Proliferative Kidney Disease (PKD) is a deadly parasitic fish disease that is classified as reportable and that has to be monitored in Switzerland. For this purpose, 15’500 potentially healthy pre-reproductive brown trout were electrofished and euthanized since 2000 to determine their PKD status by the applicant at the Centre for Fish and Wildlife Health (FIWI) alone.
This project aims to establish an alternative, non-lethal and animal-independent monitoring method based on pathogen DNA detection from water samples (an approach also called “eDNA” – environmental DNA). By establishing sensitive and reliable technical methods, and by evaluating them in the field, we will generate spatio-temporal detection probability maps and will provide the necessary expertise and tools for our implementation partners in cantonal and federal fish health and water quality monitoring to replace current monitoring practices.
OUTPUT
How did the project output contribute to the advancement of the 3Rs principle?
The successful implementation of the eDNA method to monitor T. bryosalmonae in the field also furthered replacement in the laboratory setting. In an infection experiment in our fish facility (n° BE47/2023), we replaced lethal sampling of experimental animals (as done previously) by regular monitorings of parasite concentrations in the water (as an indicator of active infection and shedding). This allowed us to use less animals in the experiment, reduce the stress for all animals in the tank entailed by catching, handling and euthanizing animals for monitoring, and allowed us to reach a higher sample size and thus better results in the main experiment.
Do the project’s results have the potential for practical implementation in 3Rs-related research?
Specific Applications: Switzerland uses a national program to monitor river and lake water quality, the National Surface Water Quality Monitoring Program (“NAWA”). NAWA aims to provide an overview of the state of surface waters in Switzerland and to monitor the effectiveness of protection measures, but also to ensure the early detection of problems and threats to aquatic communities. Therefore, assessments of the health status of aquatic animals are currently part of the efforts to reach NAWA’s aims.
The importance of fish health data in political decisions and as success control of management measures is well recognized, but fish health monitoring is currently under debate because of the huge impact on delicate fish populations and the whole ecosystem. Current PKD monitoring a) involves culling of brown trout and electrofishing-induced stress/injury in non-target species, b) is labour intense (approximately nine person-days per location) and c) requires highly educated and experienced staff for electrofishing, fish pathology, histology, and (q)PCR.
eDNA offers the opportunity for a cheaper method (approximately three person-days to sample and analyse water from up to 10 locations) without animal welfare issues. eDNA approaches are already used to monitor the presence of invasive species, but also diseases such as Proliferative Kidney Disease and crayfish plague. However, In-depth adaptations and validation experiments were required before existing tissue-based PCR assays could be applied on eDNA monitoring in the field.
Transferability and Potential Impact: The now established cost-effective, straightforward, and highly sensitive laboratory analysis workflow to detect DNA of T. bryosalmonae from water samples can also be transferred to monitor other water-borne infectious agents, like Aphanomyces astaci (causing crayfish plague), in the field. This would prevent already endangered species, like European brown trout or native European crayfish species) to be sampled for survey reasons.
The eDNA method established in the frame of the project was also already applied in research projects to identify the time point of parasite shedding in PKD infection experiments. In this way, regular sampling of brrwon trout could be replaced by regular sampling of water (e.g. experimental permssion n° BE47/2023, Immuno-physiological changes of brown trout under temperature stress in combination with PKD infection). Therefore, number of fish used in this experiment could be drastically reduced.
Challenges: The eDNA method is not suitable to detect the disease status in the animals, as it only recognizes agent DNA. The effect of the infectious agents on the animal itself can not be judged. There is a high number of infectious agents naturally present in the environment, only causing disaese if the animal host is stressed or if the agent concentration surpasses a special threshold. With eDNA, a semi-quantitative measurement of the agent concentration is possible. This can give a hint on the possible impact. In addition, eDNA cannot measure infection status of individual animals, meaning that no prevalence studies are possible using eDNA. This can only be overcome by combining different investigation strategies.
