Gut$community$analysis$$of$Atlan0c$Cod$(Gadus&morhua)$

Thomas'HA'Haverkamp1'(thhaverk@bio.uio.no),$B$Star1,S$Jento<1,$KS$Jakobsen1$

Affilia0ons$

Methods$Fishes were caught in the outer Oslo-Fjord, Norway. Guts were dissected from fishes and flushed with RNAlater. The flushed gut contents were used for DNA isolation. Amplicon sequencing was done with barcoded primers targeting the 16S rRNA V3 region. Amplicons were pooled and shot-gun sequenced using 454 technology giving 370299 reads[3]. Sequence data was cleaned from artifacts using Mothur[4]. Calculation of diversity was performed in Mothur. Taxonomic classification was done using BlastN against the SILVA SSUref NR V108 and analyzed in MEGAN[5].

Introduc0on$By far the majority of vertebrate diversity is represented by fish lineages. Nevertheless, gut communities of vertebrates have been mainly investigated in mammalian hosts despite the fact that mammals only represent a fraction of this diversity. Most fish gut communities have been studied with 16S rRNA clone libraries or culture based methods and only one pyrosequenced zebrafish gut community study – in which individual samples were pooled - is available[1,2]. Therefore, a detailed microbial community analysis of fish guts using the power of next generation sequencing on individual samples is lacking.

Here we analyze pyrosequenced amplicon libraries of the gut communities of 11 Atlantic Cod (Gadus morhua) specimens caught in the outer Oslo Fjord. We find that the most abundant taxa are shared among individuals, an observation supporting the idea of the presence of a core gut microbiome in fish species[1].

Conclusions$1.   The Cod gut community is dominated by species beloning to

the orders, Clostridiales, Bacteriodales and Vibrionales. 2.   The diversity of low abundant taxa explains most of the

diversity differences between the 11 gut communities

Results$

1$Center$for$Ecological$and$Evolu0onary$Synthesis,$Dept.$of$Biology,$University$of$Oslo,$Oslo,$Norway$

References$Acknowledgements$We$thank$$Jonathan$E.$Colman$(UMB,$Ås)$for$catching$the$fish$and$the$Norwegian$Sequencing$PlaTorm$at$UIO$for$$sequencing$of$our$samples$$(www.sequencing.uio.no).$

[1]$$Sullam$et$al.$2012,$Mol$Ecol$21:$3363\3378$[2]$Roeselers$et$al.,$2011,$ISME$J$5:1595\1608$[3]$Margulies$et$al.$2006,$Nature$437:$376\380$[4]$Schloss$et$al.$2009,$Appl$Environ$Micro$$$$$$$Biol$75:7537\7541$[5]$Huson$et$al.,$2009.$BMC$bioinforma0cs$10,$supl1:S12$

Aim$$Comparative analysis of the gut microbial community composition of the Atlantic Cod in individual fish.

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F01# F02# F03# F04# F05# F06# F07# F08# F09# F10# F11#

Order%&%Re

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Clostridiales#

Bacteroidales#

Vibrionales#

Erysipelotrichales#

Fusobacteriales#

Alteromonadales#

Desulfovibrionales#

Mycoplasmatales#

Spirochaetales#

Flavobacteriales#

VicIvallales#

Verrucomicrobiales#

Deferribacterales#

RickeLsiales#

Sphingobacteriales#

Rhodospirillales#

Other#orders#

Not#assigned#

Figure 2. Order level diversity found in Cod gut communities. Barplot with relative abundances of classified representative OTU sequences. Sequences were compared with the SILVA SSUref V108 NR database using blastN. Blastn results were analyzed with MEGAN.

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Phylum

'('Re

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Firmicutes#

Proteobacteria#

Bacteroidetes#

Fusobacteria#

Tenericutes#

Bacteria#

Spirochaetes#

Deferribacteres#

Verrucomicrobia#

LenEsphaerae#

Not#assigned#

Chordata#

Ciliophora#

candidate#division#TM6#

Acidobacteria#

Bacteria#environmental#samples#

Cyanobacteria#

Chlamydiae#

AcEnobacteria#

Figure 1. Phylum level diversity found in Cod gut communities. Barplot with relative abundances of classified representative OTU sequences. Sequences were compared with the SILVA SSUref V108 NR database using blastN. Blastn results were analyzed with MEGAN.

a$ b$

Figure 3. Sequencing effort for each of the 11 fish. a) Rarefaction curves of the 16S rRNA V3 region pyrosequences using a pairwise sequence similarity cut-off of 97%. b) Rank abundance plot showing the OTU (97%) abundance distribution in each Cod gut community.

Figure 4. Pairwise similarity matrix based on OTU (97%) community structure. Distances between samples were calculated using the Bray-Curtis similarity index. 1 means a highly similar community structure and a low beta-diversity