen submitted to the laboratory—its
proper selection, timing, collection, and handling. Equal
attention is given to the applications of aseptic and
disinfectant techniques as they relate to practical situations in the
care of patients. The manual seeks to provide practical
insight and experience rather than to detail the microbial
physiology a professional microbiologist must learn. We have
approached this revision with a view toward updating
basic procedures and reference sources. Every exercise has
been carefully reviewed and revised, if necessary, to conform
to changing practices in clinical laboratories. A new
exercise, Exercise 19, has been prepared describing modern
diagnostic techniques that use antigen detection and
nucleic acid methods. These methods are now in use in many
clinical microbiology laboratories. When relevant, antigen
detection methods have been added to the exercises, so that
the students will gain experience in their use.
Expanded sections on diagnosing microbial pathogens that require
special laboratory techniques are included in the exercises
of Section XI. Many new figures and additional
colorplates are found in this edition. These are intended to
illustrate procedures the students will use and help the beginning
student recognize the microbes they will view under the
microscope as well as the appropriate reactions for
biochemical tests they will perform.
The material is organized into four parts of increasing complexity designed to give students
first a sense of familiarity with the nature of microorganisms, then practice in aseptic cultural methods in clinical
settings. Instructors may select among the exercises or
parts of exercises they wish to perform, according to the focus
of their courses and time available. Part 1 introduces
basic techniques of microbiology. It includes general
laboratory directions, precautions for handling
microorganisms, the use of the microscope, microscopic morphology of
microorganisms in wet and stained preparations, pure
culture techniques, and an exercise in environmental
microbiology.
Part 2 provides instruction and some experience in methods for the destruction of
microorganisms, so that students may understand the
principles of disinfection and sterilization before proceeding to
the study of pathogenic microorganisms. There is an exercise
on antimicrobial agents that includes antimicrobial
susceptibility testing using the National Committee for
Clinical Laboratory Standards (NCCLS) technique, with the
latest category designations and inhibition zone
interpretations, as well as experiments to determine minimal
inhibitory concentrations by the broth dilution method,
and bacterial resistance to antimicrobial agents.
The principles learned are then applied to diagnostic microbiology in Part 3. Techniques
for collecting clinical specimens (Microbiology at the
Bedside) and precautions for handling them are reviewed. A
discussion of the Centers for Disease Control and
Prevention “standard precautions” for avoiding transmission
of bloodborne pathogens is included. The normal
flora of various parts of the body is discussed. The five
sections of this part cover the principles of diagnostic
bacteriology; the microbiology of the respiratory, intestinal,
urinary, and genital tracts; and the special techniques required for
the recognition of anaerobes, mycobacteria,
mycoplasmas, rickettsiae, chlamydiae, viruses, fungi, protozoa,
and animal parasites. Sections VIII and IX, dealing respectively
with the microbiology of the respiratory and
intestinal tracts, present exercises on the common pathogens and
normal flora of these areas, followed by exercises
dealing with methods for culturing appropriate clinical
specimens. Experiments for performing antimicrobial
susceptibility tests on relevant isolates from such specimens are also
included.
The former Part 4 has been incorporated into Part 3, reflecting the essential role of
antigen detection techniques in the routine laboratory and the
more limited use of methods for detecting serum
antibodies. Part 4 presents some simple microbiological methods
for examining water and milk.
The sequence of the exercises throughout the manual, but particularly in Part 3, is
intended to reflect the approach of the diagnostic laboratory
to clinical specimens. In each exercise, the student is led
to relate the practical world of patient care and clinical
diagnosis to the operation of the microbiology laboratory.
To learn the normal flora of the body and to appreciate
the problem of recognizing clinically significant
organisms in a specimen containing mixed flora, students collect
and culture their own specimens. Simulated clinical
specimens are also used to teach the microbiology of infection.
The concept of transmissible infectious disease becomes a
reality, rather than a theory, for the student who can see
the myriad of microorganisms present on hands,
clothes, hair, or environmental objects, and in throat, feces,
and urine. Similarly, in learning how antimicrobial
susceptibility testing is done, the student acquires insight into the
basis for specific drug therapy of infection and the
importance of accurate laboratory information.
In acquiring aseptic laboratory technique and a knowledge of the principles of
disinfection and sterilization, the student is better prepared
for subsequent encounters with pathogenic, transmissible microorganisms in professional practice. The authors
believe that one of the most valuable contributions a
microbiology laboratory course can make to patient care is to
give the student repeated opportunities to understand
and develop aseptic techniques through the handling of
cultures. Mere demonstrations have little value in
this respect. Although the use of pathogenic microorganisms
is largely avoided in these exercises, the students are
taught to handle all specimens and cultures with respect, since
any microorganism may have potential pathogenicity. To
illustrate the nature of infectious microorganisms, material
to be handled by students includes related
“nonpathogenic” species of similar morphological and cultural
appearance, and demonstration material presents pathogenic
species. Occasional exceptions are made in the case of
organisms such as pneumococci, staphylococci, or clostridia
that are often encountered, in any case, in the
flora of specimens from healthy persons. If the instructor so
desires, however, substitutions can be made for these as
well.
Teaching flexibility has been sought throughout the manual. There are 35 exercises, many
of which contain general experiments. These may be
tailored to meet the needs of any prescribed course
period, the weekly laboratory hours available, or the
interests and capabilities of individual students. The manual can be
adapted to follow any textbook on basic microbiology
appropriate for students entering the allied health field. For
the instructor’s use, a more complete listing of current
literature and other source material is provided in the
Instructor’s Manual.
Each exercise begins with a discussion of the material to be covered, the rationale of
methods to be used, and a review of the nature of
microorganisms to be studied. In Part 3, tables are frequently
inserted to summarize laboratory and/or clinical information
concerning the major groups of pathogenic microorganisms.
The questions that follow each exercise are designed to
test the ability of students to relate laboratory
information to patient-care situations and to stimulate them to read
more widely on each subject presented.
The five appendices included in previous editions of this manual have been moved to the
Instructor’s Manual to provide instructors with information
and assistance in presenting the laboratory course.
Sadly, our long-term colleague and original inspiration for this laboratory manual, Dr.
Marion Wilson, passed away during the initial stages of
this revision. We dedicate this edition to her. We are fortunate
in being joined by Dr. Paul Granato, who is responsible for
much of the new material in Exercise 19 and Sections X
and XI.
We are grateful to all those professional colleagues who gave generously of their time and
expertise to make constructive suggestions regarding
the revision of this manual. For their helpful comments
and reviews, we thankCaroline Amiet, Odessa College; John
Mark Clauson, Western Kentucky University; Angel
Gochee, Indiana University; John Ferrara, Cuyahoga
Community College; Fernando Monroy, Indiana State
University; David Stetson, University of Maine; Martin
Steinbeck, Mid-Plains Community College; and Jane
Weston, Genesse Community College. We owe special
thanks to Dr. Edward Bottone, Mount Sinai Hospital,
New York, for providing us with several of the
photographs in the color plates, Dr. Nancy Morello,
Massachusetts Bay Community College for her advice on revisions,
and to Mr. Scott Matushek, Mr. Gordon Bowie, and Ms.
Liane Duffee-Kerr of the University of Chicago for
their photographic assistance.
Finally, we acknowledge the role of McGraw-Hill in publication of this work. Their many
courtesies, extended through Jean Fornango, senior developmental editor, have encouraged and guided this new
edition, and they have been primarily responsible for
its production. For her skillful efforts and expert
assistance during the production process, we thank Sheila Frank,
project manager. We also acknowledge Laura Fuller, senior
production supervisor, Rick D. Noel, design coordinator,
Carrie K. Burger, lead photo research coordinator, and Tammy
Juran, senior media project manager, who contributed
to the style and
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