Large-scale human microbiota studies have generated key insights into a variety of disease processes, including cardiovascular disease, carcinogenesis, diabetes, hypertension, and inflammatory bowel disease. However, the utility of such studies is contingent on accurate and consistent representation of microbial communities.

Stool preservatives are commonly used to prevent shifts in microbial community composition which result in bias during the period between specimen collection and processing. This is particularly important in multisite studies where cold-chain transport—the “gold standard” for microbiota preservation—is not realistic. In the present study, researchers from the Centre for Gut Microbiota Research in Hong Kong evaluated the performance of six stool preservation solutions (Norgen, OMNI, RNAlater, CURNA, HEMA, and Shield) in inactivating infectious agents and retaining gut microbiota composition compared to immediate freezing at −80°C. 

Following ambient temperature storage of five human stool samples with the aforementioned preservatives for seven days, three hypervariable regions of the bacterial 16S rRNA gene (V1-V2, V3-V4, and V4) were amplicon sequenced. The authors observed that samples collected in Norgen’s Stool Nucleic Acid Preservative consistently showed the least shift in microbial community composition relative to −80°C standards compared with other preservatives, whilst effectively inhibiting the growth of aerobic and anaerobic bacteria. Intersubject variation was associated with the greatest differences in microbiota composition; however, the authors note that the effect of the preservative was sufficient to introduce shifts in community composition and subsequent bias in downstream applications. Further, when microbiome profiles inferred from different 16S rRNA gene regions were compared,  the authors observed differential sensitivity among primer sets.

Therein, choice of preservative for stool sample transportation and of 16S rRNA primers are important factors affecting microbial community composition. In their conclusion, the authors recommend the use of a preservative such as Norgen’s Stool Nucleic Acid Preservative that can minimize shifts in microbial community composition and inactivate bacterial growth, enabling safe and stable stool transportation in human microbiota studies.