Cod (Gadus morhua) in subdivisions 24–32, eastern Baltic ...
Poster ISME 2012 - Gut community analysis of Atlantic Cod (Gadus morhua)
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Gut community analysis of Atlan0c Cod (Gadus morhua) Thomas HA Haverkamp 1 ([email protected]), B Star 1 ,S Jento< 1 , KS Jakobsen 1 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. 0% 10% 20% 30% 40% 50% 60% 70% 80% 90% 100% F01 F02 F03 F04 F05 F06 F07 F08 F09 F10 F11 Order & Representatvie OTUs (97%) 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. 0% 10% 20% 30% 40% 50% 60% 70% 80% 90% 100% F01 F02 F03 F04 F05 F06 F07 F08 F09 F10 F11 Phylum ( Representa1ve OTUs (97%) 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
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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.
Transcript of Poster ISME 2012 - Gut community analysis of Atlantic Cod (Gadus morhua)
Gut$community$analysis$$of$Atlan0c$Cod$(Gadus&morhua)$
Thomas'HA'Haverkamp1'([email protected]),$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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Order%&%Re
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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
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Firmicutes#
Proteobacteria#
Bacteroidetes#
Fusobacteria#
Tenericutes#
Bacteria#
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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.
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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
