DOI: 10.5061/dryad.tb2rbp05m
title: Data from: Gut microbiome dysbiosis is associated with host genetics in the Norwegian Lundehund

description: A group of diseases have been shown to correlate with a phenomenon called
 microbiome dysbiosis, where the bacterial species composition of the gut
 becomes abnormal. The gut microbiome of an animal is influenced by many
 factors including diet, exposures to bacteria during post-gestational
 growth, lifestyle, and disease status. Studies also show that host
 genetics can affect microbiome composition. We sought to test whether host
 genetic background is associated with gut microbiome composition in the
 Norwegian Lundehund dog, a highly inbred breed with an effective
 population size of 13 individuals. The Lundehund has a high rate of a
 protein-losing enteropathy in the small intestine that is often reported
 as Lundehund syndrome, which negatively affects longevity and
 life-quality. An outcrossing project with the Buhund, Norrbottenspets and
 Icelandic sheepdog was recently established to reintroduce genetic
 diversity to the Lundehund and improve its health. To assess whether there
 was an association between host genetic diversity and the microbiome
 composition, we sampled the fecal microbiomes of 75 dogs of the parental
 (Lundehund), F1 (Lundehund x Buhund), and F2 (F1 x Lundehund) generations.
 We found significant variation in microbiome composition from the parental
 Lundehund generation compared to the outcross progeny. The variation
 observed in purebred Lundehunds corresponded to dysbiosis as seen by a
 highly variable microbiome composition with an elevated Firmicutes to
 Bacteroidetes ratio and an increase in the prevalence of Streptococcus
 bovis/Streptococcus equinus complex, a known pathobiont that can cause
 several diseases. We tracked several other environmental factors including
 diet, the presence of a cat in the household, living on a farm and the use
 of probiotics, but we did not find evidence of an effect of these on
 microbiome composition and alpha diversity. In conclusion, we found an
 association between host genetics and gut microbiome composition, which in
 turn may be associated with the high incidence of Lundehund syndrome in
 the purebred parental dogs.
description: In
 April to August 2021, we collected stool samples from Lundehund (P, n =
 50), Lundehund x Buhund crosses (F1, n = 8), and F1 x Lundehund crosses
 (F2, n = 22). The lower number of individuals in the F1 cohort is due to
 the challenge in finding owners of Buhund females that were willing to let
 their dog being paired with a Lundehund. Owners were instructed in how to
 collect and handle fresh naturally deposed samples, avoiding
 contamination. The stool samples were stored at room temperature in Stool
 Nucleic Acid Collection and Preservation Tubes (Norgen BioTek Corp, Cat.
 45660). In September 2021, the samples were analyzed with the ZymoBIOMICS®
 Targeted Sequencing Service (Zymo Research, Irvine, CA). The
 ZymoBIOMICS®-96 MagBead DNA Kit (Zymo Research, Irvine, CA) was used to
 extract DNA using an automated platform. Bacterial 16S ribosomal RNA gene
 targeted sequencing was performed using the Quick-16S™ NGS Library Prep
 Kit (Zymo Research, Irvine, CA). The bacterial 16S rRNA primers amplified
 the V3-V4 region of the 16S rRNA gene. The final PCR products were
 quantified with qPCR fluorescence readings and pooled together based on
 equal molarity. The final pooled library was cleaned with the
 Select-a-Size DNA Clean & Concentrator™ (Zymo Research, Irvine,
 CA), then quantified with TapeStation® (Agilent Technologies, Santa Clara,
 CA) and Qubit® (Thermo Fisher Scientific, Waltham, WA). The ZymoBIOMICS®
 Microbial Community DNA Standard (Zymo Research, Irvine, CA) was used as a
 positive control for each targeted library preparation. Negative controls
 (i.e., blank extraction control, blank library preparation control) were
 included to assess the level of bioburden carried by the wet-lab process.
 The final library was sequenced on an Illumina® MiSeq™ with a v3 reagent
 kit (600 cycles). The sequencing was performed with a 10% PhiX spike-in.
 Unique amplicon sequences variants (ASVs) were inferred from raw reads
 using the DADA2 pipeline. Potential sequencing errors and chimeric
 sequences were also removed with the DADA2 pipeline. ASVs that were
 present in only one sample, i.e., singletons, were examined for each
 cohort and removed from the dataset for clustering analysis. Taxonomy
 assignment was performed using Uclust from Qiime v.1.9.1 with the Zymo
 Research Database. Any taxa that were not represented at over 1% relative
 abundance in at least one sample were removed. Five purebred Lundehund
 dogs older than 9 years and two F2 dogs that were under medication at
 sampling were also removed from further analyses, in order to obtain a
 sample more homogenous in age and without the influence of antibiotics.
 Normalization of the data was performed by calculating the relative
 abundance for each sample by library scaling.
description: The data can be analysed with different
 softwares, we used R version 4.1.3 with RStudio version 2022.07.2 and with
 the R packages <em>Phyloseq</em> and
 <em>Microbiome</em>. More details
 about scripts are in the README.md file.