Freshwater crayfish are ecosystem engineers with a big influence on biological diversity and functioning ecosystems. Unfortunately, populations of native crayfish are declining in all of Europe. The biggest threat to european crayfish are the rising populations of invasive north-american crayfish that are transmitters for the crayfish plague Aphanomyces astaci. That Oomycet in ranked among the 100 worst invasive species world-wide and caused the extinction of whole populations of freshwater crayfish in Europe with massive effects on ecosystems. Therefore it is of great ecological significance to research how native crayfish species can resist the crayfish plague.
Project aims:
For a long time it was assumed that invertebrates lack an immune memory. For crustaceans though, first proof for an immune memory was provided through immune priming experiments. The underlaying mechanism behind that phenomena is most likely connected to epigenetic processes, which are changes on the level of gene expression. Maternal transgenerational immune priming (mTGIP) is such an epigenetic mechanism. The females brace there offspring against germs that they themselves had come in contact with. This mTGIP triggers the expression of certain immune respones gene which leads to a similar immune defense then the one of the mother.
Our aim is therefore to develop and test a vaccination for female novel crayfish(Astacus astacus), that will protect there offspring from the crayfish plague through mTGIP.
For centuries, the crayfish plague has caused great ecological damage in Germany and Europe. Regardless, a treatment to produce native crayfish with stronger immune systems was not tested so far. The availability of plague resistant stocking crayfish would significantly improve the long-term success of management strategies. Therefore the proposed vaccination method can be regarded as highly innovative and ecologically relevant. A positive result would be a milestone for the management of freshwater crayfish, that would greatly improve the success of restocking measures.
To reach our goals we plan to do our experiment in three consecutive work packages:
- Vaccination of the females before mating:
The female crayfish are injected subcutaneously of raw hyphae extract from A. astaci to intensify their immune answer without actually infecting them (the hyphae are the non-infectious stage of A. astaci). To identify the best time and dose the crayfish will be grouped in three vaccination groups: They will receive a vaccination either four weeks (Group 1) or eight weeks (Group 2) before mating. Some females will get vaccinated on both times (Group3), receiving twice the amount of vaccine. A fourth group of female breeding animals remains unvaccinated as a control. To assess the immune status of the female crayfish we take sublethal hemolymph samples for qPCR of immune genes at different time points before and after vaccination as well as before mating. - Mating under mTGIP
We use the vaccinated females for controlled mating in indoor ponds containing multiple males. After mating, the eggs of each female will be incubated in separate incubation baskets where the larvae can hatch and develop into juveniles. The immune status will be assessed using qPCR of immune genes during the development of the eggs and the offspring. The F1 generation will be raised until their first summer, separated by vaccination groups. At this point the juveniles should have reached an average size of 4 cm. This is adequate for the following controlled infection experiments to test if TGIP is sufficient. - Infection experiment to proof mTGIP
We use the F1 generation of noble crayfish for controlled infection experiments to verify the efficiency of the mTGIP approach. Therefore we conduct a controlled infection experiment on each vaccination group with 10 treated and 10 control animal from each group. We monitor the treated individuals for disease symptoms. After the infection experiment, the immune status of all crayfish will be examined with qPCR. The surviving F1-groups can be used for restocking measures.
