Table Of ContentGene Probes for Bacteria
Edited by
Alberto J. L. Macario
Everly Conway de Macario
Wadsworth Center for Laboratories and Research
New York State Department of Health
and
School of Public Health
State University of New York
Albany, New York
ACADEMIC PRESS, INC.
Harcourt Brace Jovanovich, Publishers
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Library of Congress Cataloging-in-Publication Data
Gene probes for bacteria / edited by Alberto J. L. Macario, Everly
Conway de Macario.
p. cm.
Includes bibliographical references.
ISBN 0-12-463000-6 (alk. paper)
1. Diagnostic bacteriology-Technique. 2. DNA probes—Diagnostic
use. I. Macario, Alberto J. L. II. Conway de Macario, Everly.
[DNLM: 1. Bacteria—isolation & purification. 2. Nucleic Acid
Probes. 3. Nucleic Acids-diagnostic use. QU 58 G326]
QR67.2.G46~dc20
DNLM/DLC
for Library of Congress 89-17733
CIP
Printed in the United States of America
90 91 92 93 9 8 7 6 5 4 3 2 1
To our collaborators, assistants, α Aid students
of sundry places and times.
Contributors
Numbers in parentheses indicate the pages on which the authors' contributions begin.
Peter W. Andrew (179), Department of Microbiology, University of
Leicester, Medical Sciences Building, Leicester LEI 9HN, England
G. T. Attwood (389), Department of Animal Sciences, Waite Agriculture
Research Institute, Glen Osmond, South Australia 5064, Australia
Christiane Bebear (45), Laboratoire de Bacteriologie, Hopital Pellegrin,
33076 Bordeaux, France
Lawrence H. Bopp (69), Laboratories for Bacteriology, Wadsworth Cen
ter for Laboratories and Research, New York State Department of
Health, Albany, New York 12201-0509
Graham J. Boulnois (179), Department of Microbiology, University of
Leicester, Medical Sciences Building, Leicester LEI 9HN, England
J. D. Brooker (389), Department of Animal Sciences, Waite Agriculture
Research Institute, Glen Osmond, South Australia 5064, Australia
Arunsri Chatkaeomorakot (95), Armed Forces Research Institute of Medi
cal Sciences, Bangkok 10400, Thailand
Everly Conway de Macario (485), Wadsworth Center for Laboratories and
Research, New York State Department of Health, and School of
Public Health, State University of New York, Albany, New York
12201-0509
Hirofumi Danbara (167), Department of Bacteriology, The Kitasato Insti
tute, Minato-ku, Tokyo 108, Japan
James J. Donegan (1), Enzo Biochem, Inc., New York, New York 10013
Ram Dular (417), Ontario Ministry of Health, Regional Public Health
Laboratory, Ottawa, Ontario K2A 1S8, Canada
xiii
xiv Contributors
Brigitte Dutilh1 (45), Laboratoire de Bacteriologie, Hopital Pellegrin,
33076 Bordeaux, France
Peter Echeverria (95), Armed Forces Research Institute of Medical Sci
ences, Bangkok 10400, Thailand
Sherry P. Goltz (1), Enzo Biochem, Inc., New York, New York 10013
Patrick A. D. Grimont (45), Unite des Enterobacteries, Unite 199
INSERM, Institut Pasteur, 75724 Paris, France
Roy Gross (205), Sclavo Research Center, 53100 Siena, Italy
David J. Groves (233), Department of Pathology, McMaster University,
and Department of Laboratory Medicine, St. Joseph's Hospital,
Hamilton, Ontario L8N 4A6, Canada
Alan M. Johnson (255), Department of Clinical Microbiology, School of
Medicine, Flinders University and Flinders Medical Center, Bedford
Park, South Australia 5042, Australia
Robert J. Jovell (485), Wadsworth Center for Laboratories and Research,
New York State Department of Health, and School of Public Health,
State University of New York, Albany, New York 12201-0509
Norman Kelker (1), Enzo Biochem, Inc., New York, New York 10013
R. A. Lockington (389), Department of Animal Sciences, Waite Agricul
ture Research Institute, Glen Osmond, South Australia 5064, Aus
tralia
Alberto J. L. Macario (485), Wadsworth Center for Laboratories and
Research, New York State Department of Health, and School of
Public Health, State University of New York, Albany, New York
12201-0509
S. Miller (389), Department of Animal Sciences, Waite Agriculture Re
search Institute, Glen Osmond, South Australia 5064, Australia
Margarita M. Molina (1), Enzo Biochem, Inc., New York, New York
10013
S. Notermans (353), Laboratory of Water and Food Microbiology, Na
tional Institute of Public Health and Environmental Protection, 3720
BA Bilthoven, The Netherlands
Linda M. Parsons (69), Laboratories for Bacteriology, Wadsworth Center
for Laboratories and Research, New York State Department of
Health, Albany, New York 12201-0509
Marjorie Pollice (1), Enzo Biochem, Inc., New York, New York 10013
Rino Rappuoli (205), Sclavo Research Center, 53100 Siena, Italy
Fran A. Rubin (323), Department of Bacterial Immunology, Walter Reed
Army Institute of Research, Washington, D.C. 20307-5100
J. ter Schegget (295), Department of Medical Microbiology, University of
Amsterdam, 1105 AZ Amsterdam, The Netherlands
1 Present address: Laboratoire d'Analyses Ntedicales, 218, rue Mandron, 33000 Bor
deaux, France.
Contributors XV
G. J. Schoone (295), Department of Tropical Hygiene, Royal Tropical
Institute, 1105 AZ Amsterdam, The Netherlands
Jitvimol Seriwatana (95), Armed Forces Research Institute of Medical
Sciences, Bangkok 10400, Thailand
Orntipa Sethabutr (95), Armed Forces Research Institute of Medical Sci
ences, Bangkok 10400, Thailand
Mehdi Shayegani (69), Laboratories for Bacteriology, Wadsworth Center
for Laboratories and Research, New York State Department of
Health, Albany, New York 12201-0509
W. J. Terpstra (295), Department of Tropical Hygiene, Royal Tropical
Institute, 1105 AZ Amsterdam, The Netherlands
John A. Todd (1), Enzo Biochem, Inc., New York, New York 10013
Kevin J. Towner (459), Department of Microbiology and PHLS Labora
tory, University Hospital, Nottingham NG7 2UH, England
Jacob Victor (1), Enzo Biochem, Inc., New York, New York 10013
Alfred L. Waring (69), Laboratories for Bacteriology, Wadsworth Center
for Laboratories and Research, New York State Department of
Health, Albany, New York 12201-0509
K. Wernars (353), Laboratory of Water and Food Microbiology, National
Institute of Public Health and Environmental Protection, 3720 BA
Bilthoven, The Netherlands
Bruce L. Wetherall (255), Department of Clinical Microbiology, Flinders
Medical Center, Bedford Park, South Australia 5042, Australia
Peter H. Williams (143), Department of Genetics, University of Leicester,
Leicester LEI 7RH, England
Huey-Lang Yang (1), Enzo Biochem, Inc., New York, New York 10013
Preface
Molecular genetics has advanced enormously in the past decade. It is
changing our views of physiological and pathological phenomena and,
consequently, our strategies to study and control them. The construction
of gene probes to analyze microbes is one example of these contemporary
trends.
Clinical and epidemiological applications of nucleic acid probes for
bacteria have just begun. This book attempts to furnish a comprehensive
account of the state of the art of this new area. It complements the treatise
on antibacterial monoclonal antibodies published earlier (see Introduc
tion). Both present extensive reviews of probes for bacteria. The earlier
work dealt with immunologic probes and methods; this one covers molec
ular genetics and nucleic acids, emphasizing diagnosis. The preparation
and use of nucleic acid probes for identifying bacteria in clinical specimens
and in other samples of practical or scientific interest are described.
The contributors are pioneers in their fields of expertise. They are
affiliated with academe, government, or industry, thus representing all
major interest groups. Their chapters—particularly their methods, conclu
sions, and suggestions for the future—should be of great use to profes
sionals, technicians, and R&D directors in universities, in federal and
state-dependent service and research institutes, and in private (e.g., hospi
tal) and industrial laboratories.
The probes and procedures described are applicable to a variety of areas
pertinent to medicine, veterinary sciences, environmental (sanitary) engi
neering, agronomy, zoology, and to other branches of science and bio
technology dealing with microbes. These probes and procedures have
been standardized for detecting the presence of bacteria and for identifying
xvii
xviii Preface
and classifying them in clinical specimens, environmental samples, labora
tory cultures, etc. The probes are for pathogens affecting the oral cavity,
nasopharynx, gastrointestinal and genitourinary tracts (including those
causing venereal diseases), eyes, blood and meninges, the cardiovascular,
respiratory, and immune systems, and for bacteria found in water and
foods, tissue cultures, and other ecosystems of importance.
Salient features of the book are highlighted in the Introduction. It calls
the reader's attention to chapter sections covering laboratory applications
and identifies other sections of general interest. To enhance the book's
utility, the Introduction also comprises a bibliographic update.
We would like to thank our teachers, assistants, collaborators, and
students for their continual support and for their help in the many tasks
that directly or indirectly culminated in this book. We would also like to
thank the contributors for their excellent manuscripts and the staff of
Academic Press for expert advice, unfailing support, and encouragement.
Alberto /. L. Macario
Everly Conway de Macario
Introduction: Molecular Genetics in
Diagnostic Bacteriology
Molecular genetics, including genetic engineering, has revolutionized
biology. Genes have been and are being isolated and characterized. Cur
rently, studies on how genes function and are regulated are progressing
rapidly. Techniques for manipulating and modifying genes are becoming
widespread.
These advances have permeated into virtually all branches of biology.
Examining how cells and organisms work can now be done from the
vantage point of molecular genetics. One of the many outcomes of these
innovative ideas and techniques has been the generation of nucleic acid
probes for identifying microbes. Thus, diagnostic microbiology has been
endowed with new means to accomplish its primary goals, namely, to
detect and identify viruses, bacteria, and other microorganisms in samples
of scientific or practical interest.
Diagnostic bacteriology, the central theme of this volume, is entering a
new era marked by the application of gene probes in addition to other
classical identification methods, such as culture-isolation and immunoas
says, based on polyclonal or monoclonal antibodies. This book was con
ceived along the lines of its predecessor "Monoclonal Antibodies against
Bacteria" (Volumes I-III, 1985-1986. A. J. L. Macario and E. Conway de
Macario, editors. Academic Press, Inc.). The main objective was to pro
vide a comprehensive report on the current status of gene probes for
bacteria useful in diagnostics. Which nucleic acid probes are available,
how and when to utilize them, what to expect in terms of results obtained
with their use, and how to prepare new probes are all questions addressed
in the various chapters.
xix
XX Introduction
The contributors were asked to prepare comprehensive manuscripts
covering the microorganisms of their expertise. A few bacterial species are
treated in more than one chapter in order to provide a multidimensional
picture of important microbes and to compile the knowledge gained from
different laboratories using the same or different (e.g., radioactive vs.
nonradioactive) probes or methods. To maintain unity of format and en
sure full coverage, the chapters include a uniform series of sections.
Sections I, "Introduction," and II, "Background," present the main
theme of the chapter, its origins, and pertinent problems and questions. A
brief historical overview is included in Section II, with descriptions of
microorganisms and diseases dealt with in the chapter. The purpose of this
introductory material is to familiarize the nonspecialist with topics that
need to be known to understand the subsequent sections, particularly
Section III, "Results and Discussion," in which the expertise of the
authors is displayed. Data are reported and analyzed to illustrate prepara
tion and/or use of nucleic acid probes. A critical examination of the results
leads to inferences and speculations which are summarized in Section IV,
"Conclusions." The advantages and disadvantages of nucleic acid probes
as opposed to antisera and monoclonal antibodies are discussed in Section
V, "Gene Probes versus Antisera and Monoclonal Antibodies." Here the
bibliography serves as a starting point in the search for additional informa
tion regarding modern serology and immunoassays based on monoclonal
antibodies. As such, Section V may help those unfamiliar with immunoas
says to venture into the advanced field of immunology and immunochemis-
try that originated only a few years ago with the widespread application of
the hybridoma technology.
Section VI, "Prospects for the Future," contains opinions, specula
tions, and suggestions by the authors based on their invaluable experience.
Suggestions relate to a variety of problems (methodological, biological,
pathological, environmental, biotechnological) the authors deem impor
tant for research in the near future.
Section VII is a synopsis of the chapter's content. It leads the reader into
Section VIII, "Materials and Methods," which is very important since it
gives detailed instructions for the preparation and/or utilization of the
nucleic acid probes specifically treated in the chapter. Procedures already
published in manuals available in virtually all laboratories working with
nucleic acids are only briefly mentioned or not described at all, unless
modifications have been introduced. These modifications are given in
detail and represent the authors' contribution to a field burgeoning with
innovations. Some are improvements on the original methods; others are
adaptations required to deal with unique technical difficulties posed by
special characteristics of some microorganisms. Section VIII should be
