Inactivated vaccines for tilapia
Front matter
1
Introduction
Background
Freshwater cultured fish are prone to microbial pathogens
2
Research context
3
Project organisation
3.1
Conceptual framework
3.2
Split of the work in workpackages
3.3
Workflow
3.4
Experimental designs
Vaccination
Sampling
3.5
Timelines
4
Pond, fish and husbandry
4.1
Aquaria and ponds
4.1.1
Size and Shape
4.1.2
Water parameters
4.2
Fish
4.3
Feeding regime
4.4
Acclimatation
5
Bacterial isolation
5.0.1
Extraction of fish tissues
6
Bacterial culture
6.1
Principles of bacterial culture
6.1.1
Generation of microbial growth-curves according to OD600 (λ=600 nm)
6.1.2
Live and dead cells counts using hemocytometer under light microscopy
6.2
Bacterial strains
6.2.1
Aeromonas veronii
6.2.2
Streptoccocus agalactiae
6.3
Bacterial culture
6.3.1
Laboratory settings
6.3.2
Recovery of strains
6.4
Standard A.veronii
6.5
Standard S.agalactiae
7
Formalin-killed vaccines
7.1
Production of vaccine Sa
7.2
Production of vaccine Av
7.3
Production of vaccine Sa + Av
7.4
Production of vaccine Control
8
Sample collection and Immunization
8.1
Sample collection
8.1.1
Blood collection
8.1.2
Mucus collection
8.1.3
Liver tissue collection
8.1.4
Head kidneys tissue collection
8.2
Immunization 1 (day 0)
8.3
Immunization 2 (day 21)
9
Lab assays, immune responses
9.1
ELISA for IgM and IgT
9.2
PCR of key immune genes
9.2.1
Normalization of gene expression for RT-PCR
9.3
Agglutination test
9.3.1
Qualitative detection of the presence of antibodies by agglutination
9.3.2
Quantitative antibody agglutination titration
9.4
Collect and analyze results
10
Challenge tests
10.1
Bacterial challenge 1 (day 0)
10.2
Bacterial challenge 2 (day 28)
11
Results
11.1
Gene expression
11.1.1
Liver
11.1.2
Headkidney
11.2
Systemic antibody response
11.2.1
Monovalent vaccines
11.2.2
Bivalent vaccine
11.3
Mucosal antibody response
11.3.1
Monovalent vaccines
11.3.2
Bivalent vaccine
11.4
Challenge test 1 (Unique)
11.4.1
Relative percentage survival (single dose)
11.4.2
Kaplan-Meier survival estimates for the first challenge test
11.5
Challenge test 2 (Booster)
11.5.1
Relative percentage survival (booster)
11.5.2
Kaplan-Meier survival estimates for the second challenge test
12
Discussion
References
Published with bookdown
Systemic and mucosal immune responses of fish Nile tilapia to monovalent and bivalent inactivated vaccines from Streptococcus agalactiae and Aeromonas veronii
Chapter 5
Bacterial isolation
5.0.1
Extraction of fish tissues
Organs in 5 randomly sampled juveniles using TSA/TSB or BHIA