Background and Objective

Since 1990 alders have been infected by the fungal-like pathogen Phytophthora alni. That resulted in severe damages on alder stands in forests and along river banks with fatal effects on flood defence. Outside nurseries, a control of the pathogen with chemicals is not allowed. Therefore, the aim of the project is to establish a first production of resistant alder clones of Alnus glutinosa:

- Selection and propagation of field resistant clones (TI)

- Development of a standardized inoculation method to test for resistance towards P. alni (JKI)

- Studies on the resistance mechanisms and on the tissue colonization with histological techniques (JKI)

Development on a method for molecular identification of alder clones (TI)

Approach

1. Selection of resistant individuals and populations from sites were P. alni is evident (approved selected seed stands, seed orchards, natural forests, bank protection stands). Growing of seedlings of selected progenies and clones (TI, JKI)
2. Development of a standard method to test the resistance of alders towards P. alni (JKI)
3. Testing of seed lots of approves seed orchards, approved stands and crossing families (JKI)
4. In vitro propagation of clones from individual trees showing resistance towards P. ×alni in first tests (TI)
5. Propagation of clonal material for a clone test for subsequent investigation after the project term (TI)
6. Adoption and standardization of histological techniques to study resistance mechanisms and tissue colonization in alder roots

Results

Project part A started with the allocation of alder seeds from different origins enabling access to alder genotypes during the project. Seeds from single trees in natural habitats with high infection pressure were acquired and offspring grown in a greenhouse. In addition, seeds from 21 licensed black alder seeding plantations were obtained in collaboration. Material was used for seedling production in a greenhouse. Beside black alder, grey alder seeds from a Bavarian region were acquired assumed to be resistant against P. ×alni. Alder clones (N25) were established and propagated in tissue culture from the offspring of characterized trees by the end of 2016. Genetic differentiation was achieved by a multiplex PCR with published microsatellite markers. Characteristic profiles of the clones showed that they are genetically unique. By the end of 2017, 60 further clones were established in tissue culture by TI-FG. In addition, the Belgian clone Liria W252 was established in tissue culture. Clones were propagated and rooted in vitro, transferred and assimilated ex vitro for infection experiments.

Project part B comprised the infection experiments to evaluate resistance of alder genotypes in short time. They comprised the in vitro incubation of rooted shoots, germinated seeds, truncated shoots, truncated leaves but also seedlings and potted alders cultivated under greenhouse conditions. In vitro infection
experiments with rooted shoots were superior to assess the gradual sensitivity of alder clones compared to infection experiments with potted specimen as their execution took place under constant and highly reproducible conditions with a large sample size. Infections obtained by application of defined zoospore suspensions of highly virulent P. ×alni isolates at the wounded stem base of potted alders or the incubation of truncated shoots lead to typical infection symptoms, but these experiments did not achieve a similar graduation of the symptom formation. Infection of leaf material was also less suitable due to the occurrence of partial leaf necrosis in negative controls.
Infections experiments identified clones 12/66, 12/67, 12/21 and Liria W252 as minor susceptible to P. ×alni. Clones are available for follow-up studies. The Bavarian grey alder accessions were unexpectedly highly susceptible to P. ×alni.