Molecular Identification, Systematics, and Population Structure of Prokaryotes (eBook)

Title Page 2
Copyright Page 3
Preface 4
Table of Contents 6
Contributors 11
1 Exciting Times: The Challenge to be a Bacterial Systematist 13
1.1 Introduction 13
1.2 The Early Heroes (1860-1900) 15
1.3 The Dawn of Microbial Ecology and the Continuing Struggle with Classification Systems (1900-1930) 17
1.4 Encouragement and Frustration (The Era 1930-1950) 19
1.5 Expanding the Range of Properties: The Genetic and Epigenetic Levels (1950-1980) 22
1.6 Yet Another Exciting Time: Unravelling the Genealogy(ies) of Cultured and As-Yet Uncultured Prokaryotes 25
References 28
2 DNA-DNA Reassociation Methods Applied to Microbial Taxonomy and Their Critical Evaluation 34
2.1 Introduction 34
2.2 Semantic Considerations 37
2.3 DNA-DNA Reassociation Measurement, Parameters and Methods 40
2.4 Interpretation of Results and the Boundaries for Species Circumscription 50
2.5 The Impact of DNA-DNA Hybridizations on the Conception of a Species and Changes in the Concept and/or the Definition 53
2.6 Epilogue 55
References 57
3 DNA Fingerprinting Techniques Applied to the Identification, Taxonomy and Community Analysis of Prokaryotes 62
3.1 Introduction 62
3.2 DNA Typing Methods 64
3.2.1 DNA Typing Methods Targeting the Whole Genome of a Bacterial Strain 64
Pulsed Field Gel Electrophoresis for Macro-restriction of Total DNA 64
Random Amplified Polymorphic DNA Assay 65
Amplified Fragment Length Polymorphism Analysis 67
Amplification of Repetitive Elements Dispersed through the Whole Genome 68
3.2.2 DNA Typing Methods Targeting Gene Clusters (Operons) 71
RFLP Analysis with Southern Blotting and Probe Hybridization (Ribotyping) 71
Analysis of the 165-235 Ribosomal Intergenic Spacer Region 73
3.2.3 DNA Typing Methods Targeting the 16S rRNA Gene 75
Amplified Ribosomal rDNA Restriction Analysis 75
Terminal Restriction Fragment Length Polymorphism Analysis 76
Denaturing Gradient Electrophoresis and Temperature Gradient Gel Electrophoresis 77
PCR-based Single-stranded Confirmation Polymorphism 80
References 82
4 Multiple Locus VNTR (Variable Number of Tandem Repeat) Analysis 94
4.1 Introduction 94
4.2 MLVA Origins 94
4.3 MLVA Set-up and Enrichment 95
4.3.1 Evaluation of the Potential Interest of MLVA for a Given Species 96
4.3.2 MLVA Validation 97
4.3.3 Data Management 98
4.4 Existing First-generation MLVA Assays 99
4.4.1 Mycobacterium tuberculosis 102
4.4.2 Bacillus anthracis 104
4.4.3 Yersinia pestis 105
4.4.4 Brucella sp. 106
4.4.5 Legionella pneumophila 108
4.4.6 Other Bacteria 108
4.5 Validating and Analysing MLVA Data 108
4.6 MLVA Compared to Other Methods 111
References 112
5 Bacterial Phylogeny Reconstruction from Molecular Sequences 116
5.1 Introduction 116
5.2 Species Definition 117
5.3 Bacterial Diversity 119
5.4 Phylogenetic Analysis Based on 16S rDNA Sequences 121
5.5 Phylogenetic Analysis Based on Protein Sequences 126
5.5.1 Selection of Target Proteins 126
5.5.2 Design of PCR Primers for the Amplification of Protein-encoding Genes: A Case Study with gyrB 132
5.6 Limitations in Reconstructing Phylogenetic Trees 137
5.7 Conclusion and Future Perspective 140
References 142
6 Integrated Databasing and Analysis 151
6.1 Introduction 151
6.2 Classes of Data 152
6.3 Character Type Data 153
6.3.1 Definition 153
Open and Closed Data Sets 153
Binary, Numerical, and Categorical Data 154
6.3.2 Data Transformation 155
Standardization 155
A. Rows (Entries) 157
B. Columns (Characters) 157
Dealing with Missing Values 158
6.3.3 Cluster Analysis of Character Type Data 159
6.4 Fingerprint Type Data 159
6.4.1 Definition 159
6.4.2 Preprocessing of Fingerprint Data 160
Step 1. Import and Preprocessing of the Gel Image 160
Step 2. Extracting Densitometric Curves from the Gel Image Lanes 164
Step 3. Normalization 165
Step 4. Detection of Bands/Peaks 169
6.4.3 Comparison of Fingerprint Data 171
Comparison of Densitometric Curves 172
Comparison of Band/Peak Positions 174
Dealing with Uncertain Bands 177
Optimization of Pattern Alignment 177
Choosing the Most Appropriate Coefficient 178
6.4.4 Fingerprint Techniques That Require Special Analysis Methods 182
Variable Number Tandem Repeats 182
6.5 Sequence Type Data 184
6.5.1 Definition 184
6.5.2 Assembling Sequencer Trace Files into Consensus Sequences 184
6.5.3 Alignment of Sequences 185
6.5.4 Multiple Alignment 189
6.5.5 Phylogenetic Clustering 190
6.5.6 Multi-locus Sequence Typing 190
Analysis of MLST Data 192
6.6 Matrix Type Data 193
6.7 Trend Type Data 194
6.8 Two-dimensional Gel Type Data 196
6.8.1 Analyzing 2D Gels 198
6.9 The Integrated Database 199
6.9.1 Distributed Databases and Portability of Data 199
6.10 Hierarchical Cluster Analysis 202
6.10.1 Similarity- or Distance-based Clustering Techniques 202
UPGMA and Related Clustering Algorithms 204
Neighbor Joining Technique 206
6.10.2 Phylogenetic Clustering Methods 208
6.10.3 Minimum Spanning Trees 208
MSTs and Population Genetics 210
6.11 Consensus Grouping and Classification 213
6.11.1 Concatenation of Data Sets 215
6.11.2 Averaging Resemblance Matrices 215
Harmonization of Distance Matrices 216
6.11.3 Consensus Trees 218
6.12 Error on Dendrograms 218
6.12.1 Degeneracy of Dendrograms 220
6.12.2 Dealing with Dendrogram Degeneracies 222
References 224
7 Assessment of Microbial Phylogenetic Diversity Based on Environmental Nucleic Acids 228
7.1 Introduction 228
7.2 Microbial Phylogenetics and the 16S rRNA Gene 229
7.3 16S rRNA and the Environment 231
7.4 Molecular Methodology in Microbial Ecology 233
7.5 General Considerations of Bias 237
7.6 Phylogenetic Assessment of Environmental Nucleic Acids 241
7.7 Fingerprinting 242
7.7.1 Denaturing Gradient Gel Electrophoresis 243
7.7.2 Temperature Gradient Gel Electrophoresis 244
7.7.3 Single-stranded Conformational Polymorphism 245
7.7.4 Terminal Restriction Fragment Length Polymorphism 245
7.7.5 Ribosomal lntergenic Spacer Analysis 246
7.7.6 Additional Considerations 247
7.8 Sequencing 248
7.8.1 16S rRNA Gene Libraries 248
7.8.2 Serial Analysis of Ribosomal Sequence Tags 250
7.9 Metagenomics 251
7.10 Array Technology 252
7.11 Composite Methodologies 254
7.12 Conclusion 255
References 256
8 Metagenome Analyses 269
8.1 Introduction 269
8.2 Construction and Screening of Metagenome Libraries 272
8.2.1 Small and Large Insert Libraries 273
8.2.2 High-capacity Vectors: Cosmids, Fosmids or BACs? 273
8.2.3 Library Size 275
8.2.4 Isolation and Purification of HMW DNA 276
8.2.5 Construction of Large Insert Metagenomic Libraries 277
8.2.6 Storage of Metagenomic Libraries 278
8.2.7 Screening of Metagenomic Libraries 279
8.2.8 Sequencing of Large Insert Constructs 280
8.3 Sequence Analysis 281
8.3.1 Marker Genes 281
8.3.2 End-Sequences 283
8.3.3 Cosmids, Fosmids or BACs 284
Correlation of Metagenomic Fragments 284
Functional Annotation 285
8.4 Summary, Pitfalls and Outlook 288
References 289
9 DNA Microarrays for Bacterial Genotyping 295
9.1 Introduction 295
9.2 Technical Principles 296
9.3 Applications 298
9.3.1 Comparative Genome Hybridization 298
9.3.2 Diagnostic Detection of Virulence Genes 303
9.3.3 Diagnostic Detection of Resistance Determinants 304
Detection of Resistance Genes 304
Analysis of Resistance-mediating Point Mutations 305
9.3.4 Multi-locus Sequence Typing by Hybridization 306
9.3.5 Composite Gene Detection for Epidemiological Typing 307
9.3.6 Detection of Genes Associated with Metabolic Functions 309
9.3.7 Phylogenetic Identification 311
9.3.8 Random Hybridization Fingerprinting 312
9.4 Present Limitations and Future Prospects 313
References 314
Subject Index 322