Microbial Extracellular Polymeric Substances: Characterization, Structure and Function

265 Pages·1999·11.304 MB·English

by  Jost Wingender, Thomas R. Neu (auth.), Dr. Jost Wingender, Dr. Thomas R. Neu, Prof. Dr. Hans-Curt Flemming (eds.)

Microbial Extracellular Polymeric Substances
Characterization, Structure and Function
Springer
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Heidelberg
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Jost Wingender· Thomas R. Neu· Hans-Curt Flemming
Microbial Extracellular
Polymeric Substances
Characterization, Structure and Function
With 42 Figures
Springer
Editors:
Dr. Jost Wingender
Dept. of Aquatic Microbiology
University of Duisburg and
IWW Center for Water Research
GeibelstraBe 41
D-470S7 Duisburg
Germany
Dr. Thomas R. Neu
Dept. of Inland Water Research Magdeburg
UFZ Centre for Environmental Research
Leipzig-Halle
BriickstraBe 3 a
D-39114 Magdeburg
Germany
Prof. Dr. Hans-Curt Flemming
Dept. of Aquatic Microbiology
University of Duisburg and
IWW Center for Water Research
GeibelstraBe 41
D-470S7 Duisburg
Germany
ISBN -13: 978-3-642-64277-7
Library of Congress Cataloging-in-Publication Data
Microbial extracellular polymeric substances: characterization, structure, and function 1 Jost Wingender,
Thomas R. Neu, Hans-Curt Flemming [editors].
Includes bibliographical references and index.
ISBN -13: 978-3-642-64277-7 e-ISBN -13: 978-3-642-60147-7
DOl: 10.1007/978-3-642-60147-7
1. Microbial polymers. 2. Biofilms. I. Wingender, Jost. II. Neu, Thomas Roo III. Flemming, Hans-Curt
QR92.P59.M53 1999 572'.29-dc21 99-38891
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© Springer-Verlag Berlin Heidelberg 1999
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Preface
The variety of microbial aggregates such as biofilms, flocs and sludge are kept
together by extracellular polymeric substances (EPS). They represent the con
struction material which allows the cells to maintain stable micro consortia and
to establish synergistic relationships. The EPS playa key role for the under
standing of structure, function, properties and development of microbial ag
gregates. The EPS fill and form the space between the biofllin constituents, e.g.,
prokaryotic and eukaryotic microorganisms which inhabit the aggregates. It
seems that the EPS matrix may serve as a multipurpose functional element of
microbial communities, including adhesion, structure, protection, recognition
and physiology. Due to the metabolic activity of the cells, gradients develop and
create different habitats within small distances, allowing a wide variety of or
ganisms to settle and grow in the aggregate. As many of these organisms pro
duce their specific EPS, it is not surprising that an extremely wide variety of
microbial aggregates result. All of them have in common their highly hydrated
extracellular matrix which is the place the organisms shape their very own
microhabitat in which they live.
The paramount role of the EPS has long been neglected and has been ack
nowledged only recently. In the first chapter, an attempt is made to give a de
finition and an overview of what EPS represents, of what they are composed,
what their function is, and which role they play in microbial ecology. A large
part of the book is devoted to the isolation and characterization of EPS from
different systems. The matrix does not seem to be built randomly by some bio
polymers but displays functional aspects. This is considered as well as the par
ticular aspect of cell-cell-communication which is of special interest in aggre
gates where the cells maintain high densities and close proximities.
Extracellular enzymes and polysaccharides can interact and this, supports the
view that the EPS matrix must be probably recognized as a key factor in a mo
del which considers biofllms and other microbial aggregates as an early form of
multicellular organisms.
The purpose of this book is to present information and concepts with regard
to current state of knowledge about EPS and, consequently bring the attention
of the readers to the material between the cells, which may be of more impor
tance than previously assumed. If it inspires the further understanding of the
aggregated way of microbial life, it has fulfilled its purpose.
August 1999 Jost Wingender
Thomas R. Neu
Hans-Curt Flemming
Contents
What are Bacterial Extracellular Polymeric Substances?
Jost Wingender, Thomas R. Neu, Hans-Curt Flemming 1
1 Introduction .................... . 1
2 Definitions of EPS ................. . 2
3 Composition, Secretion and Spatial Arrangement of EPS 4
4 Industrial and Clinical Importance of EPS 9
5 Bacterial Alginate - an Example of Bacterial EPS 10
6 Functions of EPS . . . . . 11
7 Ecological Aspects of EPS 13
References ....... . 15
In Situ Characterization of Extracellular Polymeric Substances (EPS)
in Biofilm Systems
Thomas R. Neu, John R. Lawrence 21
1 Introduction ....... . 22
1.1 Traditional Approaches for Studying Microbial Polysaccharides 22
l.2 Change of Paradigms .. . 22
1.3 In situ Methods ..... . 23
2 Destructive Analysis of EPS 23
2.1 Chemical Techniques 23
2.1.1 Extraction .... . 23
2.1.2 Analysis ....... . 24
2.2 Electron Microscopy . 24
2.3 Scanning Probe Techniques 26
3 Non-Destructive Analysis of EPS 27
3.1 Infrared Spectroscopy (FT-IR) . 27
3.2 Magnetic Resonance Spectroscopy (NMR) 28
3.3 Confocal Laser Scanning Microscopy (CLSM) 29
4 CLSM Approach ..... . 29
4.1 Probes for Analysis of EPS 29
4.1.1 Polysaccharides 30
4.l.2 Proteins .......... . 31
4.1.3 Nucleic Acids . . . . . . . . 32
4.2 Physicochemical Characterization of EPS Matrix 32
VIII Contents
4.2.1 Charge Distribution ...... . 32
4.2.2 Hydrophobicity/Hydrophilicity . 32
4.2.3 Permeability .......... . 33
4.3 EPS Bound and Associated Molecules 33
5 Image Analyses and Three-Dimensional Data Presentation 33
5.1 Analyses Techniques . . . . . . 34
5.2 Three-Dimensional Imaging ........ . 35
5.3 Three-Dimensional Display . . . . . . . . . . 35
6 Case Studies: In Situ Characterization of EPS 36
7 Limitations of Non-Destructive Analyses 40
7.1 FT-IR/ATR . 40
7.2 NMR 41
7.3 CLSM ... 41
References 42
Extraction of EPS
Per H. Nielsen, Andreas Jahn 49
1 Introduction. . . . 50
1.1 Definition of EPS . . 50
1.2 Composition of EPS 51
1.3 Extraction Procedure 52
2 Sampling and Pretreatment 54
2.1 Sampling and Storage 54
2.2 Homogenization. 54
2.3 Washing........ 54
3 Extraction of EPS 54
3.1 Selection of an Appropriate Extraction Method 55
3.2 Physical Methods ................ . 55
3.3 Chemical Methods . . . . . . . . . . . . . . . . . 58
3.4 Combination of Physical and Chemical Methods 61
4 Contamination by Intracellular Macromolecules 61
5 Disruption of Macromolecules . 64
6 Extraction Efficiency 65
7 Purification and Analysis 68
8 Conclusion 68
References ..... . 69
Biofilm Exopolysaccharides
Ian W. Sutherland . 73
1 Introduction 73
2 Polysaccharide Composition and Structure 76
3 Promotion of Order and Gelation 81
4 Ionic Interactions 83
5 Synergistic Effects ........ . 83
Contents IX
6 Other Functions of Biofilm Exopolysaccharides 84
7 Dissolution of Physical Structures 87
8 Conclusions 88
References 89
Regulation of Matrix Polymer in Biofilm Formation and Dispersion
David G. Davies ....................... 93
Alginate Biosynthesis in Pseudomonas aeruginosa 93
2 Regulation of Alginate Biosynthetic Genes During Biofilm
Development ... . . . . . . . . . . . . . . . . . . Ibo
3 Control of Biofilm Dispersion .......... . 104
4 Activation of Alginate Biosynthesis and Chemical
Communication in BiofIlms . . . 106
5 P. aeruginosa BiofIlm Life Cycle 110
References ........... . 112
Exopolymers of Sulphate-Reducing Bacteria
Iwona B. Beech, Rudi C. Tapper ....... . ....... 119
Analysis and Function of the EPS from the Strong Acidophile
Thiobacillus ferrooxidans
Wolfgang Sand, Tilman Gehrke 127
1 Introduction ...... . 127
2 Leaching Bacteria 129
3 Metal Sulfide Dissolution 130
4 EPS Analysis .. 132
5 Function of EPS 135
6 Conclusions 138
References 140
Physical and Chemical Properties of Extracellular Polysaccharides
Associated with Biofilms and Related Systems
Bj0rn E. Christensen 143
1 Introduction . 143
2 Biopolymer Gels 145
3 Physical Characterization of BiofIlms 146
4 Chemical and Biochemical Characterization of BiofIlms 147
5 Bacterial Alginates 149
References ........................ . 153
x
Contents
Chemical Communication Within Microbial Biofilms:
Chemotaxis and Quorum Sensing in Bacterial Cells
Alan W. Decho . . . 155
1 Introduction 155
2 Bacterial Chemotaxis and Initial Aggregations of Cells 156
3 Microspatial Patterns and Movement of Bacteria Within Biofllms 157
4 Bacterial Quorum Sensing: Induction and Regulation of Activities 158
4.1 Model Systems ................ 159
4.1.1 Homoserine Lactone (Autoinducer) Signals 159
4.1.2 The Vibrio fischeri System. . . . . . . . . . 160
4.2 Other Chemical Signals ........... 160
4.2.1 Conjugation and Pheromone-Inducible Gene Expression in
Enterococcus faecalis . . . . . . . . . . 160
4.2.2 Cyanobacteria Systems ........ 161
4.3 Functional Roles of Quorum Sensing 162
4.4 Specificity of Autoinduction Process . 162
5 Enhancement of Chemotaxis and Quorum Sensing Processes
by the Biofllm Matrix ......... 162
6 The Biofllm as a Microbial Organism 164
References ............... 165
Function of EPS
Gideon M. Wolfaardt, John R. Lawrence, Darren R. Korber 171
1 Introduction. . . . . . . . . . . . . 172
2 Role of EPS in Cellular Associations 172
2.1 Suspended and Attached Cells 172
2.2 Co aggregation, Consortial Behavior, and Floc Formation 173
2.3 Biofllm Formation . . . 174
2.4 Genetic Transfer .............. 175
3 Role of EPS in Nutrition . . . . . . . . . . 177
3.1 Production of EPS as a Nutrient Reserve 178
3.2 Accumulation of Nutrients by EPS . . . . 178
4 Role of EPS in the Interaction of Microorganisms
with their Biological, Physical, and Chemical Environment 181
4.1 EPS and the Micro-Environment . . . . . . . . . 181
4.2 EPS as a Physical Barrier to Solute Translocation . . . . . . 183
4.3 Role in Enzyme Reactions. . . . . . . . . . . . . . . . . . . 185
4.4 Role in Pathogenicity and Protection Against Host Defense
Mechanisms .................. 186
4.5 Protective Role Against Predation/Digestion 186
4.6 Microbial-Plant Associations .... 188
4.7 Effect of Environmental Conditions 189
5 EPS and the Macro-Environment 190
5.1 Stabilization of the Environment . . 190
Contents XI
5.2 Microbial-Mineral Interactions ............. . 191
5.3 Controls on Flow . . . . . . . . . . . . . . . . . . . . . . 192
5.4 Role of EPS in the Bio-Accumulation of Contaminants 193
6 Conclusions 194
References 195
Polysaccharases in Biofilms - Sources - Action - Consequences!
Ian W. Sutherland . . . . . . . . . 201
1 Polysaccharases in Biofilms 201
2 Nature of Polysaccharases . 202
3 Sources of Polysaccharases 202
3.1 The Endogenous Production of Polysaccharases 202
3.2 The Exogenous Production of Polysaccharases 205
3.3 Bacteriophage ........... . 205
4 Effects of Enzymes on Biofilms . . . 210
5 Application of Enzymes to Biofilms 212
References ............. . 213
Extracellular Enzymes Within Microbial Biofilms
and the Role of the Extracellular Polymer Matrix
Monica Hoffman, Alan W. Decho ......... . .. 217
Introduction: Importance of Extracellular Enzymes to Bacterial
Cells and Organic Matter Processing . . . . . . . . . . . . . . .. 217
2 The Microbial Biofilm and Extracellular Polymers . . . . . . .. 219
3 Extracellular Polymers as a "Sorptive Sponge" and "Colloidal Trap"
for High-Molecular-Weight Organic Matter . . . . . 220
4 Localization of Enzymes by Extracellular Polymers:
the "Lectin-Localization Model" ........... 221
5 "Extracellular Polymer Microdomains" and the Stability
of Extracellular Enzymes .................. 223
6 Environmental Influences on Enzyme Activity . . . . . . 225
7 Biofilm Induction and Regulation of Extracellular Enzymes 225
7.1 Induction of Extracellular Enzymes at the Physiological Level 226
7.2 Regulation of Enzyme Activity by Extracellular Factors 226
References ............................. 227
Interaction Between Extracellular Polysaccharides and Enzymes
Jost Wingender, Karl-Erich Jaeger, Hans-Curt Flemming 231
1 Extracellular Enzymes in Biofilms . . . . . . . . . 231
2 Mechanisms of Enzyme and Polysaccharide Secretion 235
2.1 Secretion of Enzymes 235
2.1.1 The ABC Pathway ................... . 236