Prebiotic fibers, polyphenols and other molecular components of food crops significantly affect the composition and function of the human gut microbiome and human health. The abundance of these, frequently uncharacterized, microbiome-active components vary within individual crop species. Here, we employ high throughput in vitro fermentations of pre-digested grain using a human microbiome to identify segregating genetic loci in a food crop, sorghum, that alter the composition and function of human gut microbes. Evaluating grain produced by 294 sorghum recombinant inbreds identifies 10 loci in the sorghum genome associated with variation in the abundance of microbial taxa and/or microbial metabolites. Two loci co-localize with sorghum genes regulating the biosynthesis of condensed tannins. We validate that condensed tannins stimulate the growth of microbes associated with these two loci. Our work illustrates the potential for genetic analysis to systematically discover and characterize molecular components of food crops that influence the human gut microbiome.

Waxy starches from cereal grains contain >90% amylopectin due to naturally occurring mutations that block amylose biosynthesis. Waxy starches have unique organoleptic characteristics (e.g. sticky rice) as well as desirable physicochemical properties for food processing. Using isogenic pairs of wild type sorghum lines and their waxy derivatives, we studied the effects of waxy starches in the whole grain context on the human gut microbiome. In vitro fermentations with human stool microbiomes show that beneficial taxonomic and metabolic signatures driven by grain from wild type parental lines are lost in fermentations of grain from the waxy derivatives and the beneficial signatures can be restored by addition of resistant starch. These undesirable effects are conserved in fermentations of waxy maize, wheat, rice and millet. We also demonstrate that humanized gnotobiotic mice fed low fiber diets supplemented with 20% grain from isogenic pairs of waxy vs. wild type parental sorghum have significant differences in microbiome composition and show increased weight gain. We conclude that the benefits of waxy starches on food functionality can have unintended tradeoff effects on the gut microbiome and host physiology that could be particularly relevant in human populations consuming large amounts of waxy grains.

Background:Waxy starches contain >90% amylopectin and are derived from grain crops carrying naturally-occurring mutations that block amylose biosynthesis. The absence of amylose in waxy starches produces unique physiochemical properties that are desirable for food processing, but the effects of increased amylopectin/amylose ratios in waxy starches on the gut microbiome and physiological characteristics of the host are not well characterized. Here, we used a whole-grain model with isogenic pairs of wild type sorghum lines and their waxy derivatives to test the hypothesis that major differences in amylose/amylopectin ratio produce significant effects on the human gut microbiome. Results:Fermentation of grain from waxy versus wild type derivatives produced substantial differences in overall microbiome composition, abundances of multiple taxa, and production of microbial metabolites (butyrate). Several of the taxonomic and metabolic signatures of fermentations from parental versus waxy lines were shared across fermentations with microbiomes from different human donors, including reduced levels of butyrate production and lower abundances of Roseburia and other amylolytic, butyrate-producing members of Lachnospiraceae in fermentations of waxy lines. Using a human microbiome-associated mouse model, we also detected significant differences in microbiome composition in animals fed low-fiber diets supplemented with 20% grain from isogenic pairs of parental versus waxy derivatives of sorghum. Remarkably, these microbiome changes were accompanied by significant differences in weight gain, with animals consuming waxy sorghum gaining significantly more weight. Conclusions:We conclude that the benefits of waxy starches on food functionality can have trade-off effects on the gut microbiome and host physiology that could be particularly relevant in human populations consuming large amounts of waxy grains.

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Several bioactive components of the human diet have major effects on composition and function of the gut microbiome, but no systematic framework exists for understanding variation in microbiome-active components amid the vast amount of genotypic and phenotypic variation within a given species of food crop. Here we present a powerful new approach for complex trait analysis of Microbiome-Active Traits (MATs) in food crops. Capitalizing on a novel automated in vitro microbiome screening (AiMS) methodology to quantify human gut microbiome phenotypes after fermentation of grain from genetically diverse lines, we show how microbiome phenotypes can be used as quantitative traits for genetic analysis. Quantitative Trait Locus (QTL) analysis of AiMS-based phenotypes across grain samples from 294 sorghum (Sorghum bicolor) recombinant inbred lines identified significant QTLs at 10 different genomic regions that collectively control MATs affecting 16 different microbial taxa. Segregation analysis and validation in Near-Isogenic Lines (NILs) confirmed that overlapping QTL peaks for microbiome phenotypes, seed color, and tannin concentration are driven by variation in the Tan2 (chromosome 2) and Tan1 (chromosome 4) regulators of the tannin biosynthetic pathway. Candidate genes at other QTLs suggest that variation in a diverse array of plant molecules can drive MATs.