Comparison of 95% ethanol and cards for stool metagenomic studies
Biomedical research stool sample collection methods must balance ease of use, sample preservation, and handling simplicity. 95% ethanol is commonly used due to its favorable performance, but can cause issues with shipping and DNA extraction. Collection cards are frequently used in cancer epidemiology to avoid difficulties involving solvents. However, relatively few studies use stool microbiome-specific card methods, and their performance against 95% ethanol is not well established.Here, we compared shotgun metagenomic data generated from 32 paired samples from a custom stool card and 95% ethanol-preserved sample. The agreement between methods was evaluated at the whole community and individual species level. Method-biased species were compared to a recently generated list of colorectal cancer-associated species to contextualize card performance. Stool cards generally agreed with 95% ethanol, though some taxa were differentially abundant between methods. Streptococci seem to be underrepresented systematically on cards, and Escherichia coli blooms were occasionally observed. Differences between methods were not enriched within the set of colorectal cancer-associated species.Both stool cards or 95% ethanol perform well, but differences should be accounted for, especially when mixing collection methods within a study. Community-level deviations can be regarded similarly to typical batch effects and adjusted as such, while species-level biases may require direct attention depending on phenotype of interest.