Symbioses between eukaryotes and nitrogen-fixing cyanobacteria off Brazil

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Catherine Ribeiro, Adriana Lopes dos Santos, Dominique, Marie, Frederico Brandini, Daniel Vaulot Station Biologique de Roscoff, UPMC et CNRS, France Universidade de São Paulo, Brasil Symbiosis between nitrogen-fixing bacteria and eukaryotes on the continental shelf of Brazil

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N2 fixation in the ocean Trichodesmium

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Nitrogen fixing cyanabacteria nifH gene

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Identifying UCYN-A

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UCYN-A lacks critical genes

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UCYN-A occurs in Symbiosis with Eukaryote UCYN-A Eukaryote NanoSIMS N15 enrichment

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The plot thickens 16S rRNA

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Genome of UCYN-A1 vs A2 86% similarity at AA level 135 genes

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The plot thickens again... Three types of UCYN-A Different hosts A1 A2 A3 ?

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Always more clades... A1 -> A6

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UCYN-A is major contributor to N2 fixation Trichodesmium UCYN-A

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Distribution of hosts A1 Host A2 Host CARBOM

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CARBOM cruise - Continental shelf Brazil SACW BC

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TR1 TR2 Pico and Nanoeukaryotes Picoeukaryotes Nanoeukaryotes

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16S rRNA from sorted cells Chrysophyc Clade B2 Isochrysidales Braarudosphaeraceae EF051761 - Chrysochromulina campanulifera - strain UIO J10 HM133350 - Biosope T60 16S OXY107.017 HM133259 - Biosope T41 16S PLA491.033 LK391826 - Ochromonas sp. - strain F933 HM133175 - Biosope T35 16S PLA491.033 St.98 surf PEUK clone F4 St.TRICHO surf PEUK clone B1 St.98 surf NEUK clone C2 HM133499 - Biosope T84 16S PLA491.026 HM133324 - Biosope T58 16S PLA491.010 LN735407 - Emiliania huxleyi - strain RCC 2941 KP282836 - Gephyrocapsa muellerae - strain RCC3862 LK391825 - Chrysochromulina apheles - strain F929 St.98 surf PEUK clone E5 HM133343 - Biosope T60 16S OXY107.004 HM133380 - Biosope T60 16S PLA491.021 HM133420 - Biosope T65 16S.38a AB847984 - Braarudosphaera bigelowii - strain TMRscBb1 2 2 7 15 picoplankton sequences nanoplankton sequences 69 100 99 57 88 92 99 90 100

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Strategy • Samples preserved with DMSO 10% • Two fractions sorted by flow cytometry : Pico and Nano (56 samples) • Two genes – nuclear 18S rRNA (V4) – nifH • 1 run Ilumina MiSeq 2*250 bp • Analysis with Mothur • Assignement : PR2, BLAST

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18S rRNA classes

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18S rRNA OTUs

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18S rRNA classes

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Bray distance clustering based on major 18S OTUs Dinos Mam Prym Diatoms Chrys UCYN-A1 host Ostreo + Bathy Prymnesiophyceae Prorocentrum

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Picoeukaryote communities are structured by the environment

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nifH diversity A1 A3 Tricho A2 A4

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A1 A3 Tricho A2 Bray distance clustering based on major nifH OTUs A4

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18S vs nifH

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Network analysis using 18S rRNA and nifH "Pico" network SparCC

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Conclusion On the shelf off Brazil • Two major types photosynthetic eukaryotic communities : – Braarudosphaeraceae – Ostreococcus + Bathycoccus • Nitrogen fixers – UCYN-A is dominant nitrogen fixer : A1, A2, A3, A4 • Network analysis – Recover A1 and A2 associations – A4 could have same host than A2 (Braarudosphaera) – A3 ?