Algal Toxins in Arizona, Common Misconceptions and Future Research

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Algal Toxins in Arizona, Common Misconceptions
and Future Research
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• Analysis of algal toxins in watersheds
  surrounding the Phoenix Metro Area since 2000
  (comprehensive since 2002).
• Analyses include quarterly sampling of
  microcystin, anatoxin-a, cylindrospermopsin, and
  saxitoxin (analyses performed by either Greg
  Boyer at SUNY-CESF or Paul Zimba at USDA)

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• Algal counts and IDs in addition to suites of
  physico-chemical and chemical variables
  (partnered with ADEQ and AzGF).
• Development of a large, comprehensive database.

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Rodeo-Chedeski Fire

• The largest wildfire in Arizona recorded history
  began on June 18th, 2002.
• This was a re-setting event in the Salt River
  and downstream reservoirs.

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Primary Production in Roosevelt
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Primary Production in Apache, Canyon, and Saguaro
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Potentially Toxic Cyanobacteria/Algae Found in
Salt River Reservoirs

• Aphanizomenon flos-aquae
• Anabaenopsis circularis
• Anabaena laxa
• Anabaena schremetievi
• Anabaena torulosa
• Anabaena variabilis
• Cylindrospermopsis raciborskii
• Merismopedia elegans
• Microcystis sp.
• Pseudanabaena sp.
• Oscillatoria aghardii
• Oscillatoria limnetica
• Prymnesium parvum
• Gymnodinoids
• and several more

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Research Highlights

• During 08/2000, over 140 µg/L of anatoxin-a found
  in Saguaro.
• Both anatoxin-a (33 of all fish) and microcystin
  (67 of all fish) found at toxic levels in
  bluegill and threadfin shad stomachs taken from
  Apache Reservoir during 2004.
• No anatoxin-a found in aqueous samples.

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• Numbers of potentially toxic species were
  relatively low during the 08/2000 event.
• All cylindrospermopsin results (with the
  exception of a concentrated plankton tow) were
  non-detects.
• Numbers of potentially toxic species increase
  after toxic events.

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• Anatoxin-a is readily degraded by sunlight and
  alkalinity conditions not lacking in surface
  waters in Arizona during the summer.
• Half-life may only be a few hours under
  environmental conditions in these reservoirs
  during spring and summer.

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Prymnesium parvum

• Found in Salt River Reservoirs by AzGF, Spring
  2005.
• Working with AzGF on counts and ID, sampling
  methodology, etc.

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• Caused massive fish kills in Texas since the
  1980s.
• Highly heterotrophic and toxin production may be
  used to slow prey (bacteria, other algae) prior
  to ingestion.
• Causes massive hemorrhaging of exposed gill
  tissue in fish, bivalves, and zooplankton
  eventually spreading to internal organs.

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• Initially believed to be an estuarine species
  requiring high salinities and relatively low
  water temperatures.
• Seems to be adapting to a wide variety of
  habitats.
• Environmental conditions for toxin production are
  not well known but nitrogen limitation may be a
  trigger.
• Has a dormant, encysted stage in sediments.

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Image From Texas Parks and Wildlife Dept.
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Image From Texas Parks and Wildlife Department
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P. Parvum Background in Salt River Reservoirs

• Large fish kills occurred in early June of 2004
  and 2005 and the riverine zone of Saguaro most
  affected.
• Small kills noticed beginning in March of both
  2004 and 2005.
• Riverine zone of Saguaro is, essentially,
  hypolimnetic water from Canyon.
• Canyon has pump-back storage.

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• Riverine sections of reservoirs usually very low
  in nitrogenous compounds early in the year.
• Thermal stratification and subsequent
  hypolimnetic accumulation of ammonia/ammonium
  released into downstream reservoirs may trigger a
  decrease in toxin production.
• Again, numbers alone do not directly correspond
  with toxicity.

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• Arizona Game and Fish and Applied Biosciences
  recently sponsored a meeting where several
  colleagues from Texas shared their expertise.
• We continue to collaborate with colleagues in
  Texas and elsewhere on P. parvum issues.

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Common Misconceptions re. HABs in Arizona and
Elsewhere

• Correlation between numbers of toxic species
  present and the amount of toxin being produced.
• The only way to determine amount of toxin in the
  water is through direct quantification of
  toxin(s).

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Misconceptions (cont.)

• Species produce constant or set amounts of
  toxins.
• Environmental conditions for toxin production are
  well-known.
• Algal toxins exert their influence in isolation.
• Genes coding for toxin production are known.
• No new or novel toxins exist.

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Misconceptions (cont.)

• Genomic fingerprinting, without first determining
  toxicity of all strains, will provide an
  early-warning system.
• Genomic fingerprinting provides information of
  environmental variables responsible for toxin
  production.
• Toxicity is consistent with genetic analysis.

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Misconceptions (cont.)

• All genomic analysis is created equal.
• Much exists downstream of 16S rRNA
• No correlation between 16S rRNA genetic analysis
  and toxicity of Microcystis strains (Neilan et al
  1997a)

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New Research

• In collaboration with ADEQ and AzGF.
• Co-PIs Paul Zimba (USDA) and JoAnn Burkholder
  (NCSU).

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A De-Constructionist Approach

• Simplicity and sound research design prioritized.
  
• Changes in sampling methodology to reflect
  current knowledge.

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• Composite samples taken from all reservoirs.
• Split samples between UA and Dr. Burkholder.
• Identification to species.
• Isolation of all suspect species into axenic
  cultures.
• Bioassays to screen toxin producers.
• Expose organisms to
• Individual algae
• Algal supernatant
• Mixed population

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• Quantification of toxins by Dr. Zimba
• Anatoxin-a, microcystin, cylindrospermopsin,
  saxitoxin, and prymnesin.
• Examine the role of bioactive peptides
  (chemotype differentiation of species
  producing, aeruginosins, cyanopeptolins,
  microginins, microviridins etc.).
• Examination of novel toxins
• Euglenoids
• Gymnodinoids

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Genetic Analysis of Known Toxin Producers

• Only after toxin-producing species have been
  isolated will genetic analysis occur.
• Who cares what strains are present if only a
  given few produce toxin?

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Dendrogram of cyanobacterial 16 S genes
(generated by RFLP). Scale bar similarity
coefficients. Numbers 14 clades. Symbols ?
neurotoxic ? non-toxic ?hepatotoxic ? toxicity
not determined ? type of toxicity unknown.
Lyra et al. (2001)
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Dendrogram of cyanobacterial genomic fingerprints
(generated by REP- and ERIC-PCR). Scale
similarity coefficients. Numbers 14 groups.
Symbols ? neurotoxic ? non-toxic ?
hepatotoxic ? toxicity not determined ?type of
toxicity unknown. Lyra et al., 2001
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Avoiding Hysteria

• Tempe Town Lake and Killer Microcystis
• Fish kills in Saguaro likely not due to
  cyan-toxins, or any toxin that can harm humans.
• This does not mean there are no human-health
  implications from other toxins.
• No matter how carefully-worded, several put all
  their research eggs in the C. raciborskii basket

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• Responses given to the media or public following
  an event should be based on sound research, not
  speculation.
• Its OK to say we dont know but were working
  on it
• Research not driven by alarmism.

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Summary

• Successful management of HABs can only occur
  through the enhanced understanding that research
  provides.
• Which strains/toxins
• Environmental factors (eutrophication,
  allelopathy, grazing pressure, water movement,
  etc.)

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Questions?