In a recent study published in the journal Nutrients, researchers used a novel, ex vivo Systemic Intestinal Fermentation Research (SIFR)® technology to assess the selectivity with which carrot-derived rhamnogalacturonan I (cRG-I) impacted the human gut microbiota.
The research was taken amid growing interest in developing targeted gut microbiota modulators, e.g., prebiotics, dietary interventions that improve human health.
Study: Carrot RG-I Reduces Interindividual Differences between 24 Adults through Consistent Effects on Gut Microbiota Composition and Function Ex Vivo. Image Credit: 5secondStudio/Shutterstock.com
Background
Recently, Lavelle et al. demonstrated that the gut microbiota composition is highly prone to clinically relevant interpersonal differences.
Though the classification of gut microbiota in distinct gut enterotypes is evolving in gut microbiota research, researchers are aware that Prevotella, Bacteroides, and Ruminococcus are the three top contributors to the microbiota variation in healthy individuals.
So if a host microorganism selectively uses a dietary substrate to confer a health benefit, interpersonal differences will impact the clinical outcome of interventions.
In other words, there is a need for prebiotics with such specificity that commonly abundant beneficial gut commensal microorganisms could selectively ferment them and confer health benefits. It could help alleviate interpersonal differences resulting in more predictable interventional outcomes.
Previous in vitro studies were limited by their small sample size. Also, in vivo microbiota and the microbiota that established itself for short- and long-term in the laboratory systems substantially altered their results.
On the contrary, the high throughput technology used in this study enabled the inclusion of 24 human adults in the study design, which helped the researchers compare the specificity of cRG-I with inulin (IN) and xanthan (XA), two traditional prebiotics with low and high specificities, respectively.
About the study
In the present study, researchers used SIFR technology, a bioreactor-based gut model, to test five experimental conditions for 24 human adults covering a 1.5 g/d of cRG-I (high dose).
In addition, they tested 0.3 g/d of cRG-I (low dose), no-substrate control (NSC), and 1.5 g/d IN and XA for comparisons.
The team carried out all experiments in bioreactors containing five mL of a nutritional medium–fecal inoculum blend supplemented with test compounds. After 48 hours of incubation, they collected liquid samples for microbial composition and metabolite analysis.
Specifically, they extracted short-chain fatty acids (SCFAs), propionate, butyrate, acetate, and valerate, and branched-chain fatty acids (bCFAs).
Further, the team used quantitative 16S ribosomal ribonucleic acid (rRNA) gene profiling for phylogenetic analysis of human gut microbiota samples and flow cytometry (FC) for total cell counts (cells/mL).
In this way, the researchers estimated absolute cell counts for taxonomic groups stratified by phyla, family, and operational taxonomic unit (OTU).
Furthermore, the researchers estimated α-diversity and β-diversity, i.e., a measure of species richness and dissimilarity between samples via the Chao1 diversity index and weighted Unifrac index, respectively. Notably, the β-diversity accounts for taxa-relatedness and their abundance.
The team compared the treatment effects of the NSC and test compounds using repeated measures ANOVA and Benjamini–Hochberg’s method [False Discovery Rate (FDR) = 0.05)] corrected p-values.
Finally, they performed statistical analysis on the log10-transformed values, where they considered a taxonomic group's limit of detection (LOD) equal to the overall LOD based on a prespecified procedure. Most importantly, they retained the 100 most abundant OTUs in the analysis.
Results
The team assessed fecal samples from 13 male & 11 female donors aged between 28 and 61. The study results confirmed that cRG-I treatment reduced interpersonal differences in gut microbial composition and metabolite synthesis by selective stimulation of common microbiome taxa.
The high complexity of the cRG-I chemical structure likely favored the growth of around 30 OTUs. In striking contrast, IN and XA enhanced interpersonal differences.
The cRG-I treatment enhanced taxa, such as acetate/propionate-synthesizing Bacteroidaceae spp., e.g., B. dorei and B. thetaiotaomicron, acetate-synthesizing Bifidobacteriaceae longum and B. adolescentis, and butyrate-synthesizing bacterial species, e.g., Faecalibacterium prausnitzii. Based on observed increased OTUs, B. dorei and B. longum emerged as keystone species for cRG-I fermentation.
Indeed, analyzing large sample sizes is the key to establishing a correlation between metabolites and bacterial species and understanding the mechanical impact of cRG-I on the human gut microbiota.
Lastly, compared to IN, cRG-I resulted in markedly higher acetate (+40%), total SCFA levels (+32%), and propionate (+22%). Thus, based on fundamental fermentation parameters, cRG-I led to a profound production of SCFA and a lowered gas production, translating into better tolerability.
Conclusions
The current study highlighted the need for targeted gut microbiota modulators with more anticipated outcomes because even high-specificity fibers, such as XA, could also be overly specific. Thus, test subjects lacking highly specialized keystone XA degraders did not respond to its treatment.
Conversely, the main compound tested in this study, cRG-I, could be consistently fermented by keystone species commonly abundant in human adults regardless of their enterotype.
All 24 donors tested in this study, thus, fermented cRG-I, resulting in a remarkable impact on microbiota composition and functionality, as indicated by fermentation parameters. Overall, cRG-I emerged as a medium-high-specificity dietary fiber with well-recognized benefits on human health.
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