Nutrient effects on virus and bacterial communities in the North Pacific Ocean

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Nutrient effects on virus and bacterial
communities in the North Pacific Ocean

• Ian Hewson Josh Steele
• Douglas Capone
• Jed A. Fuhrman
• University of Southern California

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Presentation Outline

• Introduction to Marine Bacteria and Viruses
• Experimental Design / Measurements
• Bacterial Production
• Bacterial Abundance
• ARISA (Community Fingerprinting)
• Virus Abundance
• Evidence for Mesocosm Death
• Conclusions
• Future Directions

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Oceanic Bacterioplankton

• Most abundant physiologically - active marine
  organisms - 106 - 107 ml-1 in surface water
• Bacteria critical to biogeochemical cycling in
  oligotrophic ocean
• nifH sequences from heterotrophic diazotrophs
  detected at HOTS site (Zehr et al. 2002)
• regeneration of DON
• Bacterial production potentially Fe limited
• Black box of bacterial community composition
  not well studied in oligotrophic ocean,
  particularly upon Fe amendment

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Marine Virioplankton

• Most abundant organism in ocean - 108- 109 ml-1
• Infect bacteria and protists
• Cause significant mortality of bacteria - 2 - 35
  d-1 (Proctor
  Fuhrman, 1991)
• Only studied in detail in last 12 years
• Previous study difficult due to size ( 20 nm )
• Virus hypothesized to influence bacteria
• Production - recycling of DOM from lysed cells
• Community diversity - killing winner of resource
  competition
• No previous studies of effects of Fe on
  virioplankton

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Microbial Iron Cycle
(Tortell, et al, 1999)
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Presentation Outline

• Introduction to Marine Bacteria and Viruses
• Experimental Design / Measurements
• Bacterial Production
• Bacterial Abundance
• ARISA (Community Fingerprinting)
• Virus Abundance
• Evidence for Mesocosm Death
• Conclusions
• Future Directions

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Mesocosm Experimental Design

• Part of Effects of Fe / P and Aeolian dust
  impacts on picoplankton Mesocosm Experiments
• MP05 - R/V KaImikai o Kanaloa - July 2002
• MP06 Biocomplexity Cruise - R/V Kilo Moana -
  September-October 2002

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Bacterial Analysis
Biomass
Activity
Diversity
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Virus and Bacterial Analyses

• Bacteria and Virus Abundance -
• SYBR Green I staining (DNA fluorochrome) and
  epifluorescence microscopy

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Bacterioplankton Production - TdR and Leu
Incorporation
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Possible Consequences of Enrichment
1. Same community becomes more productive 2.
Community replaced by more productive community -
weedy species 3. No Effect 4. Negative
Effect - community death
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Effects upon Net Production MP05
Via TdR
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Effects upon Net Production MP06
Via leu
Meso 2
Meso 1
Bacterial Production (cells ml-1 d-1)
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Effects upon specific growth rate
Specific Growth Rate ( d-1)
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Effects upon Bacterial Abundance
Bacterial Abundance (cells ml-1)
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Virus and Bacteria Community Analyses

• Bacterial community diversity -
• Bacteria from 4 L of each mesocosm at each time
  point collected on 0.22 um Durapore filter
  (prefiltered through 0.8 um GF/F to remove
  protists -gt Transported to USC frozen
• DNA extracted from durapore
• automated rRNA intergenic spacer analysis (ARISA)
  conducted on mixed-community bacterioplankton DNA

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Automated rRNA Intergenic Spacer Analysis (ARISA)
(A way to separate PCR products analytically from
complex mixtures)

• DNA extracted from mixed community
• PCR performed with one tagged primer
• Forward universal, reverse eubacterial

Products run on automated sequencer.
Shows exact sizes Each peak represents an
operational taxonomic unit
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MESO 1
T 0
Control
Fe
P
Fe P
Dust Cont
Dust
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MESO 2
T 0
Control
Fe
P
Fe P
Dust Cont
Dust
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MESO 1
1.00
0.90
0.80
0.70
0.60
0.50
0.40
0.30
Similarity Jaccard Coefficient
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MESO 2
1.00
0.90
0.80
0.70
0.60
0.50
0.40
Similarity Jaccard Coefficient
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Talk Outline

• Introduction to Marine Bacteria and Viruses
• Experimental Design / Measurements
• Bacterial Production
• Bacterial Abundance
• ARISA (Community Fingerprinting)
• Virus Abundance
• Evidence for Mesocosm Death
• Conclusions
• Future Directions

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Mesocosm death
Abundance
Production
Bacterial Production (cells ml-1 d-1)
Bacterial Abundance (cells ml-1)
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Changes in Virus Abundance
Meso 1
Meso 2
Virus Abundance (VLP ml-1)
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Talk Outline

• Introduction to Marine Bacteria and Viruses
• Experimental Design / Measurements
• Bacterial Production
• Bacterial Abundance
• ARISA (Community Fingerprinting)
• Virus Abundance
• Evidence for Mesocosm Death
• Conclusions
• Future Directions

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Conclusions

• Bacterial production stimulated significantly by
  addition of P, Fe and Dust compared to respective
  controls
• Bacterial community composition significantly
  altered by containment
• Bacterial communities in Dust incubation
  resemble Fe addition
• Virus production elevated in some mesocosms which
  are not physiologically active gt Lysogen
  Induction?

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Talk Outline

• Introduction to Marine Bacteria and Viruses
• Experimental Design / Measurements
• Bacterial Production
• Bacterial Abundance
• ARISA (Community Fingerprinting)
• Virus Abundance
• Evidence for Mesocosm Death
• Conclusions
• Future Directions

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Future Directions- MP08 and beyond

• Induction of lysogens
• Who - PFGE Virus Diversity / Distinctive band
  formation
• How many - Mitomycin C addition before / after
  comparison
• Why - Elevated physiology, contaminant, UV
  exposure
• Bacterial community function
• Insufficient to consider only 16S - functional
  groups of bacteria
• Nitrogen fixers (nifH)
• Ammonium oxidizers (amoA)
• DOM release upon lysogen induction may be
  important quantify
• mRNA siderophore production gene heterogeneity?

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Acknowledgements
Michael Schwalbach Xiaolin Liang Ximena
Hernandez Mark Brown Jill Sohm Juliette
Finzi Troy Gunderson Luisa Falcon Rachael
Foster Toby Westberry
Stephanie Jaeger Paul Morris Reni
Schimmoeler Michael Neumann Jay Burns Lia
Protopapadakis Sarah Govil Anyone else we forgot
on MP05 and MP06 and anywhere else