Abstract
BackgroundThe effects of gut microbiota on human traits are expected to be small to moderate and adding the complexity of the human diseases, microbiome research demands big sample sizes. Fecal samples for such studies are mostly self-collected by participants at home. This imposes an extra level of complexity as sample collection and storage can be challenging. Effective, low-burden collection and storage methods allowing fecal samples to be transported properly and ensuring optimal quality and quantity of bacterial DNA for upstream analyses are necessary. Moreover, accurate assessment of the microbiome composition also depends on bacterial DNA extraction method. The aim of this study was to evaluate the reliability and efficiency of the OMNIgene•GUT kit as a participant-fecal friendly collection method (storage at room temperature for 24 h (O24h) or 7 days (O7d)) in comparison to the standard collection method (Fresh, storage at 4 °C for less than 24 h) in terms of amount of variability and information content accounting for two common DNA extraction methods.ResultsFourteen fecal samples were collected from healthy individuals (7 males, 7 females). Collection and storage methods did not differ significantly in terms of DNA concentration and Shannon diversity index. Phylum relative abundance showed significant differences for Bacteroidetes, Actinobacteria and Cyanobacteria. The differences were observed between control (Fresh) and O24h methods, but not between Fresh and O7d. These differences were not seen when performing bacterial DNA quantification based on three bacterial groups: Bacteroides spp., Bifidobacterium spp. and Clostridium cluster IV, which represent three major phyla: Bacteroidetes, Actinobacteria and Firmicutes respectively. The two DNA extraction methods differ in terms of DNA quantity, quality, bacterial diversity and bacterial relative abundance. Furthermore, principal component analysis revealed differences in microbial structure, which are driven by the DNA extraction methods more than the collection/storage methods.ConclusionOur results have highlighted the potential of using the OMNIgene•GUT kit for collection and storage at ambient temperature, which is convenient for studies aiming to collect large samples by giving participants the possibility to send samples by post. Importantly, we revealed that the choice of DNA extraction method have an impact on the microbiome profiling.
Highlights
The effects of gut microbiota on human traits are expected to be small to moderate and adding the complexity of the human diseases, microbiome research demands big sample sizes
In the present study, (i) we compared two collection and storage methods (Fresh; OMNIgeneGUT) and (ii) we accounted for two common DNA extraction methods (QIA; PowerFecal® DNA Isolation Kit (PF)) experimental design can be found in Additional file 1: Figure S1 and Additional file 2: Figure S2, respectively
The DNA concentration of the samples collected with the OMNIgeneGUT kit after 7 days of storage had lower values compared to OMNIgene•GUT kit and stored for 24 h (O24h) (Table 2)
Summary
The effects of gut microbiota on human traits are expected to be small to moderate and adding the complexity of the human diseases, microbiome research demands big sample sizes. Fecal samples for such studies are mostly self-collected by participants at home. Current methods of collecting fecal samples demand high involvement from participants to set up appointments on the collection day and storing the samples in their own fridge or freezer, which can bring discomfort for the participants and potentially represent a health risk Those factors might negatively affect participants’ decision to take part in a study. Use of such kits enables participants to send their samples via regular post without the need of making appointments, refrigeration, or coldchain transportation and might increase participation for (future) research studies
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