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RNA-Seq analysis of seasonal and individual variation in blood transcriptomes of healthy managed bottlenose dolphins

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ABSTRACT

Background: The blood transcriptome can reflect both systemic exposures and pathological changes in other organs of the body because immune cells recirculate through the blood, lymphoid tissues, and affected sites. In human and veterinary medicine, blood transcriptome analysis has been used successfully to identify markers of disease or pathological conditions, but can be confounded by large seasonal changes in expression. In comparison, the use of transcriptomic based analyses in wildlife has been limited. Here we report a longitudinal study of four managed bottlenose dolphins located in Waikoloa, Hawaii, serially sampled (approximately monthly) over the course of 1 year to establish baseline information on the content and variation of the dolphin blood transcriptome.

Results: Illumina based RNA-seq analyses were carried out using both the Ensembl dolphin genome and a de novo blood transcriptome as guides. Overall, the blood transcriptome encompassed a wide array of cellular functions and processes and was relatively stable within and between animals over the course of 1 year. Principal components analysis revealed moderate clustering by sex associated with the variation among global gene expression profiles (PC1, 22 % of variance). Limited seasonal change was observed, with < 2.5 % of genes differentially expressed between winter and summer months (FDR < 0.05). Among the differentially expressed genes, cosinor analysis identified seasonal rhythmicity for the observed changes in blood gene expression, consistent with studies in humans. While the proportion of seasonally variant genes in these dolphins is much smaller than that reported in humans, the majority of those identified in dolphins were also shown to vary with season in humans. Gene co-expression network analysis identified several gene modules with significant correlation to age, sex, or hematological parameters.

Conclusions: This longitudinal analysis of healthy managed dolphins establishes a preliminary baseline for blood transcriptome analysis in this species. Correlations with hematological parameters, distinct from muted seasonal effects, suggest that the otherwise relatively stable blood transcriptome may be a useful indicator of health and exposure. A robust database of gene expression in free-ranging and managed dolphins across seasons with known adverse health conditions or contaminant exposures will be needed to establish predictive gene expression profiles suitable for biomonitoring.

Electronic supplementary material: The online version of this article (doi:10.1186/s12864-016-3020-8) contains supplementary material, which is available to authorized users.

No MeSH data available.


Top ten Gene Ontology (GO) annotations (level 6) from the 100 most highly expressed transcripts in dolphin blood. a All of the top 100 expressed genes mapping to the dolphin genome were annotated in Blast2GO. b Ninety-two of the top 100 expressed genes mapping to the de novo blood transcriptome were annotated in Blast2GO
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Fig2: Top ten Gene Ontology (GO) annotations (level 6) from the 100 most highly expressed transcripts in dolphin blood. a All of the top 100 expressed genes mapping to the dolphin genome were annotated in Blast2GO. b Ninety-two of the top 100 expressed genes mapping to the de novo blood transcriptome were annotated in Blast2GO

Mentions: Overall approximately 85 % of reads mapped back to the dolphin genome. However, only 28.5 % of reads mapped back to annotated genes in the genome with Bowtie2. This indicated that many reads mapped outside of annotated regions of the genome and were consequently excluded from further analyses; therefore improved annotation of the Ensembl dolphin genome may greatly expand data interpretation. In order to more accurately compare the dolphin blood transcriptome to the Ensembl genome, we selected a suite of 17,475 sequences, comprised of coding sequences and pseudogenes, from the genome. Pseudogenes were included after identifying reads from the transcriptome aligning to regions of the genome annotated as such. Reads mapped to 9610, 45.2 %, of these genes identified in the Ensembl dolphin genome, with FPKM > 0 in at least half the samples and an average FPKM ≥ 1, similar to the percentage expressed in human blood [35]. Among the 100 most highly expressed genes in dolphin blood (avg FPKM from globin depleted samples), 100 % were annotated and were dominated by transcripts associated with ribosomes, translation, and DNA and RNA binding (Fig. 2a). Many of these terms are also among the most highly expressed transcripts in human blood. Likewise, transcripts mapping to GO terms involved with immune response, transcription, cell cycle and proliferation, signaling, and structural components or functions are highly expressed in both human and dolphin blood [38].Fig. 2


RNA-Seq analysis of seasonal and individual variation in blood transcriptomes of healthy managed bottlenose dolphins
Top ten Gene Ontology (GO) annotations (level 6) from the 100 most highly expressed transcripts in dolphin blood. a All of the top 100 expressed genes mapping to the dolphin genome were annotated in Blast2GO. b Ninety-two of the top 100 expressed genes mapping to the de novo blood transcriptome were annotated in Blast2GO
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC5016863&req=5

Fig2: Top ten Gene Ontology (GO) annotations (level 6) from the 100 most highly expressed transcripts in dolphin blood. a All of the top 100 expressed genes mapping to the dolphin genome were annotated in Blast2GO. b Ninety-two of the top 100 expressed genes mapping to the de novo blood transcriptome were annotated in Blast2GO
Mentions: Overall approximately 85 % of reads mapped back to the dolphin genome. However, only 28.5 % of reads mapped back to annotated genes in the genome with Bowtie2. This indicated that many reads mapped outside of annotated regions of the genome and were consequently excluded from further analyses; therefore improved annotation of the Ensembl dolphin genome may greatly expand data interpretation. In order to more accurately compare the dolphin blood transcriptome to the Ensembl genome, we selected a suite of 17,475 sequences, comprised of coding sequences and pseudogenes, from the genome. Pseudogenes were included after identifying reads from the transcriptome aligning to regions of the genome annotated as such. Reads mapped to 9610, 45.2 %, of these genes identified in the Ensembl dolphin genome, with FPKM > 0 in at least half the samples and an average FPKM ≥ 1, similar to the percentage expressed in human blood [35]. Among the 100 most highly expressed genes in dolphin blood (avg FPKM from globin depleted samples), 100 % were annotated and were dominated by transcripts associated with ribosomes, translation, and DNA and RNA binding (Fig. 2a). Many of these terms are also among the most highly expressed transcripts in human blood. Likewise, transcripts mapping to GO terms involved with immune response, transcription, cell cycle and proliferation, signaling, and structural components or functions are highly expressed in both human and dolphin blood [38].Fig. 2

View Article: PubMed Central - PubMed

ABSTRACT

Background: The blood transcriptome can reflect both systemic exposures and pathological changes in other organs of the body because immune cells recirculate through the blood, lymphoid tissues, and affected sites. In human and veterinary medicine, blood transcriptome analysis has been used successfully to identify markers of disease or pathological conditions, but can be confounded by large seasonal changes in expression. In comparison, the use of transcriptomic based analyses in wildlife has been limited. Here we report a longitudinal study of four managed bottlenose dolphins located in Waikoloa, Hawaii, serially sampled (approximately monthly) over the course of 1 year to establish baseline information on the content and variation of the dolphin blood transcriptome.

Results: Illumina based RNA-seq analyses were carried out using both the Ensembl dolphin genome and a de novo blood transcriptome as guides. Overall, the blood transcriptome encompassed a wide array of cellular functions and processes and was relatively stable within and between animals over the course of 1 year. Principal components analysis revealed moderate clustering by sex associated with the variation among global gene expression profiles (PC1, 22 % of variance). Limited seasonal change was observed, with < 2.5 % of genes differentially expressed between winter and summer months (FDR < 0.05). Among the differentially expressed genes, cosinor analysis identified seasonal rhythmicity for the observed changes in blood gene expression, consistent with studies in humans. While the proportion of seasonally variant genes in these dolphins is much smaller than that reported in humans, the majority of those identified in dolphins were also shown to vary with season in humans. Gene co-expression network analysis identified several gene modules with significant correlation to age, sex, or hematological parameters.

Conclusions: This longitudinal analysis of healthy managed dolphins establishes a preliminary baseline for blood transcriptome analysis in this species. Correlations with hematological parameters, distinct from muted seasonal effects, suggest that the otherwise relatively stable blood transcriptome may be a useful indicator of health and exposure. A robust database of gene expression in free-ranging and managed dolphins across seasons with known adverse health conditions or contaminant exposures will be needed to establish predictive gene expression profiles suitable for biomonitoring.

Electronic supplementary material: The online version of this article (doi:10.1186/s12864-016-3020-8) contains supplementary material, which is available to authorized users.

No MeSH data available.