Oral swab effectiveness as a proxy for direct ruminal sampling in Holstein dairy cows is correlated with the darkness of the swab

Authors: SKARLUPKA, JOSEPH - University Of Wisconsin; COX, MADISON - University Of Washington Medical School; STEINBERGER, ANDREW - University Of Wisconsin; SBARDELLATI, DINO - University Of Wisconsin; McClure, Jennifer; BICKHART, DEREK - Hendrix Genetics; SCHEFTGEN, ANDREW - University Of Wisconsin; ZUNIGA-CHAVEZ, IBRAHIM - University Of Wisconsin; WOLFE, LUKE - University Of Wisconsin; PAGET, ERIC - University Of Wisconsin; SKADRON, CHARLES - University Of Wisconsin; ATTIPETTY, NITHYA - University Of Wisconsin; SUEN, GARRET - University Of Wisconsin

Submitted to: Journal of Animal Science and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/30/2024
Publication Date: N/A
Citation: N/A

Interpretive Summary: The community of microbes (microbiome) of a ruminant animal is a window into the animal’s ability to digest feed, produce milk, meat, and methane, and can indicate its overall health. One way to peek into the microbiome of cattle is to collect oral swabs when they are ruminating. Swab samples can represent up to 70% of what we would find in the actual rumen of the cow using more invasive methods. These oral swabs can vary in color from dark brown to nearly white, but we currently don't know what constitutes a good sample of the rumen from these swabs. Therefore, we used sequencing methods to identify and compare microbial communities from swabs of different color. To correlate swab color to the microbial communities we developed and utilized a new imaging approach to quantify the color of each swab on a light to dark scale. We found that swabs with increasing darkness scores were significantly associated with increased bacterial diversity within each sample and most of the bacteria being associated ruminal taxa. Lighter swabs produced more bacteria categorized as belonging to known cow oral and environmental taxa rather than ruminal taxa. Our analysis of the fungal microbiome found that swabs with increasing darkness scores were associated with less diversity within each individual sample, and were also significantly associated with the ruminal solids fungal community, but not with the ruminal liquid community. Our study refines the use of oral swabs as a proxy for capturing the ruminal microbiome and demonstrates that swab color is an important factor to consider when using this approach for documenting both the bacterial and fungal communities.

Technical Abstract: Using oral swabs to collect remnants of regurgitation of stomach contents during rumination in dairy cows captures up to 70% of the ruminal bacterial community, offering potential for broad-scale population-based studies on the rumen microbiome. Use of oral swabs allows for a non-invasive method of identifying cattle with lower methane emissions and greater feed efficiency through characterization of the rumen microbiome. Swabs collected from dairy cows often vary widely with respect to sample quality, likely due to several factors such as time of sample collection and cow rumination behavior, which may limit the ability of a given swab sample to accurately represent the ruminal microbiome. One such factor is the color of the swab, which can vary significantly across a sample set. Here, we hypothesize that darker-colored swabs contain more rumen contents, thereby better representing the ruminal bacterial community than lighter-colored swabs. To address this, we collected oral swabs from 427 dairy cows and rumen samples from 13 cannulated cows on a research farm in Wisconsin and subjected them to 16S rRNA sequencing. In addition, given that little is known about the ability of oral swabs to recapitulate the ruminal fungal community, we also conducted ITS sequencing of these samples. To correlate swab color to the microbiota we developed and utilized a novel imaging approach to colorimetrically quantify each swab from a range of light to dark. We found that swabs with increasing darkness scores were significantly associated with increased bacterial alpha diversity (p < 0.05). Lighter swabs exhibited greater variation in their community structure, with many identified amplicon sequence variants (ASVs) categorized as belonging to known bovine oral and environmental taxa. Our analysis of the fungal microbiome found that swabs with increasing darkness scores were associated with decreased alpha diversity (p < 0.05) and were also significantly associated with the ruminal solids fungal community, but not with the ruminal liquid community. Our study refines the utility of oral swabs as a useful proxy for capturing the ruminal microbiome and demonstrates that swab color is an important factor to consider when using this approach for documenting both the bacterial and fungal communities.