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Amino acid utilization allows intestinal dominance of Lactobacillus amylovorus

Abstract

The mammalian intestine harbors heterogeneous distribution of microbes among which specific taxa (e.g. Lactobacillus) dominate across mammals. Deterministic factors such as nutrient availability and utilization may affect microbial distributions. Due to physiological complexity, mechanisms linking nutrient utilization and the dominance of key taxa remain unclear. Lactobacillus amylovorus is a predominant species in the small intestine of pigs. Employing a pig model, we found that the small intestine was dominated by Lactobacillus and particularly L. amylovorus, and enriched with peptide-bound amino acids (PBAAs), all of which were further boosted after a peptide-rich diet. To investigate the bacterial growth dominance mechanism, a representative strain L. amylovorus S1 was isolated from the small intestine and anaerobically cultured in media with free amino acids or peptides as sole nitrogen sources. L. amylovorus S1 grew preferentially with peptide-rich rather than amino acid-rich substrates, as reflected by enhanced growth and PBAA utilization, and peptide transporter upregulations. Utilization of free amino acids (e.g. methionine, valine, lysine) and expressions of transporters and metabolic enzymes were enhanced simultaneously in peptide-rich substrate. Additionally, lactate was elevated in peptide-rich substrates while acetate in amino acid-rich substrates, indicating distinct metabolic patterns depending on substrate forms. These results suggest that an increased capability of utilizing PBAAs contributes to the dominance of L. amylovorus, indicating amino acid utilization as a deterministic factor affecting intestinal microbial distribution. These findings may provide new insights into the microbe-gut nutrition interplay and guidelines for dietary manipulations toward gut health especially small intestine health.

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Fig. 1: Structure and composition of gut microbiota in pigs fed with intact casein or hydrolyzed casein.
Fig. 2: Measurement of bacterial quantities and peptide-bound amino acids.
Fig. 3: Identification and genomic sequencing of L. amylovorus S1 from the small intestine of pigs.
Fig. 4: Growth traits and utilization of peptide-bound amino acids by L. amylovorus S1 in amino acid-rich or peptide-rich substrates in vitro.
Fig. 5: The utilization, transport and metabolism of free AAs by L. amylovorus S1 in vitro.
Fig. 6: Distinct metabolic patterns in amino acid-rich or peptide-rich substrates during in vitro cultivation.
Fig. 7: Summary of the amino acid utilization in L. amylovorus.

Data availability

The 16S rRNA gene sequence of L. amylovorus S1 has been submitted to NCBI under the accession number MT525371. The high-throughput sequencing data are available under PRJNA796201 within NCBI Sequence Read Archive. The complete sequence of L. amylovorus S1 is available at GenBank under the accession number: CP090603 (chromosome), CP090604 (plasmid 1) and CP090605 (plasmid 2).

References

  1. Chen YJ, Leung PM, Wood JL, Bay SK, Hugenholtz P, Kessler AJ, et al. Metabolic flexibility allows bacterial habitat generalists to become dominant in a frequently disturbed ecosystem. ISME J. 2021;15:2986–3004.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  2. 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.

    PubMed  PubMed Central  Article  Google Scholar 

  3. Li D, Chen H, Mao B, Yang Q, Zhao J, Gu Z, et al. Microbial biogeography and core microbiota of the rat digestive tract. Sci Rep. 2017;8:45840.

    CAS  PubMed  Article  Google Scholar 

  4. Yasuda K, Oh K, Ren B, Tickle TL, Franzosa EA, Wachtman LM, et al. Biogeography of the intestinal mucosal and lumenal microbiome in the rhesus macaque. Cell Host Microbe. 2015;17:385–91.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  5. Hynonen U, Kant R, Lahteinen T, Pietila TE, Beganovic J, Smidt H, et al. Functional characterization of probiotic surface layer protein-carrying Lactobacillus amylovorus strains. BMC Microbiol. 2014;14:199.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  6. Marti R, Dabert P, Ziebal C, Pourcher A-M. Evaluation of Lactobacillus sobrius/L. amylovorus as a new microbial marker of pig manure. Appl Environ Microbiol. 2010;76:1456–61.

    CAS  PubMed  Article  Google Scholar 

  7. Konstantinov SR, Awati AA, Williams BA, Miller BG, Jones P, Stokes CR, et al. Post-natal development of the porcine microbiota composition and activities. Environ Microbiol. 2006;8:1191–9.

    CAS  PubMed  Article  Google Scholar 

  8. Dai Z, Zhang J, Wu G, Zhu W. Utilization of amino acids by bacteria from the pig small intestine. Amino Acids. 2010;39:1201–15.

    CAS  PubMed  Article  Google Scholar 

  9. Wallace RJ. Ruminal microbial metabolism of peptides and amino acids. J Nutr. 1996;126:1326S–34S.

    CAS  PubMed  Article  Google Scholar 

  10. Yang YX, Dai ZL, Zhu WY. Important impacts of intestinal bacteria on utilization of dietary amino acids in pigs. Amino Acids. 2014;46:2489–501.

    CAS  PubMed  Article  Google Scholar 

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

    CAS  Google Scholar 

  12. 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.

    CAS  Article  Google Scholar 

  13. Hsieh CM, Yang F-C, Iannotti EL. The effect of soy protein hydrolyzates on fermentation by Lactobacillus amylovorus. Process Biochem. 1999;34:173–9.

    CAS  Article  Google Scholar 

  14. Visser J, Bos N, Harthoorn L, Stellaard F, Beijer‐Liefers S, Rozing J, et al. Potential mechanisms explaining why hydrolyzed casein‐based diets outclass single amino acid‐based diets in the prevention of autoimmune diabetes in diabetes‐prone BB rats. Diabetes/Metab Res Rev. 2012;28:505–13.

    CAS  Article  Google Scholar 

  15. Singh KM, Jisha TK, Reddy B, Parmar N, Patel A, Patel AK, et al. Microbial profiles of liquid and solid fraction associated biomaterial in buffalo rumen fed green and dry roughage diets by tagged 16S rRNA gene pyrosequencing. Mol Biol Rep. 2015;42:95–103.

    CAS  PubMed  Article  Google Scholar 

  16. Fadrosh DW, Ma B, Gajer P, Sengamalay N, Ott S, Brotman RM, et al. An improved dual-indexing approach for multiplexed 16S rRNA gene sequencing on the Illumina MiSeq platform. Microbiome. 2014;2:6.

    PubMed  PubMed Central  Article  Google Scholar 

  17. Segata N, Izard J, Waldron L, Gevers D, Miropolsky L, Garrett WS, et al. Metagenomic biomarker discovery and explanation. Genome Biol. 2011;12:R60.

    PubMed  PubMed Central  Article  Google Scholar 

  18. Schloss PD, Westcott SL, Ryabin T, Hall JR, Hartmann M, Hollister EB, et al. Introducing mothur: open-source, platform-independent, community-supported software for describing and comparing microbial communities. Appl Environ Microbiol. 2009;75:7537–41.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  19. Kim E, Yang S-M, Lim B, Park SH, Rackerby B, Kim H-Y. Design of PCR assays to specifically detect and identify 37 Lactobacillus species in a single 96 well plate. BMC Microbiol. 2020;20:96.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  20. Chen L, Luo Y, Wang H, Liu S, Shen Y, Wang M. Effects of glucose and starch on lactate production by newly isolated Streptococcus bovis S1 from Saanen Goats. Appl Environ Microbiol. 2016;82:5982–9.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  21. Yang J, Roy A, Zhang Y. Protein–ligand binding site recognition using complementary binding-specific substructure comparison and sequence profile alignment. Bioinformatics. 2013;29:2588–95.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  22. Zheng W, Zhang C, Li Y, Pearce R, Bell EW, Zhang Y. Folding non-homologous proteins by coupling deep-learning contact maps with I-TASSER assembly simulations. Cell Rep Methods 2021;1:100014.

    PubMed  PubMed Central  Article  Google Scholar 

  23. Yang Y, Faraggi E, Zhao H, Zhou Y. Improving protein fold recognition and template-based modeling by employing probabilistic-based matching between predicted one-dimensional structural properties of query and corresponding native properties of templates. Bioinformatics. 2011;27:2076–82.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  24. Yang J, Roy A, Zhang Y. BioLiP: a semi-manually curated database for biologically relevant ligand-protein interactions. Nucleic Acids Res. 2013;41:D1096–103.

    CAS  PubMed  Article  Google Scholar 

  25. Capra JA, Laskowski RA, Thornton JM, Singh M, Funkhouser TA. Predicting protein ligand binding sites by combining evolutionary sequence conservation and 3D structure. PLoS Comput Biol. 2009;5:e1000585.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  26. Dai Z, Wu Z, Jia S, Wu G. Analysis of amino acid composition in proteins of animal tissues and foods as pre-column o-phthaldialdehyde derivatives by HPLC with fluorescence detection. J Chromatogr B Anal Technol Biomed Life Sci. 2014;964:116–27.

    CAS  Article  Google Scholar 

  27. Zhang C, Yu M, Yang Y, Mu C, Su Y, Zhu W. Differential effect of early antibiotic intervention on bacterial fermentation patterns and mucosal gene expression in the colon of pigs under diets with different protein levels. Appl Microbiol Biotechnol. 2017;101:2493–505.

    CAS  PubMed  Article  Google Scholar 

  28. Benjamini Y, Hochberg Y. Controlling the false discovery rate: a practical and powerful approach to multiple testing. J R Stat Soc. 1995;57:289–300.

    Google Scholar 

  29. Lamarque M, Charbonnel P, Aubel D, Piard J-C, Atlan D, Juillard V. A multifunction ABC transporter (Opt) contributes to diversity of peptide uptake specificity within the genus Lactococcus. J Bacteriol. 2004;186:6492–500.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  30. Dai Z, Wu G, Zhu W. Amino acid metabolism in intestinal bacteria: links between gut ecology and host health. Front Biosci. 2011;16:1768–86.

    CAS  Article  Google Scholar 

  31. Morowitz MJ, Carlisle EM, Alverdy JC. Contributions of intestinal bacteria to nutrition and metabolism in the critically ill. Surgical Clin. 2011;91:771–85.

    Google Scholar 

  32. Zhang Q, Ren J, Zhao H, Zhao M, Xu J, Zhao Q. Influence of casein hydrolysates on the growth and lactic acid production of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus. Int J Food Sci Technol. 2011;46:1014–20.

    CAS  Article  Google Scholar 

  33. Davila A-M, Blachier F, Gotteland M, Andriamihaja M, Benetti P-H, Sanz Y, et al. Re-print of “Intestinal luminal nitrogen metabolism: Role of the gut microbiota and consequences for the host”. Pharmacol Res. 2013;69:114–26.

    PubMed  Article  Google Scholar 

  34. Pridmore RD, Berger B, Desiere F, Vilanova D, Barretto C, Pittet AC, et al. The genome sequence of the probiotic intestinal bacterium Lactobacillus johnsonii NCC 533. Proc Natl Acad Sci USA 2004;101:2512–7.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  35. Michalak L, Gaby JC, Lagos L, La Rosa SL, Hvidsten TR, Tétard-Jones C, et al. Microbiota-directed fibre activates both targeted and secondary metabolic shifts in the distal gut. Nat Commun. 2020;11:5773.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  36. Pereira FC, Berry D. Microbial nutrient niches in the gut. Environ Microbiol. 2017;19:1366–78.

    PubMed  PubMed Central  Article  Google Scholar 

  37. Zhang L, Wu W, Lee YK, Xie J, Zhang H. Spatial heterogeneity and co-occurrence of mucosal and luminal microbiome across swine intestinal tract. Front Microbiol. 2018;9:48.

    PubMed  PubMed Central  Article  Google Scholar 

  38. Chang PV, Hao L, Offermanns S, Medzhitov R. The microbial metabolite butyrate regulates intestinal macrophage function via histone deacetylase inhibition. Proc Natl Acad Sci USA. 2014;111:2247–52.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  39. Argyle J, Baldwin R. Effects of amino acids and peptides on rumen microbial growth yields. J Dairy Sci. 1989;72:2017–27.

    CAS  PubMed  Article  Google Scholar 

  40. Chen G, Strobel H, Russell J, Sniffen C. Effect of hydrophobicity of utilization of peptides by ruminal bacteria in vitro. Appl Environ Microbiol. 1987;53:2021–5.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  41. Soto RC, Muhammed SA, Newbold C, Stewart C, Wallace R. Influence of peptides, amino acids and urea on microbial activity in the rumen of sheep receiving grass hay and on the growth of rumen bacteria in vitro. Anim Feed Sci Technol. 1994;49:151–61.

    CAS  Article  Google Scholar 

  42. Chalova VI, Sirsat SA, O’Bryan CA, Crandall PG, Ricke SC. Escherichia coli, an intestinal microorganism, as a biosensor for quantification of amino acid bioavailability. Sensors. 2009;9:7038–57.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  43. Matthews D, Payne J. Peptides in the nutrition of microorganisms and peptides in relation to animal nutrition. In: Peptide transport in protein nutrition, New York, US: North-Holland Publishing Company; 1975. p. 1–60. vol. 37.

  44. Sanz Y, Lanfermeijer FC, Renault P, Bolotin A, Konings WN, Poolman B. Genetic and functional characterization of dpp genes encoding a dipeptide transport system in Lactococcus lactis. Arch Microbiol. 2001;175:334–43.

    CAS  PubMed  Article  Google Scholar 

  45. Hartmann T, Cairns TC, Olbermann P, Morschhäuser J, Bignell EM, Krappmann S. Oligopeptide transport and regulation of extracellular proteolysis are required for growth of Aspergillus fumigatus on complex substrates but not for virulence. Mol Microbiol. 2011;82:917–35.

    CAS  PubMed  Article  Google Scholar 

  46. Dressaire C, Redon E, Gitton C, Loubière P, Monnet V, Cocaign-Bousquet M. Investigation of the adaptation of Lactococcus lactis to isoleucine starvation integrating dynamic transcriptome and proteome information. Microb Cell Factories. 2011;10:S18.

  47. Wissenbach U, Six S, Bongaerts J, Ternes D, Steinwachs S, Unden G. A third periplasmic transport system for l‐arginine in Escherichia coli: molecular characterization of the artPIQMJ genes, arginine binding and transport. Mol Microbiol. 1995;17:675–86.

    CAS  PubMed  Article  Google Scholar 

  48. Merlin C, Gardiner G, Durand S, Masters M. The Escherichia coli metD locus encodes an ABC transporter which includes Abc (MetN), YaeE (MetI), and YaeC (MetQ). J Bacteriol. 2002;184:5513–7.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  49. Rodionov DA, Vitreschak AG, Mironov AA, Gelfand MS. Regulation of lysine biosynthesis and transport genes in bacteria: yet another RNA riboswitch? Nucleic acids Res. 2003;31:6748–57.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  50. Nguyen PT, Lai JY, Lee AT, Kaiser JT, Rees DC. Noncanonical role for the binding protein in substrate uptake by the MetNI methionine ATP Binding Cassette (ABC) transporter. Proc Natl Acad Sci USA. 2018;115:E10596–604.

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Flatley J, Barrett J, Pullan ST, Hughes MN, Green J, Poole RK. Transcriptional responses of Escherichia coli to S-nitrosoglutathione under defined chemostat conditions reveal major changes in methionine biosynthesis. J Biol Chem. 2005;280:10065–72.

    CAS  PubMed  Article  Google Scholar 

  52. Caldara M, Charlier D, Cunin R. The arginine regulon of Escherichia coli: whole-system transcriptome analysis discovers new genes and provides an integrated view of arginine regulation. Microbiology. 2006;152:3343–54.

    CAS  PubMed  Article  Google Scholar 

  53. Yvon M, Chambellon E, Bolotin A, Roudot-Algaron F. Characterization and role of the branched-chain aminotransferase (BcaT) isolated from Lactococcus lactis subsp. cremoris NCDO 763. Appl Environ Microbiol. 2000;66:571–7.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  54. Li W, Zhang Y, Li H, Zhang C, Zhang J, Uddin J, et al. Effect of soybean oligopeptide on the growth and metabolism of Lactobacillus acidophilus JCM 1132. RSC Adv. 2020;10:16737–48.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  55. Alberdi A, Andersen SB, Limborg MT, Dunn RR, Gilbert MTP. Disentangling host–microbiota complexity through hologenomics. Nat Rev Genet. 2022;23:281–97.

  56. Bordenstein SR, Theis KR. Host biology in light of the microbiome: ten principles of holobionts and hologenomes. PLoS Biol. 2015;13:e1002226.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

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Acknowledgements

This work was supported by National Natural Science Foundation of China (32030104, 31902166).

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WZ conceived, designed, supervised the study and proposed the manuscript strategy. YJ conducted the experimental work and performed statistical tests with guidance from CM and EGZ. CM wrote the manuscript draft with revisions and edits by YJ, EGZ and WZ. WZ and CM finalized the manuscript. HW and JS conducted the animal study. All the authors have read and approved the final manuscript for publication.

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Correspondence to Weiyun Zhu.

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Jing, Y., Mu, C., Wang, H. et al. Amino acid utilization allows intestinal dominance of Lactobacillus amylovorus. ISME J (2022). https://doi.org/10.1038/s41396-022-01287-8

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