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Research Article
Suman Ranjan Das
Vanderbilt University Medical Center
Corresponding Author
ORCiD: https://orcid.org/0000-0003-2496-9724
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: The cotton rat (genus Sigmodon) is an essential small animal model for the study of human infectious disease and viral therapeutic development. However, the impact of the host microbiome on infection outcomes has not been explored in this model, partly due to the lack of a comprehensive characterization of microbial communities across different cotton rat species. Understanding the dynamics of their microbiome could significantly help to better understand its role during when modeling viral infections in this small animal model.
Results: We examined the bacterial communities of the gut and three external sites (skin, ear, and nose) of two inbred species of cotton rats commonly used in research (S. hispidus and S. fulviventer) by using 16S rRNA gene sequencing, constituting the first comprehensive catalog of the cotton rat microbiome. We showed that S. fulviventer maintained higher alpha diversity and richness than S. hispidus at external sites (skin, ear, nose), but there were no differentially abundant genera. However, S. fulviventer and S. hispidus had distinct fecal microbiomes composed of several significantly differentially abundant genera. Whole metagenomic shotgun sequencing of fecal samples identified species-level differences between S. hispidus and S. fulviventer, as well as different metabolic pathway functions as a result of differential host microbiome contributions. Furthermore, the microbiome composition of the external sites showed significant sex-based differences while fecal communities were not largely different.
Conclusions: Our study shows that host genetic background potentially exerts homeostatic pressures, resulting in distinct microbiomes for two different inbred cotton rat species. Because of the numerous studies that have uncovered strong relationships between host microbiome, viral infection outcomes, and immune responses, our findings represent a strong contribution for understanding the impact of different microbial communities on viral pathogenesis. Furthermore, we provide novel cotton rat microbiome data as a springboard to uncover the full therapeutic potential of the microbiome against viral infections.
Keywords
microbiome, metagenomics, cotton rat, Sigmodon, 16S rRNA gene, gut, skin, respiratory
Figure 1
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Figure 2
Figure 2
Figure 3
Figure 3
Figure 4
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Figure 5
Figure 5
This is a list of supplementary files associated with this preprint. Click to download.
- Table1.pdf
Percentage abundance of the most abundant phyla identified with 16S rRNA gene sequencing in both S.fulviventer/S.hispidus (listed respectively).
- Table1.pdf
Percentage abundance of the most abundant phyla identified with 16S rRNA gene sequencing in both S.fulviventer/S.hispidus (listed respectively).
- FigureS1.pdf
Top 20 most abundant bacterial genera at each body site of S. hispidus and S. fulviventer. In both species of cotton rats, external sites (skin, ear, nose) shared similar dominating genera while there were notable difference in gut taxa between S. hispidus and S. fulviventer. Not all reads were able to be classified down to the genus level; the lowest taxonomic level available is reported. The letter after the classification denotes the taxonomic level (i.e., g for genus, f for family, o for order, c for class, p for phylum).
- FigureS1.pdf
Top 20 most abundant bacterial genera at each body site of S. hispidus and S. fulviventer. In both species of cotton rats, external sites (skin, ear, nose) shared similar dominating genera while there were notable difference in gut taxa between S. hispidus and S. fulviventer. Not all reads were able to be classified down to the genus level; the lowest taxonomic level available is reported. The letter after the classification denotes the taxonomic level (i.e., g for genus, f for family, o for order, c for class, p for phylum).
- FigureS2.pdf
Chao1 and Simpson alpha diversity indices of ear, nose, skin, and feces between S. fulviventer and S. hispidus. Statistical testing between each cotton rat species was performed using a Student’s t-test. Statistical testing between body sites is not shown (no significant differences across external sites). ns=P>0.05, *=P≤0.05, **=P≤0.01, ***=P≤0.001, ****=P≤0.0001.
- FigureS2.pdf
Chao1 and Simpson alpha diversity indices of ear, nose, skin, and feces between S. fulviventer and S. hispidus. Statistical testing between each cotton rat species was performed using a Student’s t-test. Statistical testing between body sites is not shown (no significant differences across external sites). ns=P>0.05, *=P≤0.05, **=P≤0.01, ***=P≤0.001, ****=P≤0.0001.
- FigureS3.pdf
PCoA plots comparing Bray-Curtis dissimilarities between body sites in both (A) S. hispidus and (B) S. fulviventer.
- FigureS3.pdf
PCoA plots comparing Bray-Curtis dissimilarities between body sites in both (A) S. hispidus and (B) S. fulviventer.
- FigureS4.pdf
Difference in body site beta diversity between S. hispidus and S. fulviventer. Clustering of samples (Bray-Curtis, OTU level) shows separation by host species (A) Ear, (B) Skin, (C) Nose, (D) Feces.
- FigureS4.pdf
Difference in body site beta diversity between S. hispidus and S. fulviventer. Clustering of samples (Bray-Curtis, OTU level) shows separation by host species (A) Ear, (B) Skin, (C) Nose, (D) Feces.
- FigureS5.pdf
Statistically significant differentially abundant bacteria genera between S. hispidus and S. fulviventer determined by GeneSelector.
- FigureS5.pdf
Statistically significant differentially abundant bacteria genera between S. hispidus and S. fulviventer determined by GeneSelector.
- FigureS6.pdf
Alpha diversity metrics of ear, nose, skin, and feces between male and female S. fulviventer and S. hispidus. Richness and diversity were determined using the following methods: A) Observed OTUs, B) Chao1 index, C) Shannon Diversity Index, and D) Simpson Diversity Index. Statistical testing between gender of each cotton rat species was performed using a one-way (feces) and two-way (external sites) ANOVA, followed by a Tukey’s post-hoc test. The p-value as a result of all comparisons is shown in the top right; pairwise comparisons that were found to be significant with the Tukey post-hoc test are denoted by a bar and asterisk above the groups being compared. Statistical testing across species is not shown. Feces were plotted separately to account for the discrepancy between the Y axes. ns=P>0.05, *=P≤0.05, **=P≤0.01, ***=P≤0.001, ****=P≤0.0001.
- FigureS6.pdf
Alpha diversity metrics of ear, nose, skin, and feces between male and female S. fulviventer and S. hispidus. Richness and diversity were determined using the following methods: A) Observed OTUs, B) Chao1 index, C) Shannon Diversity Index, and D) Simpson Diversity Index. Statistical testing between gender of each cotton rat species was performed using a one-way (feces) and two-way (external sites) ANOVA, followed by a Tukey’s post-hoc test. The p-value as a result of all comparisons is shown in the top right; pairwise comparisons that were found to be significant with the Tukey post-hoc test are denoted by a bar and asterisk above the groups being compared. Statistical testing across species is not shown. Feces were plotted separately to account for the discrepancy between the Y axes. ns=P>0.05, *=P≤0.05, **=P≤0.01, ***=P≤0.001, ****=P≤0.0001.
- FigureS7.pdf
DESeq2 results representing differential abundance of Lactobacillus species and strains between S. hispidus and S. fulviventer. Data were generated from whole metagenome sequencing.
- FigureS7.pdf
DESeq2 results representing differential abundance of Lactobacillus species and strains between S. hispidus and S. fulviventer. Data were generated from whole metagenome sequencing.
- FigureS8.pdf
Several pathways that were more active in S. fulviventer than S. hispidus were greatly contributed to by Akkermansia species. (A) A member of the superpathway of branched chain amino acid biosynthesis, that generates not only isoleucine, but also leucine and valine (ILEUSYN-PWY). (B) Degradation of starch for the generation of carbon skeletons, reductants, and ATP for anabolic bacterial fatty acid pathway initiation via pyruvate decarboxylation to acetyl CoA (PWY-1042). (C) Biosynthesis of R-4'-phosphopantothenate, the universal precursor for the synthesis of coenzyme A and acyl carrier protein. Only plants and microorganisms can synthesize pantothenate de novo; animals require a dietary supplement. Synonymous with Vitamin B5 synthesis. (PANTO-PWY). (D) A member of the superpathway of branched chain amino acid biosynthesis, that generates not only valine, but also leucine and isoleucine.
- FigureS8.pdf
Several pathways that were more active in S. fulviventer than S. hispidus were greatly contributed to by Akkermansia species. (A) A member of the superpathway of branched chain amino acid biosynthesis, that generates not only isoleucine, but also leucine and valine (ILEUSYN-PWY). (B) Degradation of starch for the generation of carbon skeletons, reductants, and ATP for anabolic bacterial fatty acid pathway initiation via pyruvate decarboxylation to acetyl CoA (PWY-1042). (C) Biosynthesis of R-4'-phosphopantothenate, the universal precursor for the synthesis of coenzyme A and acyl carrier protein. Only plants and microorganisms can synthesize pantothenate de novo; animals require a dietary supplement. Synonymous with Vitamin B5 synthesis. (PANTO-PWY). (D) A member of the superpathway of branched chain amino acid biosynthesis, that generates not only valine, but also leucine and isoleucine.
- TableS1.pdf
Differential abundance analysis of taxa between individual body site across female and male S. hispidus and S. fulviventer. Positive Log2FoldChange = higher in females; negative Log2FoldChange higher in males. There were no significant taxa in S. fulviventer feces.
- TableS1.pdf
Differential abundance analysis of taxa between individual body site across female and male S. hispidus and S. fulviventer. Positive Log2FoldChange = higher in females; negative Log2FoldChange higher in males. There were no significant taxa in S. fulviventer feces.
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Research Article
Suman Ranjan Das
Vanderbilt University Medical Center
Corresponding Author
ORCiD: https://orcid.org/0000-0003-2496-9724
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Published
View the published article at Animal Microbiome.
Version 1
Posted 19 Nov, 2020
No community comments so far
Editorial decision: Major revision
On 22 Dec, 2020
Review #2 received
Received 13 Dec, 2020
Review #1 received
Received 01 Dec, 2020
Reviewer #2 agreed
On 23 Nov, 2020
Reviewers invited
Invitations sent on 23 Nov, 2020
Reviewer #1 agreed
On 23 Nov, 2020
First submitted
On 12 Nov, 2020
Editor assigned
On 12 Nov, 2020
Submission checks complete
On 12 Nov, 2020
Editor invited
On 12 Nov, 2020
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
General Microbiology
License:
This work is licensed under a CC BY 4.0 License. Read Full License
View the published article at Animal Microbiome.
Background: The cotton rat (genus Sigmodon) is an essential small animal model for the study of human infectious disease and viral therapeutic development. However, the impact of the host microbiome on infection outcomes has not been explored in this model, partly due to the lack of a comprehensive characterization of microbial communities across different cotton rat species. Understanding the dynamics of their microbiome could significantly help to better understand its role during when modeling viral infections in this small animal model.
Results: We examined the bacterial communities of the gut and three external sites (skin, ear, and nose) of two inbred species of cotton rats commonly used in research (S. hispidus and S. fulviventer) by using 16S rRNA gene sequencing, constituting the first comprehensive catalog of the cotton rat microbiome. We showed that S. fulviventer maintained higher alpha diversity and richness than S. hispidus at external sites (skin, ear, nose), but there were no differentially abundant genera. However, S. fulviventer and S. hispidus had distinct fecal microbiomes composed of several significantly differentially abundant genera. Whole metagenomic shotgun sequencing of fecal samples identified species-level differences between S. hispidus and S. fulviventer, as well as different metabolic pathway functions as a result of differential host microbiome contributions. Furthermore, the microbiome composition of the external sites showed significant sex-based differences while fecal communities were not largely different.
Conclusions: Our study shows that host genetic background potentially exerts homeostatic pressures, resulting in distinct microbiomes for two different inbred cotton rat species. Because of the numerous studies that have uncovered strong relationships between host microbiome, viral infection outcomes, and immune responses, our findings represent a strong contribution for understanding the impact of different microbial communities on viral pathogenesis. Furthermore, we provide novel cotton rat microbiome data as a springboard to uncover the full therapeutic potential of the microbiome against viral infections.
Figure 1
Figure 1
Figure 2
Figure 2
Figure 3
Figure 3
Figure 4
Figure 4
Figure 5
Figure 5
This is a list of supplementary files associated with this preprint. Click to download.
- Table1.pdf
Percentage abundance of the most abundant phyla identified with 16S rRNA gene sequencing in both S.fulviventer/S.hispidus (listed respectively).
- Table1.pdf
Percentage abundance of the most abundant phyla identified with 16S rRNA gene sequencing in both S.fulviventer/S.hispidus (listed respectively).
- FigureS1.pdf
Top 20 most abundant bacterial genera at each body site of S. hispidus and S. fulviventer. In both species of cotton rats, external sites (skin, ear, nose) shared similar dominating genera while there were notable difference in gut taxa between S. hispidus and S. fulviventer. Not all reads were able to be classified down to the genus level; the lowest taxonomic level available is reported. The letter after the classification denotes the taxonomic level (i.e., g for genus, f for family, o for order, c for class, p for phylum).
- FigureS1.pdf
Top 20 most abundant bacterial genera at each body site of S. hispidus and S. fulviventer. In both species of cotton rats, external sites (skin, ear, nose) shared similar dominating genera while there were notable difference in gut taxa between S. hispidus and S. fulviventer. Not all reads were able to be classified down to the genus level; the lowest taxonomic level available is reported. The letter after the classification denotes the taxonomic level (i.e., g for genus, f for family, o for order, c for class, p for phylum).
- FigureS2.pdf
Chao1 and Simpson alpha diversity indices of ear, nose, skin, and feces between S. fulviventer and S. hispidus. Statistical testing between each cotton rat species was performed using a Student’s t-test. Statistical testing between body sites is not shown (no significant differences across external sites). ns=P>0.05, *=P≤0.05, **=P≤0.01, ***=P≤0.001, ****=P≤0.0001.
- FigureS2.pdf
Chao1 and Simpson alpha diversity indices of ear, nose, skin, and feces between S. fulviventer and S. hispidus. Statistical testing between each cotton rat species was performed using a Student’s t-test. Statistical testing between body sites is not shown (no significant differences across external sites). ns=P>0.05, *=P≤0.05, **=P≤0.01, ***=P≤0.001, ****=P≤0.0001.
- FigureS3.pdf
PCoA plots comparing Bray-Curtis dissimilarities between body sites in both (A) S. hispidus and (B) S. fulviventer.
- FigureS3.pdf
PCoA plots comparing Bray-Curtis dissimilarities between body sites in both (A) S. hispidus and (B) S. fulviventer.
- FigureS4.pdf
Difference in body site beta diversity between S. hispidus and S. fulviventer. Clustering of samples (Bray-Curtis, OTU level) shows separation by host species (A) Ear, (B) Skin, (C) Nose, (D) Feces.
- FigureS4.pdf
Difference in body site beta diversity between S. hispidus and S. fulviventer. Clustering of samples (Bray-Curtis, OTU level) shows separation by host species (A) Ear, (B) Skin, (C) Nose, (D) Feces.
- FigureS5.pdf
Statistically significant differentially abundant bacteria genera between S. hispidus and S. fulviventer determined by GeneSelector.
- FigureS5.pdf
Statistically significant differentially abundant bacteria genera between S. hispidus and S. fulviventer determined by GeneSelector.
- FigureS6.pdf
Alpha diversity metrics of ear, nose, skin, and feces between male and female S. fulviventer and S. hispidus. Richness and diversity were determined using the following methods: A) Observed OTUs, B) Chao1 index, C) Shannon Diversity Index, and D) Simpson Diversity Index. Statistical testing between gender of each cotton rat species was performed using a one-way (feces) and two-way (external sites) ANOVA, followed by a Tukey’s post-hoc test. The p-value as a result of all comparisons is shown in the top right; pairwise comparisons that were found to be significant with the Tukey post-hoc test are denoted by a bar and asterisk above the groups being compared. Statistical testing across species is not shown. Feces were plotted separately to account for the discrepancy between the Y axes. ns=P>0.05, *=P≤0.05, **=P≤0.01, ***=P≤0.001, ****=P≤0.0001.
- FigureS6.pdf
Alpha diversity metrics of ear, nose, skin, and feces between male and female S. fulviventer and S. hispidus. Richness and diversity were determined using the following methods: A) Observed OTUs, B) Chao1 index, C) Shannon Diversity Index, and D) Simpson Diversity Index. Statistical testing between gender of each cotton rat species was performed using a one-way (feces) and two-way (external sites) ANOVA, followed by a Tukey’s post-hoc test. The p-value as a result of all comparisons is shown in the top right; pairwise comparisons that were found to be significant with the Tukey post-hoc test are denoted by a bar and asterisk above the groups being compared. Statistical testing across species is not shown. Feces were plotted separately to account for the discrepancy between the Y axes. ns=P>0.05, *=P≤0.05, **=P≤0.01, ***=P≤0.001, ****=P≤0.0001.
- FigureS7.pdf
DESeq2 results representing differential abundance of Lactobacillus species and strains between S. hispidus and S. fulviventer. Data were generated from whole metagenome sequencing.
- FigureS7.pdf
DESeq2 results representing differential abundance of Lactobacillus species and strains between S. hispidus and S. fulviventer. Data were generated from whole metagenome sequencing.
- FigureS8.pdf
Several pathways that were more active in S. fulviventer than S. hispidus were greatly contributed to by Akkermansia species. (A) A member of the superpathway of branched chain amino acid biosynthesis, that generates not only isoleucine, but also leucine and valine (ILEUSYN-PWY). (B) Degradation of starch for the generation of carbon skeletons, reductants, and ATP for anabolic bacterial fatty acid pathway initiation via pyruvate decarboxylation to acetyl CoA (PWY-1042). (C) Biosynthesis of R-4'-phosphopantothenate, the universal precursor for the synthesis of coenzyme A and acyl carrier protein. Only plants and microorganisms can synthesize pantothenate de novo; animals require a dietary supplement. Synonymous with Vitamin B5 synthesis. (PANTO-PWY). (D) A member of the superpathway of branched chain amino acid biosynthesis, that generates not only valine, but also leucine and isoleucine.
- FigureS8.pdf
Several pathways that were more active in S. fulviventer than S. hispidus were greatly contributed to by Akkermansia species. (A) A member of the superpathway of branched chain amino acid biosynthesis, that generates not only isoleucine, but also leucine and valine (ILEUSYN-PWY). (B) Degradation of starch for the generation of carbon skeletons, reductants, and ATP for anabolic bacterial fatty acid pathway initiation via pyruvate decarboxylation to acetyl CoA (PWY-1042). (C) Biosynthesis of R-4'-phosphopantothenate, the universal precursor for the synthesis of coenzyme A and acyl carrier protein. Only plants and microorganisms can synthesize pantothenate de novo; animals require a dietary supplement. Synonymous with Vitamin B5 synthesis. (PANTO-PWY). (D) A member of the superpathway of branched chain amino acid biosynthesis, that generates not only valine, but also leucine and isoleucine.
- TableS1.pdf
Differential abundance analysis of taxa between individual body site across female and male S. hispidus and S. fulviventer. Positive Log2FoldChange = higher in females; negative Log2FoldChange higher in males. There were no significant taxa in S. fulviventer feces.
- TableS1.pdf
Differential abundance analysis of taxa between individual body site across female and male S. hispidus and S. fulviventer. Positive Log2FoldChange = higher in females; negative Log2FoldChange higher in males. There were no significant taxa in S. fulviventer feces.
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