Prepare aquaria, stock fish, acclimate 7 days
T1.1 Prepare the 4 aquaria.
T1.2 Assess population distribution parameters: calculate mean for mass, size and associated standard deviations (if the population is gaussian).
T1.3 Stock 165 juveniles fish of at least 12-13 grams per aquarium.
T1.4 Record water parameter.
T1.5 Define feeding regime.
T1.6 Acclimate 7 days.
Recovery of the two strains S.agalactiae and A.veronii using TSA/TSB for 24h
T2.1 Recovery of the two strains
Read more Wednesday, September 15
Cultivate bacteria 24h, draw growth-curves, count viable and dead cells, estimate biomass with spectrophotometer for λ=600 nm
T2.2 Stockage of bacteria for challenge test
T2.3 Production of vaccine Sa
T2.4 Production of vaccine Av
T2.5 Production of vaccine Sa + Av
T2.6 Production of vaccine Control
Bacterial isolation for different organs in 5 randomly sampled juveniles using TSA/TSB or BHIA
T1.7 Bacterial isolation: Quality control step to prove that the fish population is free of diseases
Read more Saturday, September 18
Mucus and blood swab for baseline week 0, end of acclimatation phase
T3.1 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, September 20
Mucus and blood swab for week 1
T3.3 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, September 27
Mucus and blood swab for week 2
T3.4 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, October 4
Mucus and blood swab for week 3
T3.5 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, October 11
Immunization 2: booster
T3.6 Immunization second dose: booster(Sa , Av , Sa + Av , Control )
Read more Monday, October 11
Mucus and blood swab for week 4
T3.7 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, October 18
Bacterial challenge test 1 (no booster)
T5.1 Expose to Steptoccocus agalactiae the group vaccinated with Sa
T5.2 Expose to Steptoccocus agalactiae the group vaccinated with PBS
T5.3 Expose to Aeromonas veronii the group vaccinated with Av
T5.4 Expose to Steptoccocus agalactiae the group vaccinated with Sa + Av
T5.5 Expose to Aeromonas veronii the group vaccinated with Sa + Av
T5.6 Expose to Aeromonas veronii the group vaccinated with PBS
T5.7 Daily monitoring survival rate
T5.8 Draw conclusions pt.1 Read more Tuesday, October 19
Mucus and blood swab for week 5
T3.8 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, October 25
Mucus and blood swab for week 6
T3.9 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, November 1
Mucus and blood swab for week 7
T3.10 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, November 8
Mucus and blood swab for week 8
T3.11 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, November 15
Mucus and blood swab for week 9
T3.12 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, November 22
Mucus and blood swab for week 10
T3.13 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, November 29
Mucus and blood swab for week 11
T3.14 Weekly fish swabs (to collect biomarkers). Harvest head kidney and liver on 3 fish.
Read more Monday, December 6
Lab assays to monitor immune response
T4.1 ELISA for each group of samples for IgM and IgT
T4.2 PCR of key immune genes
T4.3 Agglutination test
T4.4 Collect and analyze results
T4.5 Draw conclusions
Bacterial challenge test 2
T5.1' Expose to Steptoccocus agalactiae the group vaccinated with Sa
T5.2' Expose to Steptoccocus agalactiae the group vaccinated with PBS
T5.3' Expose to Aeromonas veronii the group vaccinated with Av
T5.4' Expose to Steptoccocus agalactiae the group vaccinated with Sa + Av
T5.5' Expose to Aeromonas veronii the group vaccinated with Sa + Av
T5.6' Expose to Aeromonas veronii the group vaccinated with PBS
T5.7' Daily monitoring survival rate
T5.8' Draw conclusions pt.2 Read more Thrusday, December 9
