Effective low-cost preservation of human stools in field-based studies for helminth and microbiota analysis. Issue 9 (August 2021)

• Record Type: Journal Article
• Authors: Stracke, Katharina; Adisakwattana, Poom; Phuanukoonnon, Suparat; Yoonuan, Tippayarat; Poodeepiyasawat, Akkarin; Dekumyoy, Paron; Chaisiri, Kittipong; Roth Schulze, Alexandra; Wilcox, Stephen; Karunajeewa, Harin; Traub, Rebecca J.; Jex, Aaron R.
• Is Part Of: International journal for parasitology. Volume 51:Issue 9(2021)
• Page Start: 741
• Page End: 748
• Publication Date: 2021-08
• Subjects: Gut microbiota -- 16S rRNA gene -- Next-generation sequencing -- Soil-transmitted helminths -- Stool preservation; Parasitology -- Periodicals; Parasitology -- Periodicals; Parasitologie -- Périodiques; Parasitology; Periodicals; Electronic journals; 571.999
• DOI: 10.1016/j.ijpara.2021.01.002
• Languages: English
• ISSNs: 0020-7519

Abstract

Graphical abstract: Highlights: Stool samples in epidemiological studies require preservation for molecular analysis. 16S rRNA gene sequencing relies on sample preservation to avoid bacterial degradation. The gut microbiome of Thai pre- and school-aged children was characterised. No variation was found in bacterial alpha-diversity among DESS, PD and frozen storage. DESS buffer effectively preserves stools containing soil-transmitted helminth DNA. Abstract: Molecular studies of gastrointestinal infections or microbiotas require either rapid sample processing or effective interim preservation. This is difficult in remote settings in low-income countries, where the majority of the global infectious disease burden exists. Processing or freezing of samples immediately upon collection is often not feasible and the cost of commercial preservatives is prohibitive. We compared fresh freezing (the 'gold standard' method), with low-cost chemical preservation in (i) a salt-based buffer consisting of DMSO, EDTA and NaCl (DESS) or (ii) 2.5% potassium dichromate (PD), for soil-transmitted helminth detection and microbiota characterisation in pre-school and school-aged children from north-western Thailand. Fresh frozen samples were frozen at −20°C on collection and maintained at −80°C within ~3 days of collection until molecular analysis, with international shipping on dry ice. In contrast, chemically preserved samples were collected and stored at ~4°C, transported on wet ice and only stored at −20°C on arrival in Australia ~8 weeks after collection, with international shipping on wet ice. DESS and PD provided better sensitivity for STH diagnosis, estimating higher infection rates (>80% for Ascaris lumbricoides and >60% for Trichuris trichiura ; versus 56% and 15% for these parasites in fresh frozen samples) and egg abundance (inferred as gene copy number estimates). All methods performed similarly for microbiota preservation, showing no significant differences in alpha-diversity based on overall richness or inverted Simpson's Index. All three methods performed similarly for RNA and protein preservation in a small subset of samples. Overall, DESS provided the best performance, with the added benefit of being non-toxic, compared with PD, hence making it particularly applicable for studies in remote and resource-poor settings.