Effective low-cost preservation of human stools in field-based studies for helminth and microbiota analysis

Katharina Stracke; Poom Adisakwattana; Suparat Phuanukoonnon; Tippayarat Yoonuan; Akkarin Poodeepiyasawat; Paron Dekumyoy; Kittipong Chaisiri; Alexandra Roth Schulze; Stephen Wilcox; Harin Karunajeewa; Rebecca J Traub; Aaron R Jex

doi:10.1016/j.ijpara.2021.01.002

Effective low-cost preservation of human stools in field-based studies for helminth and microbiota analysis

Int J Parasitol. 2021 Aug;51(9):741-748. doi: 10.1016/j.ijpara.2021.01.002. Epub 2021 Mar 26.

Affiliations

¹ The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, Australia; Department of Medical Biology, The Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, Melbourne, Victoria, Australia. Electronic address: stracke.k@wehi.edu.au.
² Department of Helminthology, Faculty for Tropical Medicine, Mahidol University, Bangkok, Thailand.
³ Social and Environmental Medicine, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand.
⁴ The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, Australia.
⁵ The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, Australia; Department of Medicine - Western Health, The University of Melbourne, Melbourne, Victoria, Australia.
⁶ Faculty for Veterinary and Agricultural Sciences, The University of Melbourne, Melbourne, Victoria, Australia.
⁷ The Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, Australia; Faculty for Veterinary and Agricultural Sciences, The University of Melbourne, Melbourne, Victoria, Australia.

PMID: 33774039
DOI: 10.1016/j.ijpara.2021.01.002

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.

Keywords: 16S rRNA gene; Gut microbiota; Next-generation sequencing; Soil-transmitted helminths; Stool preservation.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Child
Feces
Helminths*
Humans
Microbiota*
Soil
Trichuris

Substances

Soil