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Laboratory techniques in plant bacteriology
Machine generated contents note: ch. 1 Laboratory Ethics in Plant Bacteriology Laboratory: Instruments and General Guidelines -- 1.1.Ethics in the Laboratory -- 1.1.1.Cleanliness -- 1.1.2.Silence -- 1.1.3.Dress Code -- 1.1.4.Storage Cabinets -- 1.1.5.Maintenance of Laboratory Hygiene -- 1.2.Laboratory Instruments -- 1.2.1.Maintenance of Instruments -- 1.2.2.Basic Instruments -- 1.2.3.Applied Instruments -- 1.3.Necessary Miscellaneous Instruments -- 1.4.General Guidelines -- ch. 2 Confirmation of Bacterial Pathogen in Diseased Samples -- 2.1.By Ooze Test -- 2.2.By Milky Water Test -- 2.3.By String Test -- 2.4.By Staining of Crushed Disease Portion -- ch. 3 Histopathology of Bacterial Infection -- 3.1.Determination of the Location of Bacterial Pathogens in Host -- 3.2.Preparation Of Microtome Sections and Staining for Histological Studies -- 3.3.Study of Changes in Chemical Constituents of the Affected Plant Tissues -- 3.3.1.Degradation in Cellulose --
Note continued: 3.3.2.Degradation of Poetic Substances -- 3.3.3.Degradation of Protein -- 3.3.4.Degradation of Lignin -- ch. 4 Sterilization -- 4.1.Sterilization of Glassware -- 4.2.Sterilization of Medium -- 4.2.1.By Autoclave -- 4.2.2.By Filtration -- 4.3.Sterilization of Soil -- 4.3.1.Sterilization in Steam without Pressure -- 4.3.2.Sterilization with Chemical Vapors -- 4.3.3.Sterilization with Solar Heat -- 4.4.Sterilization of the Working Place -- 4.4.1.By UV Radiation -- ch. 5 Medium for Isolation of Plant Pathogenic Bacteria -- 5.1.Basic Liquid Media (Broth) for the Routine Cultivation of Bacteria -- 5.2.General Medium for Routine Cultivation of Bacteria -- 5.2.1.Preparation of Nutrient Sucrose Agar (NSA) Media -- 5.3.Selective and Differential Media -- 5.3.1.Special Medium -- 5.3.1.1.Special Media for Xanthomonas -- 5.3.1.2.Special Medium for Pseudomonas fluorescens (King's B Medium) -- 5.3.1.3.Special Medium for Isolation of Pectolylic Erwinia --
Note continued: 5.3.1.4.Special Medium for Agrobacterium -- 5.3.1.5.Yeast Glucose Chalk-Agar Medium for Maintenance of Bacterial Cultures -- 5.4.Preparation of Slants -- ch. 6 Isolation of Bacterial Plant Pathogens -- 6.1.From Diseased Plant Sample -- 6.2.From Disease Field Soil Sample -- 6.3.From Field Water Sample -- 6.4.From Infected Seed Material -- 6.4.1.Isolation of Externally Seedborne Bacteria -- 6.4.2.Isolation of Internally Seedborne Bacteria -- ch. 7 Purification of Plant Pathogenic Bacterial Cultures -- 7.1.Selection of Single Bacterial Colonies -- 7.2.Picking of Single Colonies and Subculture -- 7.3.Checking the Purity of the Isolated Culture -- ch. 8 Rapid Assessment of Plant Pathogenic Nature of Bacterial Isolates -- 8.1.By Hypersensitive Reaction on a Tobacco Plant -- 8.2.By Hypersensitive Reaction on a Non-Host Plant -- 8.3.By Reaction of Bacteria on Soft Fruits --
Note continued: ch. 9 Determination of Pathogenicity of the Isolated Bacterial Culture on Natural Host Plants -- 9.1.By Infiltration Method -- 9.2.By Swab Inoculation Method -- 9.3.By Spray Inoculation Method -- 9.4.By Root Dip Inoculation Method -- 9.5.By Vacuum Infiltration Method -- 9.6.By Pin Prick Inoculation Method -- ch. 10 Isolation and Enumeration of Microorganisms Associated with Bacterial Plant Pathogen from Soil, Rhizosphere, and Phylloplane -- 10.1.Isolation of Microorganisms from Cultivated Soil/Soils under Cropping Systems -- 10.2.Isolation of Rhizosphere Microflora -- 10.3.Isolation of Microorganism from Root Surface -- 10.4.Isolation of Microorganisms from Virgin/Uncultivated/Non-Rhizosphere Soil -- 10.5.Isolation of Phylloplane Microflora by the Serial Dilution Method -- 10.6.Study of Phylloplane Microflora by Leaf Imprint Method -- ch. 11 Preparation of Pure Cultures of Microorganisms -- 11.1.By Streak Plate Method -- 11.2.By Pour Plate Method --
Note continued: 11.3.By Spread Plate Technique -- ch. 12 Maintenance and Preservation of Bacterial Pure Cultures -- 12.1.Preservation on YGCA Media -- 12.2.Preservation in Refrigerator or Cold Room Storage -- 12.3.Preservation in Mineral Oil -- 12.4.Preservation at -40°C in Glycerol -- 12.5.Preservation in Edible/Nonedible Oils -- 12.6.Preservation by Liquid Nitrogen Method (Storage at Low Temperature) -- 12.7.Preservation by Paraffin Method -- 12.8.Preservation by Freeze-Drying (Lyophilization) Method -- ch. 13 Staining of Bacterial Cultures for Morphological Studies -- 13.1.Preparation of Bacterial Smears -- 13.2.Simple or Direct Staining of Bacteria -- 13.3.Negative Staining of Bacteria -- 13.4.Gram Staining of Bacteria -- 13.5.Acid-Fast Staining of Bacteria -- 13.6.Bacterial Cell Wall Staining -- 13.7.Cytoplasmic Membrane Staining -- 13.8.Bacterial Spore (Endospore) Staining -- 13.9.Capsule Staining -- 13.10.Flagella Staining --
Note continued: 13.11.Staining of Nuclear Material of Bacteria -- 13.12.Viability Staining of Bacteria -- 13.13.Suslow Reaction to Determine Gram-Positive and Gram-Negative Bacteria -- ch. 14 Bacterial Mobility -- 14.1.Hanging Drop Technique for Demonstrating Motility of Bacteria -- 14.2.Brownian Movement -- 14.3.Relationship of Virulence with Motility (in Erwinia carotovora var. zeae) -- ch. 15 Describing Bacterial Colony Morphology -- 15.1.Colony Type -- 15.2.Colony Elevation -- 15.3.Colony Edge -- 15.4.Internal Colony Structure ([×] 15) -- 15.5.Colony Pigments -- ch. 16 Physiological Characterization of Bacteria (Effect or Environmental Conditions on Growth of Bacteria) -- 16.1.Effect of Incubation Temperatures on Bacterial Growth -- 16.2.Determination of Thermal Death Point of Bacteria (TDP) -- 16.3.Determination of Thermal Death Time of Bacteria -- 16.4.Effect or pH on the Bacterial Growth -- 16.5.Effect of Salt Concentration on Bacterial Growth --
Note continued: 16.6.Effect of Metals on Bacterial Growth -- 16.6.1.The Oligodynamic Action of Metals (Aluminum, Copper, and Lead) on Bacteria -- 16.7.Effect of Dyes on Bacterial Growth -- 16.8.Effect of Different Wavelengths of Light on Bacterial Growth -- 16.9.Lethal Effects of Ultraviolet Radiation (UV Rays) on Bacterial Growth -- 16.10.Effect of Relative Humidity on Bacterial Growth -- ch. 17 Biochemical Tests Used in Identification of Bacteria -- 17.1.Amylase Production Test (or Demonstration of Starch Hydrolysis) -- 17.2.Cellulase Production Test (Degradation of Cellulose) -- 17.3.Production of Pectolytic Enzymes (Degradation of Pectin) -- 17.4.Hydrolysis of Gelatin, a Protein (Production of Gelatinase) -- 17.5.Casein Hydrolysis -- 17.6.Urease Test -- 17.7.Hydrogen Sulfide Production Test -- 17.8.Carbohydrate Catabolism by Microorganisms (Oxidation and Fermentation of Glucose) --
Note continued: 17.8.1.Grouping of Bacterial Isolates of Xanthomonas malvacearum on the Basis of Lactose Utilization -- 17.9.Fermentation of Carbohydrates -- 17.10.Microbial Reaction in Litmus Milk -- 17.11.Nitrate Reduction -- 17.12.Oxidase Test -- 17.13.Pigment Production -- 17.14.The IMViC Tests -- 17.14.1.Indole Production Test -- 17.14.2.Methyl-Red and Voges -- Proskauer Tests -- 17.14.3.Citrate Utilization Test -- 17.15.Catalase Test -- ch. 18 Characterization of Phytopathogenic Bacteria up to Genus -- 18.1.Agrobacterium -- 18.2.Pseudomonas -- 18.3.Ralstonia -- 18.4.Xanthomonas -- 18.5.Xylella -- 18.6.Xylophilus -- 18.7.Erwinia -- 18.8.Pantoea -- 18.9.Clavibacter (Corynebacterium) -- 18.10.Curtobacterium -- ch. 19 Differentiation of Bacterial Genus into Group -- 19.1.Differentiation of Genus Xanthomonas into Groups -- 19.2.Differentiation of Some Well-Studied Species of the Genus Pseudomonas into Group/Species and Biovars --
Note continued: 19.2.1.Characteristics Useful for Differentiation of Various Denitrifying Pseudomonas -- 19.2.2.Sources and Characteristics of Additional Pseudomonas Species (Isolated from Diseased Plants and Mushrooms) -- 19.3.Differentiation of Genus Erwinia into Groups -- 19.3.1.Differentiation of Erwinia Species by Cultural, Physiological, and Biochemical Characteristics -- 19.3.2.Differentiation of Erwinia Species Based on Acid Production from Organic Compounds -- 19.3.3.Differentiation of Erwinia Species Based on Utilization of Some Organic Compounds as a Source of Carbon and Energy -- 19.4.Differentiation of Genus Agrobacterium into Groups and Biovars -- ch. 20 Differentiation of Bacterial Group into Species -- 20.1.Differentiation of Genus Xanthomonas into Species Based on Morphological and Biochemical Characteristics -- 20.2.Differentiation of Genus Pseudomonas into Species on the Basis of Biochemical Tests --
Note continued: 20.3.Differentiation of Genus Erwinia into Species on the Basis of Biochemical Tests -- 20.4.Differentiation of Genus Pantoea into Species on the Basis of Biochemical Tests -- 20.5.Differentiation of Genus Clavibacter (Corynebacterium) into Species on the Basis of Tests -- 20.6.Differentiation of Agrobacterium tumefaciens into Biotype on the Basis of Different Tests -- ch. 21 Identification of Races of Bacterial Plant Pathogen -- 21.1.Identification of Races of X. campestris pv. campestris (Xcc) -- 21.2.Identification of Races of X. axonopodis pv. malvaceraum -- 21.3.Identification of Races of X axonopodis pv. viticola -- 21.4.Identification of Races of X. oryzae pv. oryzae -- 21.5.Identification of Races of Ralstonia solancearum -- ch. 22 Studies on Bacterial Cell Wall-Related Biological Compounds -- 22.1.Isolation of Bacterial Cell Wall (Peptidoglycan) --
Note continued: 22.1.1.Isolation and Preparation of Bacterial Cell Wall for Compositional Analysis by Ultra Performance Liquid Chromatography (UPLC) -- 22.2.Isolation of Cell Wall -- Less (L-Form) Bacteria -- 22.3.Isolation of Bacterial Exopolysaccharide (EPS) -- 22.3.1.Testing of EPS by Iodine Test -- 22.3.2.Estimation of EPS -- 22.3.3.Estimation of Sugar in EPS -- 22.3.4.Estimation of Protein in EPS -- 22.4.Isolation of Bacterial Lipid -- 22.4.1.Estimation of Bacterial Lipid -- 22.5.Determination of Glycoprotein Secretion Factor -- 22.6.Testing of EPS/GP for Induction of WS Reaction -- ch. 23 Isolation of Bacterial Metabolites -- 23.1.Extraction of Total Cultural Filtrate -- 23.2.Isolation of Bacterial Toxins -- 23.2.1.Non-Host-Specific Toxins -- 23.2.1.1.Tabtoxin -- 23.2.1.2.Phaseolotoxin -- 23.3.Isolation of Bacterial Enzymes -- 23.3.1.Isolation of Cellulases -- 23.3.1.1.Extraction of Cellulase from Pathogen (In Vitro) --
Note continued: 23.3.1.2.Extraction of Cellulase Enzyme from Diseased Plants (In Vivo) -- 23.3.1.3.Measurement of Endoglucanase -- 23.3.2.Pectinases or Pectolytic Enzymes of Pathogens -- 23.3.2.1.Extraction of Pectolytic Enzymes from Diseased Plants (In Vivo) -- 23.3.2.2.Extraction of Pectolytic Enzymes from a Pathogen (In Vitro) -- 23.3.2.3.Assaying ofPolygalacluronase -- 23.3.2.4.Assaying of Pectin Transliminases (PTE, PATE) -- 23.3.3.Assaying Pectolytic Activity of Erwinia carotovora -- 23.3.4.Determination of Pectic Enzyme Production by Xanthomonas malvacearum -- 23.4.Isolation of Bacteriocin -- 23.4.1.Bacteriocin Production -- 23.4.2.Chemical and Physical Stability of Bacteriocin -- 23.4.3.Induction of Bacteriocin Production by Mitomycin C -- 23.4.4.Detection of Antimicrobial Activity in Bacteriocin -- 23.5.Isolation of Siderophore -- 23.5.1.Siderophore Detection Assay --
Note continued: 23.5.2.Effect of Iron Concentration, Sugars, Organic and Amino Acids, and Nitrogen on Siderophores Production -- 23.5.3.Characterization of Siderophores -- ch. 24 Role of Bacterial Component in Induction of Disease Reaction or Pathogenesis -- 24.1.EPS in Induction of Disease Reaction -- 24.2.Bacterial Enzymes in Induction of Disease Reaction -- 24.2.1.Studies with Pectolytic Enzymes of Erwinia -- 24.3.Bacterial Toxins in Induction of Disease Reaction -- 24.3.1.Studies with Phaseolotoxin of Pseudomonas -- ch. 25 Enumeration of Bacterial Population -- 25.1.Enumeration (Counting) of Bacteria by Plate Count or Serial Dilution Agar Plate Technique -- 25.2.Counting of Bacterial Population by the Use of Spectrophotometer -- ch. 26 Determination of Ice-Nucleation in Plant Pathogenic Bacteria -- 26.1.Assessment of Ice Nucleation Property of Pseudomonas syringae -- ch. 27 Transmission of Plant Pathogenic Bacteria through Vector --
Note continued: 27.1.Transmission of Bacteria (Xanthomonas) through Lepidopteron Larvae (Spotted Bollworms) -- 27.2.Transmission of Bacteria (Xanthomonas) through Insect/Beetle (Red Cotton Bug) -- 27.3.Transmission of Erwinia amylovora through Bees -- 27.4.Transmission of Erwinia through Spotted Beetles -- 27.5.Transmission of Erwinia through Nematodes -- ch. 28 Determination of Perpetuation of Bacterial Plant Pathogen -- 28.1.In Soil -- 28.2.In Planting Material -- 28.3.In Seed Material -- 28.4.In Vector -- 28.5.On Implement -- ch. 29 Determination of Plant Resistance -- 29.1.Determination of Vertical Resistance -- 29.2.Determination of Horizontal Resistance -- 29.3.Determination of Horizontal Resistance by Using Single Race -- ch. 30 Identification of Bacteria by Using the Immunodiagnostic Technique -- 30.1.Preparation of Bacterial Antigens or Antigen Glycoprotein from BacteriaPathogens -- 30.2.Immunization and Preparation of Antibodies --
Note continued: 30.2.1.Effect of Immunization Period on Recovery of Immunoglobulin -- 30.2.2.Effect of Extraction Method on Recovery and Affinity of Immunoglobulin -- 30.2.3.Estimation of Effective Immunoglobulin -- 30.2.4.Estimation of Affinity of Immunoglobulin -- 30.3.Production and Characterization of Monoclonal and Polyclonal Antibodies (Evaluation of Specificity) -- 30.3.1.Validity of pAb-Xcc in Detection of the Pathogens in Seed Samples and Infected Plant Materials -- 30.4.Immunodiagnostic Techniques (Antigen -- Antibodies Reaction) -- 30.4.1.Precipitation Test -- 30.4.2.Gel Diffusion Test -- 30.4.3.Agglutination Test -- 30.4.4.Immunological Test with Markers -- 30.4.5.Immunological Electron Microscopy -- 30.4.6.Enzyme Linked Immunosorbent Assay (ELISA) -- 30.4.7.Detection of Xanthomonas campestris pv. Campestris through ELISA -- 30.4.8.Determination of the Antigenic Determinant in Bacterial Pathogen by Immunoblot Assay --
Note continued: 30.5.Use of Specific Immunoglobulin under Immunofluorescence to Detect the Latent Infection -- ch. 31 Identification of Bacteria by Using Molecular Techniques -- 31.1.Isolation of Bacterial DNA -- 31.1.1.Isolation of Bacterial Genomic DNA -- 31.1.2.Isolation of Bacterial Plasmid DNA -- 31.2.Plasmid Profiling Based Techniques -- 31.2.1.Plasmid DNA Restriction Analysis -- 31.2.2.Replicon Typing -- 31.3.Hybridization-Based Techniques -- 31.3.1.Southern Blotting Technique -- 31.3.2.Western Blotting Technique -- 31.3.3.Fluorescent In Situ Hybridization (FISH) -- 31.3.4.Microarray Technology -- 31.4.PCR Based Technique -- 31.4.1.PCR Technology -- 31.4.2.Primer Identification/Designing -- 31.4.3.Arbitrary/Random Primers Based PCR Methods -- 31.4.3.1.Randomly Amplified Polymorphic DNA Markers (RAPD) -- 31.4.3.2.Inter Simple Sequence Repeats Markers (ISSR) -- 31.4.4.Sequence Specific Primers Based PCR Methods --
Note continued: 31.4.4.1.Simple Sequence Repeats (SSR) or Microsatellite Markers -- 31.4.4.2.Single Nucleotide Polymorphism (SNP) -- 31.4.5.Modified PCR Methods -- 31.4.5.1.Multiplex PCR -- 31.4.5.2.Nested PCR -- 31.4.5.3.Real-Time PCR -- 31.4.5.4.Reverse Transcription -- PCR (RT -- PCR) -- 31.4.5.5.Quantitative Real-Time Reverse Transcription -- PCR (RT -- PCR) -- 31.4.5.6.Loop Mediated Isothermal Amplification (LAMP) -- 31.4.5.7.Cooperational PCR -- 31.4.5.8.Eric and Box PCR Amplification -- 31.5.Hybridization -- PCR Combination-Based Techniques -- 31.5.1.Amplified Fragment Length Polymorphism Markers (AFLP) -- 31.6.rRNA/DNA Based Techniques -- 31.6.1.Ribotyping or Ribosomal DNA Restriction Analysis -- 31.6.2.16S rRNA Sequencing -- 31.6.3.Internal Transcribed Spacer (ITS) rRNA Sequencing -- 31.7.Terminal Restriction Fragment Pattern (TRFP) Method -- 31.8.Pulse Field Gel Electrophoresis Technique (PFGE) -- 31.9.Guanine-Plus-Cytosine-Based Analysis --
Note continued: 31.10.Flow Cytometry for Bacterial Diagnosis -- 31.11.Deoxyribonucleic Acid Content of Bacterium, Its Relation to Virulence and Drug Resistance -- ch. 32 Bacteriophages of Plant Pathogenic Bacteria -- 32.1.Isolation of Lytic Phages -- 32.1.1.By Direct Method -- 32.1.2.Indirect Method or Enrichment Method -- 32.1.3.Detection of Phage -- 32.1.4.Characterization of Phage -- 32.2.Bacterial Phage Typing -- 32.3.Determination of Phage Titer -- 32.4.Isolation of Lysogenic Phages -- 32.5.Preservation of Bacteriophages -- ch. 33 Determination of Bacterial Sensitivity to Antibiotics and Pesticide -- 33.1.Determination of Bacterial Sensitivity by Poison Food Technique -- 33.2.Determination of Bacterial Sensitivity by Disc Assay Method -- 33.3.Determination of Minimal Inhibitory Concentration (MIC) of Antibiotics/Pesticides -- 33.4.Isolation of Antibiotics/Pesticide Resistant Mutant of Bacteria -- 33.5.Studies on Pesticide Cross Resistance in Bacteria --
Note continued: 33.6.Studies on Multidrug Resistance in Bacteria -- 33.7.Studies on Pathogenicity of Multidrug Resistance Bacteria -- 33.8.Curing of Bacterial Mutant from Antibiotic Resistance -- 33.9.Curing of Plasmid Borne Antibiotic Resistance -- ch. 34 Determination of Mutation in Bacterial Plant Pathogen -- 34.1.Isolation of Antibiotic Resistant Mutant by Gradient Plate Technique -- 34.2.Isolation of Avirulent Mutants (Mutation for Pathogenicity Character) -- 34.3.Isolation of Albino Mutant (Mutation for Pigmentation) -- 34.4.Differentiation of Virulent and Avirulent Mutant -- ch. 35 Transfer of Genetic Factor in Plant Pathogenic Bacteria -- 35.1.Transfer of Genetic Material through Transformation -- 35.2.Transfer of Genetic Material through Conjugation -- 35.3.Transfer of Genetic Material through Transduction -- ch. 36 Population Studies of Plant Pathogenic Bacteria on/in Host Plant -- 36.1.Estimation of Bacterial Population by Leaf Wash Method --
Note continued: 36.2.Estimation of Epiphytic Population by Leaf Impression Method -- 36.3.Estimation of Endophytic Population by Serial Dilution Method -- 36.4.Dynamics of Bacterial Population in Host Plant -- 36.5.Isolation of Intercellular Fluid to Study Bacterial Growth -- ch. 37 Preservation of Bacterial Disease Samples -- 37.1.Dry Preservation of Samples -- 37.2.Microwave Drying Method -- 37.3.Preservation of Samples in Preservation Solutions -- 37.4.Preservation of Leaf Spot/Blight Samples by Lamination -- 37.5.Preservation of Fruit Samples by Wax Treatment -- ch. 38 Measurement of Plant Bacterial Disease -- 38.1.Glossary to Describe Symptoms -- 38.2.Measurement of Leaf Spot/Blight Disease -- 38.3.Measurement of Wilt Disease -- ch. 39 Isolation of Antibacterial Compound -- 39.1.Preliminary Assessment Technique to Determine Presence of Antibacterial Compound in Plant by Computing Bacterial Population -- 39.2.Assessment of Antibacterial Properties of Plant --
Note continued: 39.3.Efficacy of Aqueous Extract of Medicinal Plant against Plant Pathogenic Bacteria (Xanthomonas spp.) -- 39.4.Preparation of Plants Extract in Solvents -- 39.5.Assessment and Quantification of Equivalent Antibacterial Compound in Solvent Plant Extract -- 39.6.Isolation of Antibacterial Compound from Plant Source -- 39.7.Isolation of Antibacterial Compound from Microbial Source -- 39.8.Testing Efficacy of Antibacterial Molecule -- 39.9.Identification of Antibacterial Molecule -- ch. 40 Isolation of Antibacterial Microbes -- 40.1.Isolation of Antibacterial Fungi -- 40.2.Isolation of Antibacterial Bacteria -- 40.3.Testing Antibacterial Property of Microbial Culture -- 40.4.Mass Production of Antibacterial Culture -- 40.4.1.Carrier-Based Mass Production -- 40.4.2.Liquid-Based Mass Production -- ch. 41 Preparation of Bactericidal Formulations -- 41.1.Preparation of Bactericidal Spray -- 41.1.1.Preparation of Bordeaux Mixture --
Note continued: 41.1.2.Preparation of Bactericidal Mixture -- 41.2.Preparation of Bactericidal Pastes -- 41.2.1.Preparation of Bordeaux Paste -- 41.2.2.Preparation of Rahuri Paste.
Includes bibliographical references and index.