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. 2024 Sep 2:11:1405470.
doi: 10.3389/fvets.2024.1405470. eCollection 2024.

Species-level characterization of the core microbiome in healthy dogs using full-length 16S rRNA gene sequencing

Affiliations

Species-level characterization of the core microbiome in healthy dogs using full-length 16S rRNA gene sequencing

Connie A Rojas et al. Front Vet Sci. .

Abstract

Despite considerable interest and research in the canine fecal microbiome, our understanding of its species-level composition remains incomplete, as the majority of studies have only provided genus-level resolution. Here, we used full-length 16S rRNA gene sequencing to characterize the fecal microbiomes of 286 presumed healthy dogs living in homes in North America who are devoid of clinical signs, physical conditions, medication use, and behavioral problems. We identified the bacterial species comprising the core microbiome and investigated whether a dog's sex & neuter status, age, body weight, diet, and geographic region predicted microbiome variation. Our analysis revealed that 23 bacterial species comprised the core microbiome, among them Collinsella intestinalis, Megamonas funiformis, Peptacetobacter hiranonis, Prevotella copri, and Turicibacter sanguinis. The 23 taxa comprised 75% of the microbiome on average. Sterilized females, dogs of intermediate body sizes, and those exclusively fed kibble tended to harbor the most core taxa. Host diet category, geographic region, and body weight predicted microbiome beta-diversity, but the effect sizes were modest. Specifically, the fecal microbiomes of dogs fed kibble were enriched in several core taxa, including C. intestinalis, P. copri, and Holdemanella biformis, compared to those fed raw or cooked food. Conversely, dogs on a raw food diet exhibited higher abundances of Bacteroides vulgatus, Caballeronia sordicola, and Enterococcus faecium, among others. In summary, our study provides novel insights into the species-level composition and drivers of the fecal microbiome in healthy dogs living in homes; however, extrapolation of our findings to different dog populations will require further study.

Keywords: 16S rRNA gene sequencing; PacBio; body weight; canine fecal microbiome; core microbiome; diet; dogs; geography.

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Conflict of interest statement

CR, BP, ES, GJ, ZE, JJ, AM, and HG are employees of AnimalBiome, a pet health company that maintains a veterinary canine and feline stool bank. The authors declare that this study received funding from AnimalBiome. The funder was involved in the study design, collection, analysis, interpretation of data, and the writing of this article.

Figures

Figure 1
Figure 1
Fecal microbiomes differ between verified healthy and presumed healthy dogs. Dogs in the healthy reference set were categorized as “verified healthy” (VH) if they were verified to be healthy via medical records, veterinary visits, and monthly parasite screenings, or “presumed healthy” (PH) if they were not and were reported to be healthy by their owners. (A) Number of core taxa for verified healthy vs. presumed healthy dogs. (B) PCoA ordination based on Aitchison distances showing the clustering of verified healthy vs. presumed healthy microbiomes.
Figure 2
Figure 2
Relative abundances of bacterial species comprising the core microbiome in healthy dogs. These bacterial species were found in at least 33% of dogs at a mean relative abundance >0.5%. UC, unclassified.
Figure 3
Figure 3
The number of core taxa present in canine fecal microbiomes varies with sex & neuter status, age, body weight, and diet. All dogs that were part of the dataset had BCS 4–6, and no history of antibiotics, medications, bacterial probiotics, and physical conditions. (A) Number of core taxa (max 23) for each Sex-Neuter category. (B) Proportion of the microbiome made up of core taxa regressed against age, with smooth curve overlaid to illustrate relationship between x and y. (C) Number of core taxa plotted against body weight in kg. (D) Number of core taxa for each diet category. *p < 0.05, **p < 0.01, ***p < 0.0001.
Figure 4
Figure 4
Microbiome alpha-diversity varies with dog sex & neuter status, age, body weight, and diet. (A) Chao 1 Richness by Sex-Neuter status (Female intact, Female sterilized, Male intact, Male sterilized). (B,C) Shannon diversity plotted against age in years or body weight in kg, with smooth curve overlaid to illustrate relationship between x and y. (D) Shannon diversity for each diet category. *p < 0.05, **p < 0.01, ***p < 0.0001.
Figure 5
Figure 5
Host correlates of fecal microbiome beta-diversity in healthy dogs. PCoA ordinations based on Aitchison distances color coded by (A) sex & neuter status, (B) age (yrs), (C) body weight (kg), (D) diet, or (E) geographic region. PERMANOVA R2 and p-values are shown.
Figure 6
Figure 6
Bacterial species enriched in the microbiomes of young dogs and large dogs. Results from LinDA differential abundance analyses performed at the bacterial species level. (A,B) The LinDA model included age (yrs) and body weight (kg) as continuous variables, and sex & neuter status as a categorical variable, though no statistically significant taxa emerged for the categorical predictor.
Figure 7
Figure 7
Bacterial species enriched in the microbiomes of dogs according to their diet category. Results from LinDA differential abundance analyses performed at the bacterial species level. The LinDA model included diet category (Kibble, Raw, or Cooked) as the dependent variable. (A) Dogs fed Kibble vs. Dogs fed Raw food. (B) Dogs fed Kibble vs. Dogs fed Cooked food. No bacterial taxa were differentially abundant between dogs fed cooked food compared to raw food, hence why these plots are not displayed.

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Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work was supported by AnimalBiome and by 169 backers on Kickstarter.

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