Genomics of Crucifer's Host- Pathosystem

Foreword
Preface
Acknowledgments
Contents
About the Authors
Abbreviations
1: Genomics of Crucifer´s Host-Pathosystem: Prologue
1.1 Introduction
1.2 Economic Importance of Crucifers
1.3 The State of Brassica Genomics
1.4 Crucifers´ Host Genome
1.4.1 Arabidopsis thaliana
1.4.2 Brassica carinata
1.4.3 Brassica juncea
1.4.4 Brassica napus
1.4.5 Brassica nigra
1.4.6 Brassica oleracea
1.4.7 Brassica rapa
1.4.8 Crucifer´s as Ideal Host-Pathosystem for Genomics Research
1.4.9 Utilization of Omics Approaches in Crucifer´s Host-Pathosystem
1.4.9.1 High-Quality Genome Assemblies
1.4.9.2 Transcriptomics of Virulence-Related Genes
1.4.9.3 Secretomics of Host-Pathogen Interaction
1.4.9.4 Interactomics of Host-Pathogen Interaction
1.4.9.5 Metabolomics of Crucifers´ Host-Pathosystem
1.5 Genomics of Crucifers Biotic and Abiotic Stresses
1.6 Major and Devastating Stresses of Crucifer´s Crops
1.7 Expression of Genes to Overcome the Abiotic Stresses
1.8 Differential Gene Expression in Brassica Under Abiotic Stresses
1.9 Major Pathogens of Crucifers at Global Level
1.9.1 Albugo (White Rust)
1.9.1.1 The Causal Organism
1.9.1.2 Taxonomy and Morphology
1.9.1.3 The Disease and Symptomatology
1.9.1.4 Infection, Pathogenesis, and Disease Cycle
1.9.1.5 Epidemiology
1.9.1.6 Pathogenic Variability
1.9.1.7 Host Resistance
1.9.1.8 Disease Management
1.9.2 Alternaria (Alternaria Blight)
1.9.2.1 The Disease and Causal Organism
1.9.2.2 Infection and Pathogenesis
1.9.2.3 Epidemiology
1.9.2.4 Pathogenic Variability
1.9.2.5 Fine Structures
1.9.2.6 Host Resistance
1.9.2.7 Disease Management
1.9.3 Colletotrichum (Anthracnose)
1.9.3.1 The Disease and Causal Organism
1.9.3.2 Economic Importance
1.9.3.3 Host Range
1.9.3.4 Disease Management
1.9.4 Erysiphe (Powdery Mildew)
1.9.4.1 The Disease and Causal Organism
1.9.4.2 Infection, Pathogenesis, and Disease Cycle
1.9.4.3 Fine Structures and Electron Microscopy
1.9.4.4 Epidemiology and Forecasting
1.9.4.5 Host Resistance
1.9.4.6 Disease Management
1.9.5 Fusarium (Fusarium Wilt)
1.9.5.1 Fusarium Oxysporum f. sp. Conglutinans
The Disease and Causal Organism
Host Range and Distribution
Disease Management
1.9.5.2 Fusarium Oxysporum
The Disease and Causal Organism
Disease Management
1.9.5.3 Fusarium equiseti (Corda) Sacc.
The Disease and Causal Organism
Host Range
1.9.6 Hyaloperonospora (Downy Mildew)
1.9.6.1 The Disease and Causal Organism
1.9.6.2 Fine Structures and Electron Microscopy
1.9.6.3 Pathogenic Variability
1.9.6.4 Infection, Pathogenesis, and Disease Cycle
1.9.6.5 Epidemiology
1.9.6.6 Mixed Infection or Association of Downy Mildew and White Rust
1.9.6.7 Biometabolomics
1.9.6.8 Host Resistance
1.9.6.9 Disease Management
1.9.7 Leptosphaeria (Blackleg)
1.9.7.1 The Disease and Causal Organism
1.9.7.2 Economic Importance
1.9.7.3 Distribution
1.9.7.4 Pathogen Morphology
1.9.7.5 Epidemiology
1.9.7.6 Infection, Pathogenesis, and Disease Cycle
1.9.7.7 Disease Management
1.9.8 Plasmodiophora (Clubroot)
1.9.8.1 The Disease and Causal Organism
1.9.8.2 Fine Structures and Electron Microscopy
1.9.8.3 Pathogenic Variability
1.9.8.4 Infection, Pathogenesis, and Disease Cycle
1.9.8.5 Epidemiology and Disease Forecasting
1.9.8.6 Biochemistry of Host-Pathogen Interaction
1.9.8.7 Host Resistance
1.9.8.8 Disease Management
1.9.9 Pseudocercosporella (White Leaf Spot)
1.9.9.1 The Disease and Causal Organisms
1.9.9.2 Distribution
1.9.9.3 Host Range
1.9.9.4 Economic Importance
1.9.9.5 Epidemiology
1.9.9.6 Pathogenic Variability
1.9.9.7 Disease Management
1.9.10 Pyrenopeziza (Light Leaf Spot)
1.9.10.1 The Disease and Causal Organism
1.9.10.2 Economic Importance
1.9.10.3 Host Range
1.9.10.4 Epidemiology
1.9.11 Rhizoctonia (Seedling Rot; Collar Rot; Blight)
1.9.11.1 The Disease and Causal Organism
1.9.11.2 Host Range and Disease Distribution
1.9.11.3 Pathogenic Variability
1.9.11.4 Economic Importance
1.9.11.5 Epidemiology
1.9.11.6 Disease Management
1.9.12 Sclerotinia (Stem Rot)
1.9.12.1 The Disease and Causal Organism
1.9.12.2 Economic Importance
1.9.12.3 Pathogenic Variability
1.9.12.4 Infection, Pathogenesis, and Disease Cycle
1.9.12.5 Epidemiology and Forecasting
1.9.12.6 Host Resistance
1.9.12.7 Disease Management
1.9.13 Turnip Mosaic Virus-TuMV (Turnip Mosaic)
1.9.13.1 The Disease and Causal Organism
1.9.13.2 Economic Importance
1.9.13.3 Host Range
1.9.13.4 Pathogenic Variability
1.9.13.5 Host Resistance
1.9.13.6 Disease Management
1.9.14 Verticillium (Verticillium Wilt or Verticillium Stem Striping)
1.9.14.1 The Disease and Causal Organism
1.9.14.2 Economic Importance
1.9.14.3 Infection, Pathogenesis, and Host Resistance
1.9.14.4 Disease Management
1.9.15 Xanthomonas (Black Rot)
1.9.15.1 The Disease and Causal Organism
1.9.15.2 Economic Importance
1.9.15.3 Infection, Pathogenesis, and Disease Cycle
1.9.15.4 Host Range
1.9.15.5 Disease Management
1.10 Expression of Genes to Overcome the Biotic Stresses
1.11 Differential Genes Expression in Brassica Under Biotic Stresses
1.12 Selection of Reference Genes for qRT-PCR Normalization in Brassica Napus Under Various Stress Conditions
1.13 Signaling System of Crucifers´ Host-Pathosystem in Response to Multiple Stresses
1.14 Genomics of Crucifers´ Host Resistance
References
2: Crucifer´s Pathogens Genome
2.1 Introduction
2.2 Brassica-Albugo Genome
2.2.1 Introduction
2.2.2 Albugo candida Genome
2.2.3 Evolution of Obligate Parasitism in Albugo-Arabidopsis
2.2.4 Genome Introgression of Albugo candida Races
2.2.4.1 Single Race Isolates of Albugo candida are Host-Specific
2.2.4.2 Genome Assemblies of Albugo candida Isolates
2.2.4.3 Genome-Wide Similarity Between Races with Non-overlapping Host Range
2.2.4.4 Mosaic-Like Genome Structure of Albugo candida Races
2.2.4.5 Intrarace Diversity Suggests Clonal Propagation After Creation of Novel Adapted Allele Repertoires
2.2.4.6 Sequential Infections Abolish Host Specificity of Susceptibility to Albugo candida Infection
2.3 Brassica-Alternaria Genome
2.3.1 Introduction
2.3.2 Alternaria brassicae Genome
2.3.3 Comparative Genomics of Alternaria Species
2.3.4 Genomic Features of Alternaria brassicae and Alternaria alternata Isolates
2.3.5 Phylogenomics of Alternaria brassicae and Alternaria brassicicola Within Alternaria Genus
2.3.6 Comparative Analysis of Alternaria alternata Isolates Obtained from Different Hosts
2.3.7 Bioinformatic Analysis of Expressed Sequence Tags in Compatible Alternaria-Brassica Interaction
2.3.8 Characterization of Histidine Kinase Gene AbNIK1 from Alternaria Conferring High Dicarboximide and Phenylpyrrole Resista...
2.3.9 Phylogenetic Relationship of Alternaria species Determined With Nuclear Ribosomal DNA Sequences
2.4 Brassica-Colletotrichum Genome
2.4.1 Introduction
2.4.2 Genome Sequencing and Assembly
2.4.3 Transcriptome Analysis
2.4.4 Genetic Transformation
2.5 Brassica-Erysiphe Genome
2.5.1 Introduction
2.5.2 Present Status of Crucifers´ Powdery Mildew Pathogen Genome
2.5.3 Phylogenetics of Powdery Mildew Pathogens
2.5.3.1 Sequence Analysis of the ITS rDNA
2.5.3.2 Sequence Comparison of DNA Encoding the 5.8S rRNA, ITS1, and ITS2
2.6 Brassica-Fusarium Genome
2.6.1 Introduction
2.6.2 Fusarium oxysporum Genome
2.7 Brassica-Hyaloperonospora Genome
2.7.1 Introduction
2.7.2 Hyaloperonospora Genome
2.7.2.1 Characteristics of Hyaloperonospora arabidopsidis Genome
2.7.2.2 Genome Architecture of Hyaloperonospora brassicae Isolates for Virulence
2.7.2.3 Genome Assemblies of Hyaloperonospora brassicae Isolates and Their Functional Characteristics
2.7.2.4 Draft Genome Assemblies from Whole-Genome Sequences of Two Hyaloperonospora brassicae Isolate Samples Differing in Fie...
2.7.3 Phylogeny of Hyaloperonospora
2.7.4 Major Species Clusters in Hyaloperonospora
2.7.5 Hyaloperonospora arabidopsidis on Arabidopsis
2.8 Brassica-Leptosphaeria Genome
2.8.1 Introduction
2.8.2 Leptosphaeria maculans Genome
2.8.3 Leptosphaeria species Genomes
2.8.4 Identification of a New Subclade of Leptosphaeria from Brassica
2.8.5 Gene Loss in Leptosphaeria maculans
2.8.6 Contrasting Genomic Organization of Leptosphaeria species
2.9 Brassica-Plasmodiophora Genome
2.9.1 Introduction
2.9.2 Early Phase of Plasmodiophora brassicae Genomics
2.9.3 Main Features of the Nuclear Genome of Plasmodiophora
2.9.4 Mitochondrial Genome, Structure, and Gene Content
2.9.5 Nuclear Genome Annotation
2.9.6 Metabolic Pathway Database
2.9.7 Prediction of New Pathways
2.9.8 Advantages of Plasmodiophorid Genomics
2.9.9 Distinguishing Features of Plasmodiophora brassicae Genome
2.9.10 Evolution and Phylogeny of Plasmodiophora brassicae
2.9.11 Gene´s Discovery of Plasmodiophora brassicae Isolates
2.9.12 The cDNA Cloning Approaches to Identify Plasmodiophora brassicae Genes
2.9.13 Plasmodiophora brassicae Gene Structure
2.10 Brassica-Sclerotinia Genome
2.10.1 Introduction
2.10.2 Early Phase of Sclerotinia sclerotiorum Genome
2.10.3 Phylogeny of Sclerotinia and Related Genera
2.10.4 Expression of Genes for Enzymes Activity
2.10.5 Expression of Calcineurin Genes for Pathogenesis
2.10.6 The Main Features of the Sclerotinia sclerotiorum Genome
2.10.6.1 Genome Sequences
2.10.6.2 Transcriptomic and Secretomic Analysis
2.10.7 Genome Architecture of Sclerotinia sclerotiorum
2.10.7.1 A Complete New Assembly of the Sclerotinia sclerotiorum Genome
2.10.7.2 Gene Annotations in the New Sclerotinia sclerotiorum Genome
2.10.7.3 Identification of a Seven-Member Effector Gene Family in Sclerotinia sclerotiorum
2.11 Brassica-Turnip Mosaic Virus Genome
2.11.1 Introduction
2.11.2 Genome Sequence of Turnip Mosaic Virus
2.11.3 The Turnip Mosaic Virus Genome Characteristics and the Virus Phylogeny
2.11.4 Genome Analysis of Turnip Mosaic Virus Co-infecting Brassica
2.11.5 Phylogenetic Analysis
2.11.6 Mutations in Turnip Mosaic Virus Genomes Adapted to Raphanus sativus
2.11.7 The Turnip Mosaic Virus Genetic Diversity
2.12 Brassica-Xanthomonas Genome
2.12.1 Introduction
2.12.2 Regulation of Secretion Systems of Xanthomonas Genome
2.12.3 Function of Type III Effector Genes of Xanthomonas
2.12.4 Virulence and Pathogenicity Effectors/Genes of Xanthomonas
2.12.5 Characterization of Xanthomonas Genome for Virulence Genes
2.12.6 Genomic Diversity in the Xanthomonas campestris species
References
3: Genomics of Host-Pathogen Interaction
3.1 Introduction
3.2 Brassica-Albugo Pathosystem
3.2.1 Introduction
3.2.2 R Genes Function During Albugo Pathogenesis
3.2.3 Molecular Mechanisms of Host-Pathogen Interaction
3.2.4 Regulation of Host-Pathogen Interaction by R Genes
3.3 Brassica-Alternaria Pathosystem
3.3.1 Introduction
3.3.2 Identification of Species-Specific Genes of Alternaria with Putative Roles in Virulence
3.3.3 Transcription Factors Associated with Pathogenesis
3.3.4 The AbPf2 Gene Activates Pathogenicity in Alternaria brassicicola
3.3.5 Identification, Cloning, and Sequencing of Virulence Genes
3.3.6 Identification of Pathogenicity Factors/Genes
3.3.7 Microarray Analysis of Brassica Infected with Alternaria
3.3.8 Changes in Host Cell Ultrastructure and Transcriptome Reprogramming
3.3.9 Defense-Related Genes are Activated in Susceptible Interaction
3.4 Brassica-Arabidopsis-Colletotrichum Pathosystem
3.4.1 Introduction
3.4.2 Mitogen-Activated Protein (MAP) Kinase and cAMP/PKA Signaling Pathway for Pathogenesis
3.4.3 Identification of Effectors/Genes during Pathogenesis
3.4.4 Role of MAPK Gene in Development of Anthracnose Disease
3.4.5 Molecular Events during Host-Pathogen Interaction
3.5 Brassica-Erysiphe Pathosystem
3.5.1 Introduction
3.5.2 Genomics of Powdery Mildew Pathogenesis
3.5.3 The MLO Proteins (Genes) in Powdery Mildew Pathogenesis
3.5.4 Host Transcriptional Regulation of Powdery Mildew Pathogenesis
3.5.5 Transcriptional Programming of Powdery Mildew Pathogenesis
3.5.5.1 Transcriptional (Genes) Regulation and Expression in Response to Powdery Mildew Infection
3.5.6 Genes Governing Powdery Mildew Pathogenesis
3.5.7 Function of Lifeguard Protein (LFG) in Powdery Mildew Pathogenesis
3.5.8 Gene Expression Levels of Healthy and Powdery Mildew-Infected Plants
3.6 Brassica-Hyaloperonospora Pathosystem
3.6.1 Introduction
3.6.2 Genes Governing Pathogenesis
3.6.3 Accommodation of Infection Structures during Pathogenesis
3.7 Brassica-Leptosphaeria Pathosystem
3.7.1 Introduction
3.7.2 Molecular Mechanisms of Pathogenesis
3.7.3 Characterization of Genome and Transcriptomes During Interaction Between Brassica and Leptosphaeria Species
3.7.4 Green Fluorescent Protein as a Reporter in the Brassica-Leptosphaeria Interaction
3.8 Brassica-Plasmodiophora Pathosystem
3.8.1 Introduction
3.8.2 Molecular Biology of Plasmodiophora brassicae Pathogenesis
3.8.3 Gene Functions at the Host-Pathogen Interaction
3.8.4 The Plasmodiophora brassicae Genome and Life Stage-Specific Transcriptomics
3.8.5 Functional Characterization of Genes During Pathogenesis
3.8.6 Effector Proteases
3.8.6.1 A-Cysteine-Rich Proteins
3.8.6.2 The Rxlr Motifs
3.8.6.3 Chitin-Binding Domains
3.8.6.4 Protease/Protease Inhibitors
3.8.6.5 Nuclear Localization Domains
3.8.6.6 The Pexel Motif
3.8.6.7 Plant Pathogenic Plasmodiophorids
3.8.6.8 Role of Effectors (Genes) in Clubroot Life Cycle
3.8.6.9 A Coherent Pipeline to Identify Effectors in Future
3.8.7 Role of a Serine Protease Pro 1 in Plasmodiophora brassicae Pathogenesis
3.8.8 Role of Lipids in Plasmodiophora brassicae Pathogenesis
3.8.9 Relationship of Endogenous Glucosinolate Content to Plasmodiophora brassicae Infection
3.8.10 Effect of Host Resistance on Plasmodiophora brassicae Pathogenesis
3.8.11 Molecular Mechanisms of Pathogenesis
3.8.12 Host Gene Expression During the Primary Phase of Plasmodiophora brassicae Interaction
3.8.13 Differential Expression of Genes During Infection of Plasmodiophora brassicae
3.8.14 Differential Expression of Host Shoots and Roots Genes During Plasmodiophora brassicae Infection
3.8.15 Expression of Hypoxia Genes in Arabidopsis Roots Infected by Plasmodiophora brassicae
3.8.16 Expression of SWEET Genes During Plasmodiophora brassicae Infection
3.8.17 Expression of Host-Specific Genes
3.8.17.1 Brassica rapa
3.8.17.2 Arabidopsis thaliana
3.8.18 Expression of miRNAs in Clubroot Pathogenesis
3.8.19 Expression of Immunophilins During Plasmodiophora brassicae Infection
3.8.20 Analysis of Host Response Through Transcriptome Data
3.8.20.1 Transcriptome Analysis of Brassica-Plasmodiophora Interaction at Early Infection Stage
3.8.21 Manipulation of Host Defense/Resistance
3.8.22 Role of G-Protein-Coupled Receptors (GPCR) Signal Transduction Pathways in Plasmodiophora brassicae Pathogenesis
3.9 Brassica-Sclerotinia Pathosystem
3.9.1 Introduction
3.9.2 Role of Enzymes in Pathogenesis
3.9.3 Expression of Genes During Pathogenesis
3.9.4 Molecular Biology of Sclerotinia sclerotiorum for Regulation of Growth, Virulence, and Pathogenesis
3.9.4.1 Regulation of Mycelial Growth and Virulence
3.9.4.2 Signaling Events for Sclerotial Formation
3.9.4.3 Control of Apothecial and Ascospore Development
3.9.4.4 Regulation of Pathogenesis
3.9.4.5 Changes in Sclerotinia sclerotiorum Transcriptome During Infection of Brassica napus
3.9.5 Identification of an Arabidopsis NLR Genes Conferring Susceptibility in Brassica to Sclerotinia
3.9.6 Survival Factor 1 Contributes in Pathogenesis of Sclerotinia sclerotiorum
3.9.7 Differential Expression of Multiple Metabolic Enzymes on Different Hosts During Sclerotinia Pathogenesis
3.9.7.1 Differential Gene Expression in Brassica Genotypes in Response to Sclerotinia Infection
3.9.7.2 Differential Gene Expression in Brassica napus Cultivars Infected with Sclerotinia sclerotiorum
3.9.8 Integrated mRNA, sRNA, and Degradome Sequencing Reveal Complex Responses to Sclerotinia Infection in Brassica
3.9.9 The Interplay of Apoptosis and Autophagy in the Pathogenesis of Sclerotinia sclerotiorum
3.9.10 A Secretory Protein of Sclerotinia sclerotiorum Suppresses Host Resistance in Early Stage of Pathogenesis
3.9.11 Mechanisms of Oxaloacetate Acetyl Hydrolase Gene of Sclerotinia in Pathogenesis
3.9.12 Functional Characterization of BNaNPR1 Gene Family During Sclerotinia Pathogenesis
3.9.13 The Effector-Like Proteins Regulate Sclerotinia Pathogenesis
3.9.14 Role of Copper in Brassica-Sclerotinia Interaction
3.9.15 Genome-Wide Alternative Splicing Profile of Sclerotinia During the Colonization of Diverse Host Families
3.10 Brassica-Turnip Mosaic Virus Pathosystem
3.10.1 Introduction
3.10.2 Molecular Mechanism of Turnip Mosaic Virus Pathogenesis
3.10.3 The Turnip Mosaic Virus Pathogenicity-Determinant Genes
3.10.4 The BnSGS3 Gene in Brassica has Differential Effects on the Pathogenesis of Viruses
3.10.5 Identification of Pathogenesis Modulating Genes in Arabidopsis-Turnip Mosaic Virus Interaction
3.11 Brassica-Verticillium Pathosystem
3.11.1 Introduction
3.11.2 Differential Expression of Proteins During Verticillium Pathogenesis of Brassica
3.12 Brassica-Xanthomonas Pathosystem
3.12.1 Introduction
3.12.2 Secretion System of Xanthomonas Genome Regulates Pathogenicity
3.12.3 Functional Characterization of Type III Effector Genes of Xanthomonas in Pathogenesis
3.12.4 Virulence and Pathogenicity Effectors/Genes of Xanthomonas
3.12.5 Characterization of Xanthomonas Gene for Virulence
3.12.6 Hormonal Signaling in the Pathways of Biotrophy-Necrotrophy Transition of Xanthomonas Infection in Brassica
3.13 Impact of Evolutionary Genomics on Host-Pathogen Interaction
3.14 Challenges of Genomics of Host-Pathogens Interaction
3.15 Advantages of Gene Fragments to Whole-Genome Analysis
References
4: Molecular Detection and Identification of Pathogens, Pathotypes, and Genes
4.1 Introduction
4.2 Use of qRT-PCR Normalization in Crucifers Under Various Stress Conditions
4.3 Brassica-Albugo Pathogen
4.3.1 Sequence Capture Detection of Albugo Sequences from Infected Leaves
4.3.2 The PenSeq Investigation of DNA Sequence Variation in Pathogen Genotypes from Field
4.3.3 Identification of Recombination Break Points of Different Races
4.3.4 The PenSeq Identification of Race Structure with Phylogenetic Lineage
4.3.5 A PCR-Based Assay for Detection of Albugo
4.4 Brassica-Alternaria Pathogen
4.4.1 A Diagnostic Biomarker for the Identification of Alternaria brassicae
4.4.2 Identification of Pathotypes Using Molecular Techniques
4.4.2.1 Genetic Variability
4.4.2.2 Proteome Analysis
4.4.2.3 Biochemical Variations
4.5 Brassica-Colletotrichum Pathogen
4.5.1 Identification of Effectors/Genes of Colletotrichum for Virulence
4.6 Brassica-Erysiphe Pathogen
4.6.1 Molecular Quantification of Powdery Mildew
4.6.1.1 Plant Material and Inoculation Methods
4.6.1.2 Genomic DNA Extraction
4.6.1.3 Quantitative Real-Time PCR
4.6.1.4 Spore Counts
4.6.2 Detection of Genes Expression in Response to Powdery Mildew Infection
4.6.3 Phylogenetics of Erysiphe
4.6.3.1 Sequence Analysis of the ITS rDNA
4.6.3.2 Sequence Comparison of DNA Encoding the 5.8S rRNA, ITS1, and ITS2 Regions
4.7 Brassica-Hyaloperonospora Pathogens
4.7.1 Molecular Identification of Hyaloperonospora parasitica Virulence Using DNA Fingerprinting
4.7.2 Identification of Downy Mildew R Genes
4.8 Brassica-Leptosphaeria Pathogen
4.8.1 Combined LMD and RNA Sequencing Improves Pathogen Detection at the Earliest Stages of Infection
4.8.2 Molecular Identification of Leptosphaeria Subclades
4.8.3 Genomic Prediction to Identify Blackleg R Genes of Brassica
4.9 Brassica-Plasmodiophora Pathogen
4.9.1 Identification of Virulence Genes of Plasmodiophora
4.9.2 The qPCR-Based Detection of Plasmodiophora Spread in Soil
4.9.3 Identification of Race/Pathotype-Specific Molecular Markers
4.9.4 Molecular Identification of Plasmodiophora Pathotype Effector/Gene
4.10 Brassica-Pseudocercosporella Pathogen
4.10.1 Phylogenetic Analysis of Pseudocercosporella Isolates for Virulence
4.11 Brassica-Rhizoctonia Pathogen
4.11.1 Molecular Detection of Rhizoctonia solani Virulence Distribution in Soil
4.12 Brassica-Sclerotinia Pathogens
4.12.1 Rapid Identification of an Arabidopsis NLR Gene Conferring Susceptibility to Sclerotinia sclerotiorum
4.13 Brassica-Turnip Mosaic Virus Pathogen
4.13.1 Detection and Differentiation of Brassica Virus by Multiplex Reverse Transcription Polymerase Chain Reaction
4.13.2 Detection and Identification of Turnip Yellows Virus in Brassica by DAS-ELISA Kits
4.13.3 Identification of Turnip Mosaic Virus From Brassica napus by DAS-ELISA
4.13.4 Molecular Detection, Identification, and Phylogenetic Analysis of Turnip Yellows Virus (TuYV)
4.13.5 The Turnip Mosaic Virus Detection, Preservation, and Identification by ELISA
4.13.6 The Turnip Mosaic Virus Biological and Serological Detection
4.14 Brassica-Verticillium Pathogen
4.14.1 Identification and Differentiation of Verticillium Species by Simplex and Multiplex PCR Assays
4.14.2 Molecular Characterization of Verticillium longisporum on the Basis of a Group-I Intron in SSU-rRNA Gene
4.14.3 Molecular Characterization of Verticillium Isolates
4.15 Brassica-Xanthomonas Pathogen
4.15.1 Identification of Virulence Effectors/Genes of Xanthomonas
4.15.2 Novel Genes Identification Through Molecular Approaches
4.16 Molecular Identification of Pathotype and Pathogen Elicitors/Effectors
4.17 The qRT-PCR Analysis of Brassica Reference Genes Under Various Stresses
4.18 Disentangling Hidden Histories in Genes and Genomes of Hosts and Pathogens
4.19 Identification of R Genes Homologous DNA Fragments in Arabidopsis thaliana
4.20 Comparisons Between Different Diagnostic Approaches
References
5: Pathogenomics of Pathogenic Variability
5.1 Introduction
5.2 Brassica-Albugo Pathogenomics
5.2.1 Polyploidy as a Contributor to Albugo Race Diversification and Evolution
5.2.2 Pathogenic Diversity in Albugo
5.2.3 Phylogenetic Relationship Between Albugo Species
5.2.4 Identification of Pathogenic Variability in Albugo candida
5.2.5 Virulence Spectrum of Virulence Genes/Pathotypes/Races
5.2.6 Phylogenetic Relationship of Pathotypes/Races
5.2.7 Phylogenetic Analyses and Nucleotide Sequence Identities
5.2.8 Characterization of Albugo candida Virulence Genes on Crucifers
5.2.9 Inheritance of Avirulence in Albugo candida
5.3 Brassica-Alternaria Pathogenomics
5.3.1 Genetic Variation Among Alternaria brassicae Isolates
5.3.2 Genetic Structure of Alternaria Populations Suggest the Occurrence of Sexual Recombination
5.3.3 Mutation of Amr1 Gene Causes Increased Virulence
5.3.4 The Cause of Increased Virulence in Δamr1 Mutants
5.3.5 Evolution of Virulence in Alternaria brassicicola
5.3.6 Identification of Pathological Variations/Pathotypes
5.3.6.1 Symptomatological Variations
5.3.6.2 Morphological and Cultural Variations
5.3.7 Identification of Pathotypes Using Molecular Approaches
5.3.7.1 Genetic Variability
5.3.7.2 Proteome Analysis
5.3.8 Biochemical Variations
5.3.9 Variations in Fungicidal and Plant Extracts Sensitivity
5.3.10 Virulence Variability of Alternaria species on Crucifers
5.3.11 Identification and Nomenclature of Pathotypes
5.4 Brassica-Colletotrichum Pathogenomics
5.4.1 Expression of Virulence Effectors/Genes
5.5 Brassica-Erysiphe Pathogenomics
5.5.1 Identification of Pathotypes of Erysiphe
5.6 Brassica-Hyaloperonospora Pathogenomics
5.6.1 Identification of Virulence Effectors/Genes/Pathotypes
5.6.2 Identification of Host Differentials and Nomenclature of Pathotypes
5.6.3 Heterothallism and Homothallism
5.6.4 Hybridization of Hyaloperonospora Isolates
5.7 Brassica-Leptosphaeria Pathogenomics
5.7.1 Evolution of Virulence in Leptosphaeria
5.7.2 Avirulence Alleles of Leptosphaeria maculans in Europe
5.7.3 Genetic Variability in Leptosphaeria maculans from France
5.7.4 Characterization of Avirulent-Mutated AvrLm Gene of Leptosphaeria
5.7.5 Identification of Genome-Wide DNA Variants and SNP Haplotypes of Leptosphaeria maculans Pathotypes
5.8 Brassica-Plasmodiophora Pathogenomics
5.8.1 Differentials for Identification of Pathotypes
5.8.2 Genome Comparison of Plasmodiophora brassicae Pathotypes
5.8.3 Proteomics of Plasmodiophora brassicae Pathotypes for Potential Effectors (Genes)
5.8.4 Phylogenetic Analysis of Pathotypes
5.8.5 Polymorphisms in Plasmodiophora brassicae
5.8.6 Molecular Marker for Identification of Plasmodiophora brassicae Pathotypes
5.8.7 Identification of Geographical Isolates of Plasmodiophora brassicae by Ribosomal DNA Sequence Polymorphisms
5.8.8 Race and Isolate-Specific Molecular Marker
5.8.9 Pathogenic and Genetic Variation in Plasmodiophora brassicae Pathotypes
5.8.10 Molecular Identification of Pathotypes and Pathogen Elicitors/Effectors
5.8.11 Role of Pathogenic Variability in the Genetic Complexity of Clubroot Resistance (CR) Genes
5.8.12 Whole Genome DNA Similarity and Pathotype Population Structure of Plasmodiophora brassicae
5.8.13 Genotyping of Plasmodiophora brassicae Reveals Distinct Population
5.9 Brassica-Pseudocercosporella Pathogenomics
5.9.1 Identification of Virulence Factors/Races
5.10 Brassica-Sclerotinia Pathogenomics
5.10.1 Genetic Diversity of Sclerotinia sclerotiorum Isolates
5.10.2 Aggressiveness of Sclerotinia sclerotiorum Isolates
5.10.3 Genetic Diversity in Sclerotinia sclerotiorum Population from Brassica juncea
5.10.4 Genetic Diversity of Sclerotinia sclerotiorum in the North Central United States
5.10.5 Genetic Diversity and Structure of Sclerotinia sclerotiorum Populations Detected by Random Amplified Polymorphic DNA Ma...
5.10.6 Identification of Virulence Factors
5.10.6.1 Pathogenicity
5.10.6.2 Genomic Factors
5.10.6.3 Mycelial Compatibility
5.10.6.4 Genetic Diversity of Isolates
5.10.6.5 Population Biology
5.11 Brassica-Turnip Mosaic Virus Pathogenomics
5.11.1 Pathological and Biological Characteristics of Turnip Mosaic Virus Isolates
5.12 Brassica-Verticillium Pathogenomics
5.12.1 Molecular Characterization of Verticillium Isolates from Crucifers
5.13 Brassica-Xanthomonas Pathogenomics
5.13.1 Identification of Pathotypes of Xanthomonas
5.13.2 Genetic Diversity in Virulence of Xanthomonas Isolates from Brassica
5.14 Evolution of Virulent Effectors of Pathogens
5.15 Challenges of Pathotype Identification, Classification, and Utilization
5.16 Challenges of Genetic Variation in Pathogenic Populations
References
6: Biometabolomics of Crucifer´s Host-Pathosystem
6.1 Introduction
6.2 Brassica-Albugo Host-Pathosystem
6.2.1 Regulation of Photosynthesis, Carbohydrate Metabolism, and Respiration
6.2.2 Induction of Hypertrophy by Hormonal Regulation
6.2.3 Differential Production of Lipid Profiles in Albugo species Sporangia
6.2.4 Differential Activity of Enzymes in Infected and Healthy Host Tissues
6.3 Brassica-Alternaria Host-Pathosystem
6.3.1 Genomic Regulation of Alternaria Brassicae Secondary Metabolites/Toxins
6.3.2 Genetic Control of Destruxin B Production by Alternaria brassicae
6.3.3 Induction of Biomolecular Changes in Brassica by Alternaria Infection
6.3.4 Alternaria-Arabidopsis Pathosystem as Model to Predict Disease Pathophysiology
6.3.5 Transcriptional Factor Amr1 Regulates Melanin Biosynthesis and Virulence in Alternaria
6.3.6 Production of Metabolites by Alternaria Related to Pathogenesis
6.3.7 Alternaria Metabolites Influences Host Physiology, Biochemistry, and Molecular Process
6.3.7.1 Effect of Alternaria Toxin on Host Physiology
6.3.7.2 Effect of Alternaria Toxins on Host Biochemistry
6.3.7.3 Effect of Alternaria Toxin on Molecular Process
6.3.7.4 Role of Metabolites in the Infection Process of Alternaria
6.3.7.5 Biomolecular Alteration During Brassica-Alternaria Interaction
6.4 Brassica-Colletotrichum Host-Pathosystem
6.4.1 Biosynthesis of Biomolecules During Pathogenesis
6.5 Brassica-Erysiphe Host-Pathosystem
6.5.1 Host Metabolism and Increased Ploidy Level of Mesophyll Cells Regulates Powdery Mildew Infection
6.5.2 Powdery Mildew Infection and Pathogenesis Regulates Transcriptional Events
6.6 Brassica-Hyaloperonospora Host-Pathosystem
6.6.1 Induction of Biometabolites Regulates Pathogenesis
6.7 Brassica-Leptosphaeria Host-Pathosystem
6.7.1 Expression of Metabolic Genes of Brassica During Leptosphaeria Pathogenesis
6.7.2 Modulation of Host Metabolism by Leptosphaeria Effector Genes During Pathogenesis
6.8 Brassica-Plasmodiophora Host-Pathosystem
6.8.1 Role of Aliphatic Glucosinolates Metabolism in Clubroot Pathogenesis
6.8.2 Induction and Translocation of Sugars in Clubroot Tissues
6.8.3 Role of Host Hormone Metabolism in Clubroot Development
6.8.4 Plasmodiophora Induces Alterations of Host Cell Wall Rigidity by Metabolic and Physiological Changes
6.8.5 Genomics of Nutrient Acquisition Pathways
6.8.6 Prediction of New Metabolic Pathways During Plasmodiophora Pathogenesis
6.8.7 Induction of Signaling Molecules and Genes During Gall Formation
6.9 Brassica-Sclerotinia Host-Pathosystem
6.9.1 Metabolites Associated with Sclerotinia Pathogenesis
6.9.2 Identification of Cell Wall Degrading Enzymes of Sclerotinia
6.9.3 Differential Expression of Genes by Sclerotinia for Virulence
6.9.4 Role of BnaNPR1 Gene Family During Sclerotinia Pathogenesis
6.10 Brassica-Xanthomonas Host-Pathosystem
6.10.1 Alteration of Hormonal Balance in Brassica Cultivars During Xanthomonas Pathogenesis
References
7: Genomics of Crucifer´s Host-Pathosystem at a Glance
7.1 Introduction, Importance, and Scope of Crucifers´ Host-Pathosystem
7.2 Characterization of Brassica Species Genome
7.3 Impact of Pathogen Genome Sequencing
7.4 Pathogens Genome Assembly and Characteristics
7.5 Genomic Regulation of Crucifers´ Host-Pathogen Interaction
7.6 Genomic Mechanism of Crucifers´ Host-Pathogen Interaction
7.7 Importance and Scope of Molecular Detection and Identification
7.8 Detection and Identification of Pathogens, Pathotypes, and Effector Genes with Molecular Approach
7.9 Significance of Pathogenic Variability
7.10 Genomics of Pathogenic Variability
7.11 Metabolic Events of Crucifers´ Host-Pathosystem
7.12 Crucifers´ Host-Pathosystem Metabolomics
7.13 Genomics Application to Develop Resistant and Genetically Modified (GM) Crops Against Multiple Stresses
7.13.1 High-Quality Genome Assembles
7.13.2 Genomics of Host-Pathosystem
7.13.3 Genetical and Molecular Mechanisms of Host Resistance
7.13.4 Genetically Modified (GM) Crops to Manage Biotic Stresses
7.14 Prospects of Indian Mustard Genetically Modified (GM) Crop
7.15 Crucifers´ Host-Pathosystem Genomics Methodology
References
8: Protocols to Study Host-Pathosystems
8.1 Introduction
8.2 Brassica-Albugo Pathogenesis Protocols
8.2.1 Use of DNA Sequence Approach Under Field Conditions
8.2.1.1 Preparation of Samples for Reconstruction Experiments
8.2.1.2 Read Alignment and Consensus Sequences Calling
8.2.1.3 Phylogenetic, Nucleotide Diversity, and Recombination Analyses
8.2.1.4 Evaluation of Albugo candida Heterozygosity and Ploidy Level
8.2.1.5 Microbiome Analysis
8.2.2 A PCR Assay for Detection of Albugo candida DNA
8.2.2.1 DNA Extraction, Sequencing, and Primer Design
8.2.2.2 Primer Effectiveness and Specificity Trials
8.3 Brassica-Alternaria Pathogenesis Protocols
8.3.1 Alternaria Genome Sequencing
8.3.1.1 Genome Sequencing and Assembly
8.3.1.2 mRNA Sequencing and Transcript Reconstruction
8.3.1.3 Gene Prediction and Annotation
8.3.1.4 Orthology and Whole-Genome Phylogenetic Analysis
8.3.1.5 Relationship of TEs and Repeat-Rich Regions to Genes and Gene Clusters
8.3.2 The PCR-Based Assay for Detecting Alternaria in Seed
8.3.2.1 Preparation of Seed Samples
8.3.2.2 The DNA Manipulation
8.3.2.3 The PCR-Based Assay
8.3.3 Microarray Analysis of Brassica-Alternaria Interactions
8.3.3.1 Photosynthetic Pigment Content
8.3.3.2 Chlorophyll a Fluorescence
8.3.3.3 Statistical Analysis
8.3.4 Biochemical and Molecular Analysis for Mechanism of Resistance in Brassica to Alternaria
8.3.4.1 Plant Growth, Testing, and Sampling
8.3.4.2 Biochemical Assays
8.3.4.3 Assessment of ROS Generation (H2O2, HO*, and O2-)
8.3.4.4 Assessment of Plant Redox Enzymic Activities
8.3.4.5 Accumulation of Free Phenols and Deposition of Lignins
8.3.5 Virulence Variability of Alternaria species Infecting Crucifers´ species
8.3.5.1 Alternaria spp. Isolates and Preparation of Inoculum
8.3.5.2 Virulence Tests
8.3.5.3 Disease Assessment
8.3.5.4 Experimental Design and Statistical Analysis
8.4 Brassica-Colletotrichum Pathogenesis Protocols
8.4.1 Characterization of MAPK Gene During Brassica-Colletotrichum Interaction
8.4.1.1 Fungal Strains, Plants, and Culture Condition
8.4.1.2 Bioinformatics Data and Programs
8.4.1.3 The ChMK1 Gene Replacement and Complementary
8.4.1.4 DNA Manipulation and Southern Blot Analysis
8.4.1.5 RNA Manipulation and qRT-PCR
8.4.1.6 Characterization of ChMK1 Deletion Transformant and Wild-Type Strain
8.4.1.7 Cell Wall Sensitivity Assay
8.5 Brassica-Erysiphe Pathogenesis Protocols
8.5.1 Molecular Identification of Anamorphic Powdery Mildews (Erysiphales)
8.5.1.1 Primer Design and Test Specimen Methods
8.5.1.2 DNA Extraction and PCR
8.5.1.3 Anamorph-Teleomorph Connection
8.5.2 The DNA Sequence Analysis
8.5.3 The DNA Marker Analysis
8.5.4 Histological Assessment of Erysiphe cruciferarum Growth
8.6 Brassica-Hyaloperonospora Pathogenesis Protocols
8.6.1 Molecular Marker for Identification of Hyaloperonospora parasitica
8.6.1.1 Hyaloperonospora parasitica Isolates and Conidia Isolation
8.6.1.2 Brassica oleracea Fungal Pathogens Selection and Growth
8.6.1.3 DNA Isolation
8.6.1.4 ITS Amplification
8.6.1.5 ITS Restriction Assay
8.6.1.6 ITS2 Sequencing
8.6.1.7 Internal Primer Design and Multiplex PCR
8.6.2 The DNA Finger Printing of Hyaloperonospora parasitica
8.6.3 The DNA Isolation
8.6.3.1 Fungal Isolates
8.6.3.2 Isolates from Host Plants
8.6.3.3 Polymerase Chain Reaction (PCR) Materials
8.6.3.4 Amplification Conditions
8.6.3.5 Hybridization Conditions
8.7 Brassica-Leptosphaeria Pathogenesis Protocols
8.7.1 Characterization of Leptosphaeria Isolates
8.7.1.1 Fungal Population to Identify Avirulence Gene Frequencies
8.7.1.2 Plants and Inoculation
8.7.1.3 Fungal Reference Isolates
8.7.1.4 Scoring
8.7.2 Genetic Variability in Leptosphaeria
8.7.2.1 Sampling of Leptosphaeria maculans Field Populations
8.7.2.2 Pathogenicity Tests
8.7.2.3 Mating Type Determination
8.7.2.4 Mini-Satellite Analysis
8.7.2.5 Statistical Analysis
8.7.3 Phylogenetic Relationship of R-Loci in Brassica to Leptosphaeria
8.7.3.1 Sample Collection, DNA Extraction, and PCR Amplification
8.7.3.2 Statistical Data Analysis
8.7.4 Transcriptome Analysis of Brassica-Leptosphaeria Pathosystem
8.7.4.1 Plant Growth and Inoculation
8.7.4.2 Tissue Processing
8.7.4.3 Sectioning and Laser Microdissection
8.7.4.4 Library Construction
8.7.4.5 Data Analysis
8.7.4.6 Real-Time qPCR Experiments
8.7.4.7 Jasmonic Acid Measurements
8.8 Brassica-Plasmodiophora Pathogenesis Protocols
8.8.1 The PCR Detection of Clubroot Pathogen
8.8.2 The PCR Technique to Identify Pathogen
8.8.3 Detection of Plasmodiophora brassicae Using Loop-Mediated Isothermal DNA Amplification
8.8.4 A Real-Time PCR Assay to Detect Plasmodiophora brassicae Under Field Conditions
8.8.4.1 In China
8.8.5 Bioassay Technique of Wallenhammar et al. (2012)
8.8.5.1 The DNA Extraction from Infected Roots, Fungal Material, and Plant Material
8.8.5.2 The DNA Extraction from Soil
8.8.5.3 Primers and Probes
8.8.5.4 Real-Time PCR
8.8.5.5 Quantification
8.8.5.6 Evaluation of Inhibition and Repeatability
8.8.5.7 Distribution of Plasmodiophora brassicae Inoculum and Risk Assessment
8.8.6 Bioassay Technique of Jin-ping et al. (2013)
8.8.6.1 The DNA Extraction from Infected Roots, Fungal Material, Bacterial Material, and Soil
8.8.6.2 Real-Time PCR Assay
8.8.6.3 Sensitivity and Reproducibility of Real-Time PCR
8.8.6.4 The Detection of Artificially and Naturally Infested Soils
8.8.7 Identification of Pathotype-Specific Genes by PCR and Quantitative PCR
8.8.8 Quantification and Control Assessment of Clubroot with qPCR
8.8.9 Assessment of the Response of Lime-Based Fertilizers with qPCR
8.8.10 The PCR and Quantitative PCR Analysis of Infested Soil Samples
8.8.11 Gene Expression (Transcriptomic) Analysis
8.8.12 The qPCR-Based Assessment of Plasmodiophora Spread in Soil
8.8.12.1 Soil Sampling
8.8.12.2 DNA Extraction from Soil
8.8.12.3 Real-Time qPCR
8.8.12.4 Quantification of Plasmodiophora brassicae Spores
8.8.12.5 Cultivation Guidelines Based on qPCR Assay
8.8.12.6 Statistical Analysis and Data Integration
8.8.13 The Plasmodiophora brassicae Genome Sequencing and Transcriptomic Analysis
8.8.13.1 The DNA and RNA Extraction
8.8.13.2 Genome Sequencing and Annotation
8.8.13.3 The RNA-Sequencing and Transcriptomic Analysis
8.8.13.4 Hormonal Signature Analysis
8.8.13.5 Detection of SNPs and Indels
8.9 Brassica-Sclerotinia Pathogenesis Protocols
8.9.1 Transcriptome Analysis of Brassica-Sclerotinia Pathosystem
8.9.1.1 Plant Material and Fungal Strain
8.9.1.2 Plant Growth and Inoculation
8.9.1.3 The RNA Extraction, Library Preparation, and Sequencing
8.9.1.4 Quality Control and Read Mapping to the Reference Genome
8.9.1.5 Differential Gene Expression Analysis
8.9.1.6 Functional Classification and Enrichment Analyses of DEGs
8.9.1.7 Differential Gene Expression Data Validation
8.9.2 Immunoassay for Early Detection of Sclerotinia sclerotiorum
8.9.3 A Polymerase Chain Reaction (PCR) Assay for the Detection of Inoculum of Sclerotinia sclerotiorum
8.10 Brassica-Turnip Yellow Mosaic Virus (TuMV) Pathogenesis Protocols
8.10.1 Detection of Brassica Viruses by Multiplex Reverse Transcription Polymerase Chain Reaction
8.10.1.1 Plant Material and Recombinant Plasmids
8.10.1.2 RNA Extraction
8.10.1.3 Reverse Transcription and PCR
8.10.1.4 Cloning and Sequencing
8.10.1.5 Primer Design and Optimization of Annealing Temperatures
8.10.1.6 Sensitivity of Multiplex RT-PCR
8.10.1.7 Detection of Virus Genotypes in Field Samples by Multiplex RT-PCR
8.10.2 Quantitative RT-PCR Assay for Detection of BnSGS3 Gene
8.10.3 Biological and Serological Testing for the Presence of Cruciferous Viruses
8.10.3.1 The Turnip Yellow Mosaic Virus (TuMV) Full Genome Sequencing, Recombination, and Phylogenetic Analysis
8.10.3.2 Phytoplasma Identification by 16s rRNA Gene Analyses
8.10.3.3 Phytoplasma Multigene Sequence Analysis
8.10.3.4 Phylogenetic Analysis of Phytoplasma Genes
8.11 Brassica-Verticillium Pathogenesis Protocols
8.11.1 Molecular Characterization of Verticillium Isolates
8.11.1.1 Fungal Isolates
8.11.1.2 The DNA Extraction
8.11.1.3 The PCR Amplification and Cloning
8.11.1.4 The DNA Sequencing and Data Analysis
8.12 Brassica-Xanthomonas Pathogenesis Protocols
8.12.1 Determination of Defensive Metabolites in the Phenylpropanoid Pathways
8.12.1.1 Total Phenolic and Flavonoid Contents
8.12.1.2 Soluble and Insoluble Tannin Contents
8.12.1.3 Proanthocyanidin and Total Hydroxycinnamic Acid (THA) Contents
8.12.1.4 Glutathione and NADPH Redox
8.12.1.5 Isolation of Total RNA and Quantitative Real-Time PCR
8.12.2 Genotyping of Xanthomonas campestris pv. campestris
8.12.2.1 Genomic DNA Extraction
8.12.2.2 Repetitive Element PCR Fingerprinting
8.12.2.3 The Multilocus Sequence Analysis
8.12.2.4 Pathogenicity
8.12.2.5 Statistical Analysis
8.13 Brassica-Mixed Infection Diagnosis Protocols
8.13.1 Multispectral Imaging for Presymptomatic Identification of Cruciferous Diseases
8.13.2 The RAPD-Based Molecular Diagnosis of Mixed Infections
References
9: Future Research Priorities of Crucifers´ Host-Pathosystem
9.1 Introduction
Index