Understanding the role of amino acid sources in the small intestinal microbial dominance


The mammalian gut harbors heterogeneous distribution of microbes along the gastrointestinal tract. Although it is recognized that the environmental differences along the gastrointestinal tract are likely responsible for this, the underlying mechanisms behind the dominance of specific microbial taxa in the small intestine is not well studied. As demonstrated in our earlier study, piglets have a high abundance of lactic acid bacteria in the small intestine (mainly Lactobacillus and Streptococcus), while microbes suggested to play a role in complex carbohydrate degradation such as Prevotella show high proportions in the large intestine (Mu et al. 2017). This represents a typical signature of microbial composition in the mammalian gut. However, the rationales responsible for the microbial distribution are rarely understood, mostly due to the complex ecosystem in the gut and a diverse array of host-microbe interaction which represents a great challenge to the microbiome research over the past 20 years. Zoetendal and colleagues have found that fast uptake and conversion of simple carbohydrates by fast growing microbes (mostly Streptococcus) are driving the small intestinal microbial ecosystem in humans (Zoetendal et al. 2012), implicating the relationship between substrate availability and microbial composition.

Our group has spent tremendous efforts since 2008 in understanding how the gut microbes in the small intestine utilize amino acids in believing that dietary nutrients are not only used for host, but also for the tiny gut bugs. In other words, amino acid utilization may underlie a possible dialogue of host-microbe symbiosis.

Initially, we found species-level heterogeneity in amino acid utilization of gut microbes in the digesta of the small intestine, e.g. Streptococcus sp. and Megasphaera elsdenii-like bacteria are predominant in utilizing lysine, threonine, arginine and glutamate (Dai et al. 2010). Afterwards, we uncovered  compartment-level heterogeneity in amino acid utilization, namely, the microbes in gut lumen and epithelial walls also differ in amino acid utilization (Yang et al. 2014). These findings convinced us that we are on the right way of dissecting the hetrogeneity of microbial abundance in the gut. Recently, we discovered that the microbes from the small intestine of pigs grow better in medium containing peptides as only nitrogen source than free amino acids (Liu et al. 2018). The significance of this finding is somehow underestimated until we further found that diet with peptide-bound amino acids selectively enriched Lactobacillus species in the small intestine, as presented in the ISME Journal (Jing et al. 2022).

Protein hydrolysate is a dietary ingredient containing peptide-bound amino acids with bioactive function.  We demonstrated that compared to intact casein, dietary casein hydrolysate stimulated gastric transit and affected microbial composition in the small intestine of pigs (Shen et al. 2020). In the following study, the enrichment of Lactobacillus, notably Lactobacillus amylovorus in the small intestine raises our interest in determining the how and why these taxa became dominant after dietary casein hydrolysate. With the increase of Lactobacillus amylovorus, peptide-rich substrates also became increased in the small intestine. We then question if the increased peptide substrate may give arise to specific taxa. The simple answer is YES.

Through an in depth genomic analysis of  L. amylovorus S1, which was significantly increased in abundance after dietary casein hydrolysate in vivo, we identified a diverse collection of genes encoding transporters and metabolic enzymes that are involved in the amino acid metabolism by L. amylovorus S1. The functional potential was further validated by analyzing the gene expression of amino acid transporters and amino acid disappearance rate during batch cultivation in media containing different forms of nitrogen susbtrate.  L. amylovorus S1 grew better with peptide-rich substrates than amino acid-rich substrates as nitrogen source, as reflected by its enhanced growth and upregulation of genes encoding threonine-, phyenylalanine-, valine-, and lysine-containing peptide utilization, as well as  peptide transporters such as oligopeptide ABC transporter-binding protein. In addition to peptide utilization, the utilization of free amino acids (e.g. methionine, valine, lysine) were enhanced simultaneously with peptide-rich substrates.

Additionally, lactate was elevated when L. amylovorus S1 was cultivated with peptide-rich substrates while acetate was elevated when it was cultivated with amino acid-rich substrates, indicating distinct metabolic patterns depending on substrate forms. Overall, these findings suggest that an increased capability of utilizing peptide-bound amino acids contributes to the dominance of L. amylovorus, with amino acid utilization as a deterministic factor affecting intestinal microbial distribution.

The microbial dominance along the gut can determine their overall performance and functionality in gut nutrition and health. Herein, the approach we reported provides a framework to explain the dominance and subsequently the performance of key taxa in the small intestine, which could be further applied to other microbes and investigated in future. Given that a peptide-rich intestinal microenvironment provides competitive advantage for Lactobacillus and notably L. amylovorus, our findings implicate the significance of nutritional manipulation by peptide-rich diet to regulate intestinal microenvironment. These findings also help understand the microbe-gut nutrition interplay and provide guidelines for dietary manipulations towards gut health especially small intestine health.



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Jing Y, Mu C, Wang HS, Shen JH, Zoetendal EG, Zhu W. Amino acid utilization allows intestinal dominance of Lactobacillus amylovorus. ISME J. 2022; in press

Liu J, Mu C, Yu K, Zhu W. Effect of two different casein hydrolysates on small intestinal bacteria of growing pigs. Acta Microbiologica Sinica. 2018;58:63-72.

Mu C, Yang Y, Su Y, Zoetendal EG, Zhu W. Differences in microbiota membership along the gastrointestinal tract of piglets and their differential alterations following an early-life antibiotic intervention. Front Microbiol. 2017;8:797.

Shen J, Mu C, Wang H, Huang Z, Yu K, Zoetendal EG, et al. Stimulation of gastric transit function driven by hydrolyzed casein increases small intestinal carbohydrate availability and its microbial metabolism. Mol Nutr Food Res. 2020;64:2000250.

Zoetendal E G, Raes J, Van Den Bogert B, et al. The human small intestinal microbiota is driven by rapid uptake and conversion of simple carbohydrates. ISME J. 2012;6(7): 1415-26.