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Genetic and environmental factors influencing Microcystis bloom toxicity

Genetic and environmental factors influencing Microcystis bloom toxicity. Juli Dyble NOAA Great Lakes Environmental Research Lab Ann Arbor, MI. Great Lakes . Saginaw Bay. western Lake Erie. Great Lakes as an aquatic resource. Largest supply of freshwater in the world

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Genetic and environmental factors influencing Microcystis bloom toxicity

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  1. Genetic and environmental factors influencing Microcystis bloom toxicity Juli Dyble NOAA Great Lakes Environmental Research Lab Ann Arbor, MI

  2. Great Lakes Saginaw Bay western Lake Erie

  3. Great Lakes as an aquatic resource • Largest supply of freshwater in the world • 80% of US freshwater supply • Drinking water supply for 40 million US and Canadian citizens • Over 500 beaches for swimming and recreation

  4. Aphanizomenon Oscillatoria Microcystis Anabaena Cylindrospermopsis Common cyanobacterial HAB genera in the Great Lakes

  5. Microcystis in the Great Lakes Lake Erie, Put-In-Bay, Sept 2006 Lake Erie, South Bass Island, Sept 2006

  6. Microcystis in the Great Lakes P abatement programs (Great Lakes Water Quality Agreement) Dreissenid mussel introduction Dominant member of phytoplankton community Blooms frequent and abundant High P input to system (detergents, fertilizers, septic) 1970 1980 1990 2000 Decrease in chlorophyll, increased water clarity Blooms rare Return of Microcystis blooms up to 90% phytoplankton community Present Abundant Microcystis blooms, July - Sept

  7. Zebra mussel (Dreissena polymorpha) • Quagga mussel (Dreissena bugensis) • Introduced by ballast water from eastern Europe

  8. Impacts of zebra mussels on Microcystis • promote growth of toxic Microcystis strains • Nutrient excretion • provide sufficient energy • for growth of toxic strains • change N:P ratios • Grazing pressure • selective rejection of • toxic Microcystis strains • consume more palatable • species Zebra mussels

  9. Current projects Microcystis sp. • Map microcystin concentrations and Microcystis cell numbers in Saginaw Bay and western Lake Erie • Identify environmental factors promoting microcystin production • Develop rapid methods for detection of toxic Microcystis • Accumulation in fish

  10. What makes a cyanobacterial bloom toxic? Mostly toxic strains Mostly non-toxic Light Nutrients Temperature Trace metals Not producing toxin Producing toxin • Shift in community composition • Stimulation of toxin production by environmental factors

  11. What makes a cyanobacterial bloom toxic? 10-1000 fold change in toxicity Mostly toxic strains Mostly non-toxic • Shift in community composition • Stimulation of toxin production by environmental factors 2-10 fold change in toxicity (Zurawell et al 2005) Not producing toxin Producing toxin

  12. Intracellular total microcystin by HPLC August 2004 Saginaw Bay Put-In-Bay 58 µg L-1 western Lake Erie

  13. Distribution of microcystin congeners Relative toxicity: - LR and –LA = 4x more toxic than –YR 10x more toxic than -RR

  14. Advantages of molecular techniques • Differentiate morphologically identical strains • toxic and non-toxic strains • Identify geographic origin of strains and genetic diversity of populations • Rapid detection

  15. Identifying toxic strains of Microcystis • All toxin-producing strains of Microcystis contain genes for microcystin production: mcyA-J • Presence of mcyB = strain able to produce toxin Absence of mcyB = non-toxic Pearson et al 2004

  16. Multiplex PCR for toxic Microcystis Saginaw Bay western Lake Erie M mcyB ITS M = molecular weight marker Number of colonies Basin # mcyB total (ITS) Saginaw 36 40 Erie 4 16 % microcystin producers 90% 25%

  17. Distribution of toxic Microcystis Stations with mcyB km 0 10 20

  18. Distribution of C. raciborskii in the US L. Yale L. Eustis Silver L. L. Griffin L. Harris Little L. Harris Trout L. L. Dora L. Beauclair L. Ola L. Carlton L. Monroe L. Jesup + + + + + + + + + + Cylindrospermopsis specific nifH primers

  19. western Lake Erie Maumee Bay Quantitative PCR for enumerating toxic Microcystis colonies Applications • measure temporal variation in proportion of toxic strains • biweekly sampling at 3 locations in western Lake Erie • identifying conditions under which cells are actively producing toxin (expressing mcyB) • zebra mussel grazing • changes in nutrients and light • Tie into circulation models to predict distribution of toxic Microcystis strains and forecast water quality

  20. Goal • Develop predictive capabilities for presence of toxic cyanobacterial blooms in Great Lakes recreational and drinking water supplies

  21. Thanks ……. • Center of Excellence for Great Lakes and Human Health (Oceans and Human Health Initiative) • Gary Fahnenstiel (NOAA-GLERL) • Hank Vanderploeg (NOAA-GLERL) • Pat Tester, Wayne Litaker (NOAA-Beaufort) • Dave Millie (Florida Institute of Oceanography) • Crew of the R/V Laurentian

  22. Monthly averages Monthly averages 1000 2 800 600 Toxins (ng g-1) Toxins (ng g-1) 1 400 200 0 0 Muscle Liver 22 22 22 24 24 24 22 22 22 9 9 9 Jun Jun Jul Jul Aug Aug Oct Oct Microcystin concentrations in PerchLake Erie, summer 2006 ng toxin (g dry mass)-1 Microcystin concentration of concern for routine fish consumption = 7.7 ng g-1

  23. Phylogenetic tree based on mcyB Erie D7 Erie F2 99 Erie F2B 56 Erie D2 Microcystis botrys 93 Microcystis aeruginosa) 69 Saginaw C3E Saginaw C3E 91 79 Saginaw A4W 100 Erie D2 Microcystis aeruginosa Microcystis viridis Erie F2B 54 Microcystis aeruginosa 87 Saginaw A4W Erie F4 98 Erie E2 Microcystis aeruginosa Microcystis botrys 100 64 Erie D2 100 Microcystis aeruginosa Microcystis sp. 70 Saginaw C5 Saginaw C3E Saginaw B1 Saginaw B1 Saginaw A4W Saginaw B1 Saginaw C5 Erie D7 100 Erie F2B 0.1 substitutions per site mcyB1(C) cluster  MC-RR mcyB1(B) cluster  MC-LR Designations according to Mikalsen et al 2003

  24. Purpose of cyanotoxins? • Secondary metabolites • no role in primary metabolism, growth or reproduction, but have somehow evolved to benefit the organism • includes alkaloids, polyketides and nonribosomal peptides • Anti-grazer, antibacterial, antifungal • Chemoattractant for microbial community • Unlikely that production of such a major cellular constituent would be retained through evolution if not of biological significance

  25. MICROCYSTIN PRODUCTION Environmental factors influencing growth and toxin production in Microcystis NUTRIENTS LIGHT HAB GROWTH WATER TEMPERATURE WIND SPEED GRAZING

  26. Summary • Microcystin concentrations above the WHO drinking water standards are common in Saginaw Bay and western Lake Erie • Highest close to lake edges where increased human exposure • Multiple strains of Microcystis are present and toxicity may be related to genetic composition of community • Designed assays for detecting toxic strains • Currently working to identify environmental conditions that regulate microcystin production

  27. Measuring microcystins • HPLC(high performance liquid chromatography) • Distinguishes between variants • Based on retention times • Dependent on availability of standards • Lab-based • Detection limit: ~0.1 µg/L Peak # 1 - RR 2 - LA 3 - YR 6 - LR Harada et al 1999

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