| Acknowledgements | 第7-8页 |
| 摘要 | 第8-10页 |
| Abstract | 第10-12页 |
| List of Abbreviations | 第27-29页 |
| Chapter 1 General introduction | 第29-33页 |
| Chapter 2 Review of literature | 第33-81页 |
| 1. History of Parasitology | 第33页 |
| 2. Helminth Classification | 第33-34页 |
| 3. Gastrointestinal nematode (GIN) | 第34-46页 |
| 3.1. History of H. contortus nomenclature | 第35页 |
| 3.2. Taxonomy | 第35-36页 |
| 3.3. Haemonchus contortus life cycle | 第36-39页 |
| 3.4. Hypobiosis | 第39-40页 |
| 3.5. Pathogenesis | 第40-42页 |
| 3.6. Common diagnostic techniques-determining how "wormy" sheep are! | 第42-46页 |
| 3.6.1. FAMACHA(?) eye color chart system | 第42-44页 |
| 3.6.2. Body condition scores | 第44-45页 |
| 3.6.3. Modified McMaster egg counting for nematode egg quantitation | 第45-46页 |
| 3.6.4. Packed cell volume | 第46页 |
| 4. Immune response to GIN parasites | 第46-48页 |
| 4.1. Ovine immunity | 第46-47页 |
| 4.2. Host response to Haemonchus infection | 第47-48页 |
| 5. Factors affecting host response to parasitic infection | 第48-49页 |
| 6. Distribution and importance | 第49页 |
| 7. Ruminants (pregastric and postgastric microbial action) | 第49-51页 |
| 8. History of Sheep | 第51-52页 |
| 9. Characteristics of Sheep | 第52页 |
| 10. Economic importance of sheep | 第52页 |
| 11. Sheep versus goats | 第52-54页 |
| 12. Abomasal anatomy | 第54-57页 |
| 13. Pathophysiology | 第57-58页 |
| 14. Microbial communities | 第58-73页 |
| 14.1. The necessities of microbial world | 第58-59页 |
| 14.2. History and technology evolution | 第59页 |
| 14.3. Sanger sequencing and next generation sequencing (or high-throughput sequencing) | 第59-60页 |
| 14.4. The different sequencing strategies | 第60页 |
| 14.5. Amplicon sequencing | 第60-65页 |
| 14.5.1. 16S rRNA gene sequencing | 第60-65页 |
| 14.6. Factors affecting gastrointestinal tract (GIT) microbiota | 第65-67页 |
| 14.7. Microbial community associated with eukaryotes | 第67-73页 |
| 14.7.1. Symbiotic bacteria | 第68-71页 |
| 14.7.2. Bacteria associated with nematodes | 第71-73页 |
| 15. RNA sequencing | 第73-81页 |
| 15.1. The need for RNA-seq | 第73-74页 |
| 15.2. RNA Sequencing-tool for gene expression analysis | 第74-76页 |
| 15.3. Transcriptional Profiling | 第76-77页 |
| 15.4. RNA-seq experiment, data generation, and analysis | 第77-81页 |
| 15.4.1. RNA extraction | 第77页 |
| 15.4.2. Library preparation | 第77-78页 |
| 15.4.3. Clustering and sequencing | 第78页 |
| 15.4.4. Data processing and quality control | 第78页 |
| 15.4.5. Differential Gene Expression (DGE) Analysis | 第78页 |
| 15.4.6. Functional annotation enrichment analyses | 第78页 |
| 15.4.7. Quantitative reverse transcriptase-PCR (RT PCR) analysis for validation of RNA-seq results | 第78-81页 |
| Chapter 3 Nucleic acid protocols: Extraction and optimization | 第81-95页 |
| 1. Introduction | 第81-82页 |
| 2. Materials and Methods | 第82-88页 |
| 2.1. Reagents | 第82页 |
| 2.2. Equipments | 第82页 |
| 2.3. Reagent setup | 第82页 |
| 2.4. Procedure | 第82-88页 |
| 2.4.1. Grinding in liquid nitrogen (Mortar and pestle) | 第82-84页 |
| 2.4.2. Repetitive pipetting: | 第84-86页 |
| 2.4.3. Glass-bead grinding | 第86-88页 |
| 3. Results and Discussion | 第88-95页 |
| 3.1. Eukaryotes,such as Eimeria spp | 第88页 |
| 3.2. Eukaryotes, such as BALB/c mice | 第88-89页 |
| 3.3. Prokaryotic examples; E. Coli | 第89-95页 |
| Chapter 4 Electrical cream separator coupled with vacuum filtration for the purification of eimerian oocysts and trichostrongylid eggs | 第95-115页 |
| 1. Introduction | 第95-97页 |
| 2. Materials and methods | 第97-108页 |
| 2.1. Eimerian oocyst propagation | 第97-98页 |
| 2.2. Seed-and- recovery experiment for validation of the current method by using different irrevelant matrices | 第98页 |
| 2.3. Trichostrongylid egg preparation | 第98-99页 |
| 2.4. Purification methods | 第99-108页 |
| 2.4.1. Oocyst and egg concentration by the electrical cream separator machine | 第99-100页 |
| 2.4.2. Cream separator machine assembling procedure | 第100-101页 |
| 2.4.3. Procedural steps for coping with the egg-/oocyst- containing faecal material | 第101页 |
| 2.4.4. Vacuum filtration for residual faecal material removal | 第101-102页 |
| 2.4.5. Cream separator machine coupled with vacuum filtration procedure | 第102-104页 |
| 2.4.6. Oocyst and egg separation on a discontinuous sucrose gradient | 第104-107页 |
| 2.4.7. Recovery rate | 第107页 |
| 2.4.8. Viability assay | 第107页 |
| 2.4.9. Optical microscopic observation | 第107-108页 |
| 2.4.10. Statistical analysis | 第108页 |
| 3. Results | 第108-112页 |
| 3.1. Cream separator machine coupled with vacuum filtration for separation of oocysts and eggs | 第108页 |
| 3.2. Optimal dilution ratio, and minimal and maximal recovery rate determination | 第108-109页 |
| 3.3. Effect of diverse extraneous matrices on oocysts' recovery efficiency | 第109页 |
| 3.4. Viability assay | 第109-110页 |
| 3.5. Combined sucrose step-density gradient centrifugation-vacuum filtration methods for oocyst and egg isolation | 第110-111页 |
| 3.5.1. Maximal and minimal recovery rates | 第110-111页 |
| 3.5.2. Viability assay | 第111页 |
| 3.5.3. Microscopic examination of concentrated and recovered coccidian oocysts and nematode eggs | 第111页 |
| 3.6. Comparison of milk cream separator with that of the modified sucrose gradient | 第111-112页 |
| 4. Discussion | 第112-115页 |
| Chapter 5 Gastric microbial community and ovine host response varies with early and late stages of Haemonchus contortus infection | 第115-133页 |
| 1. Introduction | 第115-117页 |
| 2. Materials and Methods | 第117-121页 |
| 2.1. Animals and parasites | 第117页 |
| 2.2. Parasite infections | 第117-118页 |
| 2.3. Abomasal histopathologic examination | 第118页 |
| 2.4. Ruminal papilla counting | 第118页 |
| 2.5. Omasal fold counting | 第118页 |
| 2.6. PH value measurements | 第118页 |
| 2.7. Sample collection, DNA extraction, amplification, and processing of samples for high-throughput sequencing | 第118-120页 |
| 2.7.1. Sample collection | 第119页 |
| 2.7.2. Microbial DNA extraction | 第119页 |
| 2.7.3. Amplicon generation and high-throughput sequencing | 第119-120页 |
| 2.8. Statistical and bioinformatics analysis | 第120-121页 |
| 3. Results | 第121-129页 |
| 3.1. Haemonchus infection-associated histopathologic changes of the abomasum | 第121-122页 |
| 3.2. Haemonchus infection-associated microhabitat changes | 第122-123页 |
| 3.3. Haemonchus infection-associated abomasal and ruminal microbial changes | 第123-129页 |
| 3.3.1. The 16S rRNA amplicon metagenomic analysis of abomasal/ruminal bacteria | 第123-126页 |
| 3.3.2. Diversity index analysis of microbial community | 第126-127页 |
| 3.3.3. The analysis of microbial community structure and dominant taxa | 第127-129页 |
| 4. Discussion | 第129-133页 |
| Chapter 6 Exploring the microbial community (microflora) associated with ovine Haemonchus contortus (macroflora) field strains | 第133-153页 |
| 1. Introduction | 第133-134页 |
| 2. Materials and methods | 第134-139页 |
| 2.1. Animals and parasites | 第134-135页 |
| 2.2. Ruling out bacterial contamination associated with the parasite surfaces | 第135-136页 |
| 2.3. DNA extraction, amplification and processing of samples for high-throughput sequencing | 第136-138页 |
| 2.3.1. Microbial DNA extraction | 第136页 |
| 2.3.2. Amplicon generation and high-throughput sequencing | 第136-138页 |
| 2.4. Statistical and bioinformatics analysis | 第138-139页 |
| 3. Results | 第139-151页 |
| 3.1. Parasitological tests, FAMACHA(?) scores and body condition scoring | 第139页 |
| 3.2. Morphological forms of the ovine H. contortus life-cycle stages | 第139-140页 |
| 3.3. Investigation of parasite surfaces for abomasal microbial contamination | 第140页 |
| 3.4. Larval-stage of H. contortus displaying internalized transgenic Escherichia coli | 第140-141页 |
| 3.5. Diversity index analysis of microbial community | 第141-147页 |
| 3.5.1. Alpha rarefaction curves | 第141-146页 |
| 3.5.2. Beta diversity | 第146-147页 |
| 3.6. The microbial community composition of the ovine abomasum and rumen in the context of H. contortus infection at the genus level | 第147页 |
| 3.7. The analysis of microbial community structure and dominant taxa associated with distinct life-cycle stages of H. contortus | 第147-151页 |
| 3.8. The analysis of microbial community structure associated with individual adult female worms of H. contortus | 第151页 |
| 4. Discussion | 第151-153页 |
| Chapter 7 Early and late gene expression profiles of the ovine mucosa in response to Haemonchus contortus infection employing Illumina RNA-seq technology | 第153-177页 |
| 1. Introduction | 第153-154页 |
| 2. Materials and Methods | 第154-158页 |
| 2.1. Animals and parasites | 第154-155页 |
| 2.2. Ovis aries genome reference for mapping reads | 第155页 |
| 2.3. Parasite infections | 第155-156页 |
| 2.3.1. Worm recovery procedures | 第155页 |
| 2.3.2. Faecal nematode egg counts | 第155-156页 |
| 2.4. RNA seq experiment | 第156-158页 |
| 2.4.1. RNA extraction | 第156页 |
| 2.4.2. Library preparation | 第156页 |
| 2.4.3. Clustering and sequencing | 第156-157页 |
| 2.4.4. Data processing and quality control | 第157页 |
| 2.4.5. Differential Gene Expression (DGE) Analysis | 第157页 |
| 2.4.6. Functional annotation enrichment analyses | 第157页 |
| 2.4.7. Quantitative Reverse Transcriptase-PCR (RT PCR) Analysis | 第157-158页 |
| 3. Results | 第158-172页 |
| 3.1. Parasitological data | 第158页 |
| 3.2. RNA-seq data quality control | 第158-159页 |
| 3.3. Massively sequencing and aligning to the reference genome | 第159-161页 |
| 3.4. RNA-seq analysis reveals distinct gene expression signatures among groups | 第161-164页 |
| 3.5. Further analysis of the immune Repertoire | 第164页 |
| 3.6. Selected genes with diverse functions, such as Gal-15,-4 and SPP1 for further analysis | 第164-169页 |
| 3.7. GO and KEGG enrichment analyses of differentially expressed genes | 第169-170页 |
| 3.8. Validation of transcriptome results by real-time PCR | 第170-172页 |
| 4. Discussion | 第172-177页 |
| Chapter 8 An ex vivo abomasal ovine model to study the immediate immune response in the context of Haemonchus contortus larval-stage | 第177-203页 |
| 1. Introduction | 第177-179页 |
| 2. Materials and Methods | 第179-183页 |
| 2.1. Animals and parasites | 第179页 |
| 2.2. Ovis aries genome reference for mapping reads | 第179页 |
| 2.3. Preparation of haemonchine larval inoculum | 第179页 |
| 2.4. Preparation of ovine gastric content supplement | 第179-180页 |
| 2.5. Preparation of media | 第180页 |
| 2.6. Ovine abomasal explant preparation and incubation | 第180页 |
| 2.7. Histological studies of the ovine abomasal explants | 第180页 |
| 2.8. Cell cytotoxicity assay | 第180-181页 |
| 2.9. RNA seq experiment | 第181-183页 |
| 2.9.1. RNA extraction | 第181页 |
| 2.9.2. Library preparation | 第181-182页 |
| 2.9.3. Clustering and sequencing | 第182页 |
| 2.9.4. Data processing and quality control | 第182页 |
| 2.9.5. Differential Gene Expression (DGE) Analysis | 第182页 |
| 2.9.6. Functional annotation enrichment analyses | 第182页 |
| 2.9.7. Quantitative Reverse Transcriptase-PCR (RT PCR) Analysis | 第182-183页 |
| 3. Results | 第183-199页 |
| 3.1. Histomorphology of the ovine abomasal explants | 第183-185页 |
| 3.2. Cell cytotoxicity assay | 第185页 |
| 3.3. RNA-seq data quality controL | 第185-188页 |
| 3.4. Massively sequencing and aligning to the reference genome | 第188-191页 |
| 3.5. RNA-seq analysis reveals distinct gene expression signatures between explants | 第191-196页 |
| 3.6. Further analysis of the immune Repertoire | 第196页 |
| 3.7. Go and KEGG enrichment analyses of differentially expressed genes | 第196-198页 |
| 3.8. Validation of transcriptome results by real-time PCR | 第198-199页 |
| 4. Discussion | 第199-203页 |
| Chapter 9 Innovation and General conclusion | 第203-205页 |
| References | 第205-251页 |
| Additional information | 第251-497页 |
| 1. Supplementary Information (Chapter 5) | 第253-302页 |
| 2. Supplementary information (Chapter 6) | 第302-333页 |
| 3. Supplementary information (Chapter 7) | 第333-455页 |
| 4. Supplementary information (Chapter 8) | 第455-497页 |
| Publications and achievements | 第497-500页 |
| Publications | 第497-499页 |
| International Patent | 第499页 |
| Awards | 第499-500页 |
| Editorial Board Members | 第500页 |
| 1. American Research Journal of Biotechnology | 第500页 |
| 2. International Journal of Agricultural Research and Crop Sciences Eidtor Board | 第500页 |
| 3. Agrotechnology Journal | 第500页 |
| 4. Palliative Medicine and Nursing | 第500页 |
