Do Allergen Challenges Alter the Skin and Gut Microbiome in msAD dogs

The objectives of this study are to evaluate the shifts in the microbiome in experimentally sensitized dogs predisposed to atopic dermatitis (AD). This study will lead to a better understanding of the canine skin microbiome during flares of atopic dermatitis. Ten Dermatophagoides farinae (Df)-sensitized Maltese-beagle crossbred atopic dogs will be challenged with Df-house dust mites by epicutaneous applications of mite allergens via painting. This model is considered a relevant model to study mechanisms of atopic skin lesion development and will serve to evaluate the dynamics of the cutaneous microbiota flora during flares of AD. The lyophilized D. farinae house dust mites used in this study will be bleached to eliminate bacterial DNA in the surface and intestinal tract and suspended in mineral oil. The dogs will be challenged via allergen painting in the right axilla. In order to have a baseline sample, skin and rectal swabs will be collected from the dogs prior to sensitization. Skin swabs from the sensitized site (right axilla) will be collected and skin lesions will be evaluated at 12h, 24h (time of development of first lesions), every 24 hours for 10 days, and at 14d, 21d and 28d post challenge. Swabs from the contralateral site (left axilla) will be collected at 48h, 10d, and 28d post challenge. Additionally, rectal swabs will be collected at 48h and 10d post challenge and at any time that dogs enrolled in this study develop diarrhea. Genomic DNA will be extracted from sterile swabs collected from the skin and from the rectum. Bacterial tag-encoded FLX-titanium amplicon pyrosequencing (bTEFAP) will be based upon the V1–V3 region (E. coli position 27–519) of the 16S rRNA gene and the data will be analyzed using the phylogeny-based unweighted Unifrac distance metric. Differences in the proportions of bacterial taxa between different timepoints pre and post-challenge and between challenged site and contralateral site will be will be compared statistically by repeated ANOVA and paired t-tests, respectively.