1. Mycorrhiza activity in Cypripedium calceolus roots Ieva Druva-Lūsīte, Dace Kļaviņa National Botanic Garden, Latvia 17-19 May 2013, Rende, Italy

2. Cypripedium calceolus is included in Red Data Book of Latvia as vulnerable species and listed on Annexes II and IV of the Habitats Directive and Appendix II of the Berne Convention.

3. Background of study Imature seeds Seedling Seedling after cold storage Mycorrhization Seedlings with first leaves Second season after ex vitro

4. The Objective To evaluate the seasonal dynamic of mycorrhiza activity in C.calceolus roots in natural habitats and in National Botanic Garden area.

5. Spread of C.calceolus and sampling places 1. Sloka 2. Ķemeri 3. NBG 4. Silzemnieki

6. Population of C. calceolus in all territory of Latvia Numbers

7. Large clamps and juveniles in samplig places %

8. Sloka pH HCl 5.1 Organic matter: 37% Vitality of population: poor Habitat: mixed broad- leaved - coniferous forest

9. Ķemeri pH HCl 7.1 Organic matter: 20% Vitality of population: good Habitat: broad-leaved deciduous forest

10. Silzemnieki pH HCl 6.4 Organic matter: 19% Vitality of population: excellent Habitat: broad-leaved deciduous forest

11. Arboretum of National Botanic Garden pH HCl 6.3 3-4 years old seedlings Habitat: arboretum with Corylus avelana

12. Method of mycorrhiza estimation Mycorrhiza Manual 2001, www2.dijon.inra.fr/mychintec/Protocole/protoframe.html Values put into the computer program “Mycocalc” to calculate M% - intensity or activity of the mycorrhizal colonisation in the root system. N=50 eyeshots in one repeat, minimum – three repeats. Roots cleared in 10% KOH for 1h; roots rinsed with water; stained in 0.05% trypan blue for 5 min; stored in lactoglicerol.

13. Mycorrhiza activity in Sloka M%

14. Mycorrhiza activity in Ķemeri M%

15. Mycorrhiza activity in Silzemnieki M%

16. Mycorrhiza activity in arboretum of NBG M%

17. Mycorrhiza activity in dormant Cypripedium calceolus in Ķemeri M%

18. Conclusions Mycorrhiza activity was higher in the begining of vegetation season in good and excellent orchid populations. Then it slowly decreased till August. Mycorrhiza activity showed stability of habitat and orchid population in Silzemnieki. In average mycorrhiza activity in Sloka was a comparatively higher than in arboretum of National Botanic Garden. Next step – to investigate mycorrhiza species in C.calceolus roots.

19. Cypripedium calceolus200x 400x 200x

20. Acknowledgements This study was funded by Latvian Environmental Protection Fund. Soil chemical analyses made at Laboratory of Plant Mineral Nutrition, Institute of Biology, University of Latvia. Thanks for permission to use microscope to Laboratory of Mycology and Plant Pathology, Latvian State Forest Research Institute "Silava“.