中草藥（TCM）和芽孢桿菌（BS）對(duì)尼羅羅非魚生長(zhǎng)、免疫應(yīng)答、抗病力及耐受性的影響
發(fā)布時(shí)間：2022-01-02 23:32
當(dāng)前,羅非魚養(yǎng)殖面臨的主要問(wèn)題是其對(duì)病原體感染的敏感性如無(wú)乳鏈球菌,以及環(huán)境脅迫因素,如缺氧、冷和熱應(yīng)激等。目前研究表明,天然免疫刺激劑的使用對(duì)魚類具有增強(qiáng)其免疫力和提高其生存環(huán)境耐受性的潛力。本研究對(duì)中草藥（TCM,黃芪、當(dāng)歸和山楂混合比例為1:1:1 w/w）、芽孢桿菌組合（BS,枯草芽孢桿菌和地衣芽孢桿菌混合比例為1:1 w/w）及中草藥-芽孢桿菌聯(lián)合（TCMBS）的功效進(jìn)行探索,獲得TCM、BS及TCMBS最佳添加劑量,并將其作為羅非魚養(yǎng)殖中潛在的免疫增強(qiáng)劑以確定它們對(duì)改善羅非魚生長(zhǎng)和免疫的作用,其研究結(jié)果為更好地了解中草藥和芽孢桿菌在改善羅非魚生長(zhǎng)和免疫方面的作用機(jī)制奠定基礎(chǔ)。該研究用BS、TCM和TCMBS分別投喂羅非魚,測(cè)定其生長(zhǎng)指標(biāo)和免疫指標(biāo),其投喂配方如下:在單種投喂組（TCM或BS）實(shí)驗(yàn)中,魚投喂含有10 g kg-1 TCM的飼料或3 g kg-1（BS3）、5 g kg-1（BS5）、7 g kg-1（BS7）和10 g kg-1（BS10）的飼料;在T...
【文章來(lái)源】：廣東海洋大學(xué)廣東省
【文章頁(yè)數(shù)】：156 頁(yè)
【學(xué)位級(jí)別】：博士
【文章目錄】：
Abstract
摘要
1 General Introduction
1.1 Background of Study
1.2 Literature Review on Immunostimulant Effects in Fish
1.2.1 Effects of Immunostimulant on Fish Growth
1.2.2 Duration of Immunostimulants Administration
1.2.3 Humoral Indices
1.2.4 Stress Response in Fish
1.2.5 Hepatic Parameters
1.2.6 Expression of Immune-Related Genes
1.2.7 Contribution of Immunostimulants on Disease Resistance
1.3 Purpose of Study
1.4 Significance of the Study
2 General Materials and Methods
2.1 Technical Routes of Study
2.2 Preparation of Experimental Diets
2.3 Fish Management
2.4 Growth Performance
2.5 Sample Collection and Preparation
2.6 Analysis of Humoral Immune Parameters
2.6.1 Lysozyme(LYZ)
2.6.2 Protease Activity
2.6.3 Antiprotease Activity
2.6.4 Immunoglobulin M(IgM)
2.6.5 Superoxide dismutase(SOD)Activity Assay
2.6.6 Catalase Activity(CAT)
2.6.7 Assessment of Hepatic Parameters
2.6.8 Alkaline Phosphatase(ALP)activity
2.6.9 Total Antioxidant Capacity(T-AOC)
2.7 Gene Expression
2.7.1 RNA Extraction and cDNA Synthesis
2.7.2 Quantitative Real-Time Polymerase Chain Reaction(qRT-PCR)
2.8 Challenge Experiment
2.8.1 Preparation of Inoculum
2.8.2 Calculation of50%Endpoint(Lethal Dose50)
2.8.3 Infection Test
2.9 Statistical Analysis
3 Effect of BS on Growth,Immune Response,and Resistance Against Streptococcus Agalactiae of Nile Tilapia
3.1 Introduction
3.2 Materials and Methods
3.2.1 Diet Preparation
3.2.2 Experimental Set-up and Fish Management
3.2.3 Growth Performance
3.2.4 Probiotic Viability in Feed and Intestines of Tilapia
3.2.5 Sample Collection
3.2.6 Analysis of Serum Immune Parameters
3.2.7 Gene Expression
3.2.8 Challenge Test
3.2.9 Statistical Analysis
3.3 Results
3.3.1 Response to Feed Intake and Growth Performance
3.3.2 Viability of Probiotic BS in Diets and Intestines
3.3.3 Humoral Immune Response
3.3.4 Gene Expression
3.3.5 Challenge Test
3.4 Discussion
4 Effects of TCM on the Growth,Immune Responses,and Resistance Against Streptococcus Agalactiae of Nile Tilapia
4.1 Introduction
4.2 Materials and Methods
4.2.1 Experimental Diets
4.2.2 Experimental Set-up and Fish Management
4.2.3 Growth Parameters
4.2.4 Blood and Tissue Sample Collection
4.2.5 Analysis of Serum Immune Parameters
4.2.6 Gene Expression
4.2.7 Challenge Test
4.2.8 Statistical Analysis
4.3 Results
4.3.1 Growth Performance
4.3.2 Hepatoprotective Effect TCM Supplementation
4.3.3 Immune Response
4.3.4 Challenge Test
4.4 Discussion
5 Effects of TCMBS on the Growth,Immune Response,and Resistance Against Streptococcus Agalactiae of Nile Tilapia
5.1 Introduction
5.2 Materials and Methods
5.2.1 Experimental Diet
5.2.2 Experimental Set-up and Fish Management
5.2.3 Blood and Tissue Sample Collection
5.2.4 Growth Performance and Morphometric Indexes
5.2.5 Analysis of Serum Immune Parameters
5.2.6 Gene Expression
5.2.7 Challenge Test
5.2.8 Statistical Analysis
5.3 Results
5.3.1 Growth Performance and Morphometric Indexes
5.3.2 Humoral Immune Parameters
5.3.3 Expression of Immune-related Genes
5.3.4 Infection Test
5.4 Discussion
6 Effects of TCMBS Application on Skin Immunity of Nile Tilapia
6.1 Introduction
6.2 Materials and Methods
6.2.1 Experimental Diet
6.2.2 Experimental Set-up and Fish Management
6.2.3 Sample Collection
6.2.4 Analysis of Skin Mucus Physico-chemical Parameters
6.2.5 Analysis of Skin Mucus Immune Parameters
6.2.6 Gene Expression
6.2.7 Bactericidal Activity
6.2.8 Statistical Analysis
6.3 Results
6.3.1 Physico-chemical Parameters in Skin Mucus
6.3.2 Enzyme Activities in Skin Mucus
6.3.3 Bactericidal Activity of Skin Mucus
6.3.4 Description of Gene Expression in Skin
6.4 Discussion
7 Modulation of NADPH Oxidase Genes in Neutrophils Following Stimulation by TCMBS3 in Nile tilapia
7.1 Introduction
7.2 Materials and Methods
7.2.1 Preparation of Diets
7.2.2 Experimental Set-up and Fish Management
7.2.3 Tissue Collection
7.2.4 Neutrophil Isolation
7.2.5 ROS Test
7.2.6 RNA Extraction,cDNA Synthesis and qRT-PCR Analysis
7.2.7 Cloning and Sequencing of the Tilapia NADPH oxidase Genes
7.2.8 Sequence Analysis
7.2.9 Infection test
7.2.10 Statistical Analysis
7.3 Results
7.3.1 Characterization of NADPH Oxidase Genes
7.3.2 Evolutional Relationship of Tilapia NADPH Oxidase Genes
7.3.3 Tissue Expression of NADPH Oxidase Genes
7.3.4 TCMBS Induces the Expression of NADPH Oxidase Genes in Splenic Neutrophils
7.3.5 ROS production
7.3.6 Expression Analysis of Antioxidant Genes
7.4 Discussion
8 Effects of TCM and BS on the Stress Tolerance of Nile tilapia
8.1 Introduction
8.2 Materials and Methods
8.2.1 Experimental Diet
8.2.2 Experimental Set-up and Fish Management
8.2.3 Time point I:Pre-exposure
8.2.4 Time point II:Stress Exposure
8.2.5 Sample Collection and Measurement
8.2.6 Gene Expression
8.2.7 Statistical Analysis
8.3 Results
8.3.1 ffects of Hypoxia,Cold,and Heat Stressors on Hemato-Immunological Parameters
8.3.2 Effects of Hypoxia,Cold,and Heat Stressors on Stress Parameters
8.3.3 Effects of Hypoxia,Cold,and Heat Stressors on Tissue Damage Parameters
8.3.4 Effects of Hypoxia,Cold,and Heat Stressors on Antioxidant Parameters
8.3.5 Effects of Hypoxia,Cold,and Heat Stressors on the Expression of HSP70 and HIF-1αmRNAs
8.4 Discussion
9 General Conclusions and Recommendations
9.1 Conclusions
9.2 Recommendations
References
Supplementary Table
List of Abbreviations
Acknowledgement
Authors’Profile
Supervsiors’Profile
本文編號(hào)：3565101
【文章來(lái)源】：廣東海洋大學(xué)廣東省
【文章頁(yè)數(shù)】：156 頁(yè)
【學(xué)位級(jí)別】：博士
【文章目錄】：
Abstract
摘要
1 General Introduction
1.1 Background of Study
1.2 Literature Review on Immunostimulant Effects in Fish
1.2.1 Effects of Immunostimulant on Fish Growth
1.2.2 Duration of Immunostimulants Administration
1.2.3 Humoral Indices
1.2.4 Stress Response in Fish
1.2.5 Hepatic Parameters
1.2.6 Expression of Immune-Related Genes
1.2.7 Contribution of Immunostimulants on Disease Resistance
1.3 Purpose of Study
1.4 Significance of the Study
2 General Materials and Methods
2.1 Technical Routes of Study
2.2 Preparation of Experimental Diets
2.3 Fish Management
2.4 Growth Performance
2.5 Sample Collection and Preparation
2.6 Analysis of Humoral Immune Parameters
2.6.1 Lysozyme(LYZ)
2.6.2 Protease Activity
2.6.3 Antiprotease Activity
2.6.4 Immunoglobulin M(IgM)
2.6.5 Superoxide dismutase(SOD)Activity Assay
2.6.6 Catalase Activity(CAT)
2.6.7 Assessment of Hepatic Parameters
2.6.8 Alkaline Phosphatase(ALP)activity
2.6.9 Total Antioxidant Capacity(T-AOC)
2.7 Gene Expression
2.7.1 RNA Extraction and cDNA Synthesis
2.7.2 Quantitative Real-Time Polymerase Chain Reaction(qRT-PCR)
2.8 Challenge Experiment
2.8.1 Preparation of Inoculum
2.8.2 Calculation of50%Endpoint(Lethal Dose50)
2.8.3 Infection Test
2.9 Statistical Analysis
3 Effect of BS on Growth,Immune Response,and Resistance Against Streptococcus Agalactiae of Nile Tilapia
3.1 Introduction
3.2 Materials and Methods
3.2.1 Diet Preparation
3.2.2 Experimental Set-up and Fish Management
3.2.3 Growth Performance
3.2.4 Probiotic Viability in Feed and Intestines of Tilapia
3.2.5 Sample Collection
3.2.6 Analysis of Serum Immune Parameters
3.2.7 Gene Expression
3.2.8 Challenge Test
3.2.9 Statistical Analysis
3.3 Results
3.3.1 Response to Feed Intake and Growth Performance
3.3.2 Viability of Probiotic BS in Diets and Intestines
3.3.3 Humoral Immune Response
3.3.4 Gene Expression
3.3.5 Challenge Test
3.4 Discussion
4 Effects of TCM on the Growth,Immune Responses,and Resistance Against Streptococcus Agalactiae of Nile Tilapia
4.1 Introduction
4.2 Materials and Methods
4.2.1 Experimental Diets
4.2.2 Experimental Set-up and Fish Management
4.2.3 Growth Parameters
4.2.4 Blood and Tissue Sample Collection
4.2.5 Analysis of Serum Immune Parameters
4.2.6 Gene Expression
4.2.7 Challenge Test
4.2.8 Statistical Analysis
4.3 Results
4.3.1 Growth Performance
4.3.2 Hepatoprotective Effect TCM Supplementation
4.3.3 Immune Response
4.3.4 Challenge Test
4.4 Discussion
5 Effects of TCMBS on the Growth,Immune Response,and Resistance Against Streptococcus Agalactiae of Nile Tilapia
5.1 Introduction
5.2 Materials and Methods
5.2.1 Experimental Diet
5.2.2 Experimental Set-up and Fish Management
5.2.3 Blood and Tissue Sample Collection
5.2.4 Growth Performance and Morphometric Indexes
5.2.5 Analysis of Serum Immune Parameters
5.2.6 Gene Expression
5.2.7 Challenge Test
5.2.8 Statistical Analysis
5.3 Results
5.3.1 Growth Performance and Morphometric Indexes
5.3.2 Humoral Immune Parameters
5.3.3 Expression of Immune-related Genes
5.3.4 Infection Test
5.4 Discussion
6 Effects of TCMBS Application on Skin Immunity of Nile Tilapia
6.1 Introduction
6.2 Materials and Methods
6.2.1 Experimental Diet
6.2.2 Experimental Set-up and Fish Management
6.2.3 Sample Collection
6.2.4 Analysis of Skin Mucus Physico-chemical Parameters
6.2.5 Analysis of Skin Mucus Immune Parameters
6.2.6 Gene Expression
6.2.7 Bactericidal Activity
6.2.8 Statistical Analysis
6.3 Results
6.3.1 Physico-chemical Parameters in Skin Mucus
6.3.2 Enzyme Activities in Skin Mucus
6.3.3 Bactericidal Activity of Skin Mucus
6.3.4 Description of Gene Expression in Skin
6.4 Discussion
7 Modulation of NADPH Oxidase Genes in Neutrophils Following Stimulation by TCMBS3 in Nile tilapia
7.1 Introduction
7.2 Materials and Methods
7.2.1 Preparation of Diets
7.2.2 Experimental Set-up and Fish Management
7.2.3 Tissue Collection
7.2.4 Neutrophil Isolation
7.2.5 ROS Test
7.2.6 RNA Extraction,cDNA Synthesis and qRT-PCR Analysis
7.2.7 Cloning and Sequencing of the Tilapia NADPH oxidase Genes
7.2.8 Sequence Analysis
7.2.9 Infection test
7.2.10 Statistical Analysis
7.3 Results
7.3.1 Characterization of NADPH Oxidase Genes
7.3.2 Evolutional Relationship of Tilapia NADPH Oxidase Genes
7.3.3 Tissue Expression of NADPH Oxidase Genes
7.3.4 TCMBS Induces the Expression of NADPH Oxidase Genes in Splenic Neutrophils
7.3.5 ROS production
7.3.6 Expression Analysis of Antioxidant Genes
7.4 Discussion
8 Effects of TCM and BS on the Stress Tolerance of Nile tilapia
8.1 Introduction
8.2 Materials and Methods
8.2.1 Experimental Diet
8.2.2 Experimental Set-up and Fish Management
8.2.3 Time point I:Pre-exposure
8.2.4 Time point II:Stress Exposure
8.2.5 Sample Collection and Measurement
8.2.6 Gene Expression
8.2.7 Statistical Analysis
8.3 Results
8.3.1 ffects of Hypoxia,Cold,and Heat Stressors on Hemato-Immunological Parameters
8.3.2 Effects of Hypoxia,Cold,and Heat Stressors on Stress Parameters
8.3.3 Effects of Hypoxia,Cold,and Heat Stressors on Tissue Damage Parameters
8.3.4 Effects of Hypoxia,Cold,and Heat Stressors on Antioxidant Parameters
8.3.5 Effects of Hypoxia,Cold,and Heat Stressors on the Expression of HSP70 and HIF-1αmRNAs
8.4 Discussion
9 General Conclusions and Recommendations
9.1 Conclusions
9.2 Recommendations
References
Supplementary Table
List of Abbreviations
Acknowledgement
Authors’Profile
Supervsiors’Profile
本文編號(hào)：3565101
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