1. The Bracken Fern (Pteridium aquilinum) Toxic effects on surrounding organisms and environment By: Peter Andriakos

2. General Information Wide distribution  Globally extensive  Widest distribution of any fern genus  Among most common plants on the planet Locally intensive  Rapid invasion of de-forested areas  Cover increasing at a global level  Major problems in the UK, Scotland, Wales, South America

3. General Info. (cont.) Persistence  spreads via rhizome  widespread underground rootstock  forms expansive stands, dense thickets Resilience  limited only by extreme cold, altitude  observed growing in wide range of soil pH  highly successful dispersal abilities

4. Human Bracken Fern Consumption  Bracken fiddleheads harvested  Many cultures throughout history Maori (NZ) herbal remedy, food  Eastern Asia (Japan, China, Korea) staple vegetable  Still utilized today as a foodstuff  Toxic effects are now known continues to be utilized

5. Non- Human Consumption Animal consumption  Domestic Herbivores Restricted feed availability Will consume readily  Major problem Toxic effects on animals Indirect effects on humans Toxic effects observed in all animal species known to consume Pteridium aquilinum Cows consuming Pteridium aquilinum while grazing

6. Toxic and Carcinogenic Effects of Bracken Fern  Wide variety of toxic effects observed vary by species among other factors  Several known toxins isolated from Bracken several carcinogenic others mutagenic  Experimental determination of toxicity studies conducted with laboratory animals myriad of syndromes observed again, vary by species

7. Bracken carcinogens in the human diet (Mahmood Shahin, Barry L. Smith, Arungundrum S. Prakash) An all encompassing article… Bracken Fern issues seen in animals Human health risks Primary carcinogenic principal Mode of carcinogenic action Cancer model

8. Toxic syndromes in animals  Numerous acute, toxic syndromes observed induced thiamine deficiency acute hemorrhagic syndrome  Severity dependant on… species and age of animal quantity/quality of plant consumed consumption rate

9. Acute hemorrhagic syndrome  seen in ruminants  degenerative change in more rapidly dividing cells  epithelial necrosis - larnyx, pharynx, small intestine  bone marrow aplasia -Platelet production ceases -“Hemorrhagic crisis” occurs -Leucopenia, thrombocytopenia, granulocytopenia Death occurs in a matter of weeks

10. Chronic Toxic Syndromes Chronic toxicity in animals also observed…  Bright blindness seen in sheep (ruminant) (Watson et al., 1965) retinal stenosis, atrophy (Watson et al.,1972)  Enzootic hematuria Tumors in the bladder mucosa hemorrhaging in bladder wall Pamukcu et al., 1967  Carcinomas upper digestive tract

11. Laboratory Animal Experimentation  Rats first report of carcinogenic potential (Evans, Mason. 1965) Diets containing Bracken powder, fronds, rhizomes Higher incidence of tumor formation vs. control Fronds vs. rhizomes (Hirono et al.,1973) Duration of exposure  critical factor Subjects fed 33% dried bracken (Hirono et al., 1970) 4 months vs. 8 months

12.  Mice feeding trials, dried bracken (Yasuda et al.,1974) rib anomalies, sternebrae fusion Tumor formation Carcinogenic effects of cow milk (Pamukcu et al., 1978)  Other experimental animals… Guinea Pigs Japanese Quail Egyptian Toads

13. Human Health Risks  Indirect effects of animal consumption milk obtained from bracken fed cattle leaching in to water supply aerial dispersion of spores  Esophageal carcinomas observed Japan (Kamon et al., 1975)  Gastric cancer frequency Wales (Galpin et al., 1990) Costa Rica (Villalobos-Salazar et al., 1989) Brazil (Marliere et al., 1995)

14. Toxic Compounds  Numerous molecules isolated  Carcinogenic, mutagenic  Quercetin  mutagen  Ptaquiloside (PT)  1 0 carcinogenic principle Ptaquiloside molecule Quercetin molecule

15. Ptaquiloside (PT) Principal carcinogen in Bracken Norsesquiterpene glucoside Difficult to isolate Carcinogenicity confirmed by Hirono et al. in 1984 Various other experimental confirmations

16. PT Action Mechanism Proposed scheme of PT reaction pathway

17. Carcinogenic basis of PT  Carcinogenesis  initial DNA damage DNA alkylation (adenine, guanine) Adenine Guanine DNA Structure

18. PT Cancer Model Multistage model for bracken-induced carcinogenesis

19. Occurrence of the carcinogenic Bracken constituent ptaquiloside in fronds, topsoils, and organic soil layers in Denmark (Rasmussen, Kroghsbo, Frisvad, Hansen)  relevance  human uptake via watersheds  Investigate occurrence of PT in fronds, topsoil materials  Multivariate data analysis

20. Materials/Methods  20 populations chosen in Denmark 3 sub-sites at each location  Sample at end of growing season Soil + plant material Dried  milled  stored @ 4 0 C  Frond height and density measured Map of Denmark, study sites indicated

21. Soil Horizons http://www.dpi.vic.gov.au/CA25677D007DC87D/LUbyDesc/AG1060a/$File/AG1060a.gif Soil Horizon Diagram  Focus was on topsoil layers  Horizons O and A 1

22.  Other measurements taken Soil pH Organic Carbon Content Bracken Biomass Precipitation level Light exposure Turnover rate  Partial Least Square Regression Analysis (PLSR) Performed on all variables less PT content Correlate parameters with PT content in fronds, horizons  PT analysis Fronds, litter, O/A horizons Extraction using de-ionized H 2 0 “cleaning" of sample with a resin Conversion to pterosin B Liquid chromatograph utilized

23. Results Ptaquiloside content  PT content in… Fronds  110 - 3800 [μg g -1 ], mean = 550 [μg g -1 ] O horizons  0.09 - 6.43 [μg g -1 ], mean = 0.39 [μg g -1 ] A horizons  0.011 – 0.713 [μg g -1 ], mean = 0.031 [μg g -1 ]

24. Results PLSR findings (+)(-) Light exposureFrond height Turnover Rate Carbon Content Fronds (+)(-) PrecipitationAmt. of Litter Turnover Rate Stand Size O horizons (+)(-) Soil pHPrecipitation Stand Size Easting A horizons

25. Conclusions  Definitive evidence that PT is found in topsoils beneath Bracken stands  Possibility that leaching does occur  High precipitation areas most susceptible to watershed contamination

26. Questions Raised  How concerned should a local human population be?  Should Bracken management be implemented? – Has been in some areas…  Do these strategies need to be re-evaluated for their efficacy? – Bracken cover is increasing rapidly…  Need to think about Bracken management in agriculture from an environmental point of view…