scott gifford, oscar sosa, dan repeta, ed delong methylotrophs dominate dilution-to-extinction...
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Scott Gifford, Oscar Sosa, Dan Repeta, Ed DeLong
Methylotrophs Dominate Dilution-to-Extinction Cultures Enriched with Marine Dissolved Organic Matter
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Particulate carbon
Dissolved Organic Carbon (DOC)
750 GT C
10-20 GT C
660 GT C
CO2
CO2
Release Mechanisms
LysisGrazingLeakage
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Dissolved Organic Carbon (DOC)
660 GT C
labile semi-labile refractory
?
lability
hr -days 1,000 – 2000 years
HMW
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Dissolved Organic Carbon (DOC)
DOC
Chemical Characterization
Dan Repeta
Rene Boiteau
Biology
Oscar Sosa
Ed DeLong
What is the chemical composition of DOC?
Which microorganisms consume DOC?
What are the genes involved in DOC degradation?
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www.nelha.org
Karl et al 1998
HMW DOM collection
Tangential flow filtration with 1kDa filter
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Ultrafiltration to obtain DOM
Courtesy Dan Repeta
Chemical Characterization
Isolates
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transfer transfer106 cells ml-1
5-10 cells ml-1
incubate
[DOM]FALC
ON
1
B
C
D
E
F
2 3 4 5 6 7 8A
inoculated well
control well
Dilution to extinction culturing
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Dil-to-Ext with a DOM twist
DOM
North Pacific : NEHLA
InoculantNorth Atlantic: Nahant Bay
BostonNahant
Bay
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noDOM 4X DOM 10X DOM total
wells inoculated 840 840 840 2520 controls 54 54 54 162
growth >105 inoculated 1 40 52 93 controls 0 0 0 0
50 to 100 100 to 500 500 to 1000 >10000
5
10
15
20
25
30
35
40
no DOM
4X DOM
10X DOM
cells per ml
wells
<105 105 5 X 105 106 or >
Screen wells for growth with flowcytometry
side scatter
log green fluorescence
Flow cytometry
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100x DOM
20x DOM
5x DOM
no DOM
days
cells per ml
Isolates have a distinct growth response to DOM
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Isolate Phylogeny (small subunit rDNA)
93 Isolates
88 obtain 16S rDNA
10 Gammaproteobacteria
5 Alteromonads3 SAR922 Vibrios
78 Betaproteobacteria
78 unclassified Methylophilales_ otu_2801_
88% of isolates are methylotrophs
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Isolate growth on methanol….
cells per ml
days
No MeOH 150 µM methanol
3
2
1
cells per ml X 106
No MeOH 150 µM methanol
days
Isolates can degrade C1 compounds
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McCarren et al 2010: Gamma: Alteromonads Gamma: Methylophaga
Current Study: Gamma: Alteromonads Beta: Methylophilales
DOM
transcripts
Same TaxaSame Functional Group
microcosm
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Alteromonads
Methylotrophs
CH3 CH3
HMW DOC
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TranscriptomicsGenomics
Draft genomes for 73 isolates
Substrate experiments
Purified DOMC1 compoundsComplex Carbs. porphyrin, xylan pectin, etc.,
Current work…..
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Isolates have a distinct growth response to DOM
Conclusions
Dilution cultures enriched with DOM produced more isolates
Hypothesize methylotrophs may have ability to use C1 compounds from DOM polymer
Isolate collection is dominated by methylotrophs
Acknowledgements
Jamie BeckerRene BoiteauChon Martinez
Tsultrim PaldenJohn EppleySara Lincoln
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