In vivo testing of wild type and attenuated salmonid alphaviruses with and without a GFP reporter gene
1. Purpose
The main goal of this study is to use GFP-reporter viruses to study salmonid alphavirus (SAV3) infection in Atlantic salmon fry. The use of fluorescent reporter viruses will allow us to study the route of entry and spread of the virus throughout the body of the fish.
2. Distress
SAV3 infection in salmon fry of this size usually gives a very high mortality. Signs of illness include a decrease in food intake and changes in swimming behavior. The fish will be monitored several times each day for signs of illness, and sick fish will be removed from the tanks and humanly euthanized.
3. Expected benefit
The development of biological reagents that will help to better understand the effects of SAV3 infection in Atlantic salmon. In addition, we hope that fluorescent imaging of live fish at various timepoints during the experiment will identify better humane endpoints that can be applied in subsequent experiments.
4. Number of animals and what kind
This experiment will use 400 Atlantic salmon fry
5. How to adhere to the 3Rs
Replacement: Much of the initial works with these recombinant GFP viruses has been done in cell lines. However, in order to study viral entry, replication, and spread in one of its natural host species, experiments with fish now have to be performed. It is not yet possible to study these aspects of SAV3 biology in cell culture systems.
Reduction: We have designed the experiment to minimize the number of fish used.
Refinement: The fish will be kept in approved facilities and will be handled by well-trained personnel according to our best-practice SOPs and routines. We have developed good procedures and methods for infection experiments using salmon fry. The fish will be anesthetized before any injections, and time out of water will be minimized. The fish will be monitored several times every day by trained personnel and moribund and sick fish will be removed and humanely euthanized.
The main goal of this study is to use GFP-reporter viruses to study salmonid alphavirus (SAV3) infection in Atlantic salmon fry. The use of fluorescent reporter viruses will allow us to study the route of entry and spread of the virus throughout the body of the fish.
2. Distress
SAV3 infection in salmon fry of this size usually gives a very high mortality. Signs of illness include a decrease in food intake and changes in swimming behavior. The fish will be monitored several times each day for signs of illness, and sick fish will be removed from the tanks and humanly euthanized.
3. Expected benefit
The development of biological reagents that will help to better understand the effects of SAV3 infection in Atlantic salmon. In addition, we hope that fluorescent imaging of live fish at various timepoints during the experiment will identify better humane endpoints that can be applied in subsequent experiments.
4. Number of animals and what kind
This experiment will use 400 Atlantic salmon fry
5. How to adhere to the 3Rs
Replacement: Much of the initial works with these recombinant GFP viruses has been done in cell lines. However, in order to study viral entry, replication, and spread in one of its natural host species, experiments with fish now have to be performed. It is not yet possible to study these aspects of SAV3 biology in cell culture systems.
Reduction: We have designed the experiment to minimize the number of fish used.
Refinement: The fish will be kept in approved facilities and will be handled by well-trained personnel according to our best-practice SOPs and routines. We have developed good procedures and methods for infection experiments using salmon fry. The fish will be anesthetized before any injections, and time out of water will be minimized. The fish will be monitored several times every day by trained personnel and moribund and sick fish will be removed and humanely euthanized.