Pangenome-based design of strain-specific primers enables precise monitoring of bacteria in human microbiome intervention trials

pangenome-based-design-of-strain-specific-primers-enables-precise-monitoring-of-bacteria-in-human-microbiome-intervention-trials
Pangenome-based design of strain-specific primers enables precise monitoring of bacteria in human microbiome intervention trials

References

  1. Larsen, I. S. et al. Experimental diets dictate the metabolic benefits of probiotics in obesity. Gut Microbes 15, (2023).

  2. Nicola, T. et al. A lactobacilli-based inhaled live biotherapeutic product attenuates pulmonary neutrophilic inflammation. Nat. Commun. 15, (2024).

  3. Leroy, F. & De Vuyst, L. Lactic acid bacteria as functional starter cultures for the food fermentation industry. Trends Food Sci. Technol. 15, 67–78 (2004).

    Google Scholar 

  4. Legein, M. et al. Modes of Action of Microbial Biocontrol in the Phyllosphere. Front. Microbiol. 11, 544057 (2020).

    Google Scholar 

  5. Backer, R. et al. Plant Growth-Promoting Rhizobacteria: Context, Mechanisms of Action, and Roadmap to Commercialization of Biostimulants for Sustainable Agriculture. Front. Plant. Sci. 9, 402666 (2018).

    Google Scholar 

  6. Lebeer, S. et al. Functional analysis of lactobacillus rhamnosus GG pili in relation to adhesion and immunomodulatory interactions with intestinal epithelial cells. Appl. Environ. Microbiol. 78, 185–193 (2012).

    Google Scholar 

  7. de Vries, M. C., Vaughan, E. E., Kleerebezem, M. & de Vos, W. M. Lactobacillus plantarum—survival, functional and potential probiotic properties in the human intestinal tract. Int. Dairy. J. 16, 1018–1028 (2006).

    Google Scholar 

  8. FDA. Early Clinical Trials with Live Biotherapeutic Products: Chemistry, Manufacturing, and Control Information; Guidance for Industry. (2016).

  9. Vargason, A. M. & Anselmo, A. C. Live Biotherapeutic Products and Probiotics for the Skin. Adv. NanoBiomed Res. 1, 1–6 (2021).

    Google Scholar 

  10. Hemmerling, A. et al. Effect of the vaginal live biotherapeutic LACTIN-V (Lactobacillus crispatus CTV-05) on vaginal microbiota and genital tract inflammation among women at high risk of HIV acquisition in South Africa: a phase 2, randomised, placebo-controlled trial. Lancet Microbe. 6, 1–12 (2025).

    Google Scholar 

  11. Smillie, C. S. et al. Strain Tracking Reveals the Determinants of Bacterial Engraftment in the Human Gut Following Fecal Microbiota Transplantation. Cell. Host Microbe. 23, 229–240e5 (2018).

    Google Scholar 

  12. Kumar, R. et al. Identification of donor microbe species that colonize and persist long term in the recipient after fecal transplant for recurrent Clostridium difficile. NPJ Biofilms Microbiomes 3, (2017).

  13. Lebeer, S. et al. Selective targeting of skin pathobionts and inflammation with topically applied lactobacilli. Cell. Rep. Med. 3, 100521 (2022).

    Google Scholar 

  14. Cohen, C. R. et al. Randomized Trial of Lactin-V to Prevent Recurrence of Bacterial Vaginosis. N Engl. J. Med. 382, 1906–1915 (2020).

    Google Scholar 

  15. Van Holm, W. et al. Antimicrobial potential of known and novel probiotics on in vitro periodontitis biofilms. npj Biofilms Microbiomes. 9, 1–12 (2023).

    Google Scholar 

  16. De Boeck, I. et al. Lacticaseibacillus rhamnosus GG in a chewable colonizes the nose and facilitates local immune benefits in allergic rhinoconjunctivitis patients. Microbiol Spectr 0, (2025).

  17. Wendel, U. Assessing Viability and Stress Tolerance of Probiotics—A Review. Front. Microbiol. 12, 818468 (2022).

    Google Scholar 

  18. Capozzi, V., Fragasso, M. & Russo, P. Microbiological Safety and the management of microbial resources in artisanal foods and beverages: The need for a transdisciplinary assessment to conciliate actual trends and risks avoidance. Microorganisms 8, (2020).

  19. Ashaolu, T. J. Safety and quality of bacterially fermented functional foods and beverages: a mini review. Food Qual. Saf. 4, 123–127 (2020).

    Google Scholar 

  20. Bron, P. A., Grangette, C., Mercenier, A., De Vos, W. M. & Kleerebezem, M. Identification of Lactobacillus plantarum Genes That Are Induced in the Gastrointestinal Tract of Mice. J. Bacteriol. 186, 5721 (2004).

    Google Scholar 

  21. Spacova, I. et al. Expression of fluorescent proteins in Lactobacillus rhamnosus to study host–microbe and microbe–microbe interactions. Microb. Biotechnol. 11, 317–331 (2018).

    Google Scholar 

  22. Turpin, P. E., Maycroft, K. A., Bedford, J., Rowlands, C. L. & Wellington, E. M. H. A rapid luminescent-phage based MPN method for the enumeration of Salmonella typhimurium in environmental samples. Lett. Appl. Microbiol. 16, 24–27 (1993).

    Google Scholar 

  23. Fasanello, V. J., Liu, P., Botero, C. A. & Fay, J. C. High-throughput analysis of adaptation using barcoded strains of Saccharomyces cerevisiae. PeerJ 8, e10118 (2020).

    Google Scholar 

  24. Fleischmann, R. D. et al. Whole-Genome Random Sequencing and Assembly of Haemophilus influenzae Rd. Sci. (80-). 269, 496–512 (1995).

    Google Scholar 

  25. Parks, D. H. et al. An ongoing census of bacterial and archaeal diversity through a phylogenetically consistent, rank normalized and complete genome-based taxonomy. Nucleic Acids Res. 50, D785–D794 (2022).

    Google Scholar 

  26. Anderson, B. D. & Bisanz, J. E. Challenges and opportunities of strain diversity in gut microbiome research. Front. Microbiol. 14, 1–8 (2023).

    Google Scholar 

  27. Dijkshoorn, L., Ursing, B. M. & Ursing, J. B. Strain, clone and species: Comments on three basic concepts of bacteriology. J. Med. Microbiol. 49, 397–401 (2000).

    Google Scholar 

  28. Hill, C. & International Scientific Association for Probiotics and Prebiotics (ISAPP). What is a strain in microbiology and why does it matter? ISAPP Sci. Blog (2022).

  29. Van Rossum, T., Ferretti, P., Maistrenko, O. M. & Bork, P. Diversity within species: interpreting strains in microbiomes. Nat. Rev. Microbiol. 18, 491–506 (2020).

    Google Scholar 

  30. Pasolli, E. et al. Large-scale genome-wide analysis links lactic acid bacteria from food with the gut microbiome. Nat. Commun. 11, 1–12 (2020).

    Google Scholar 

  31. De Boeck, I. et al. Randomized, double-blind, placebo-controlled trial of a throat spray with selected Lactobacilli in COVID-19 outpatients. Microbiol. Spectr. 10, (2022).

  32. De Boeck, I. et al. Lactobacilli Have a Niche in the Human Nose. Cell. Rep. 31, 107674 (2020).

    Google Scholar 

  33. Lebeer, S. et al. A citizen-science-enabled catalogue of the vaginal microbiome and associated factors. Nat. Microbiol. 8, 2183–2195 (2022).

    Google Scholar 

  34. Wuyts, S. et al. Carrot juice fermentations as man-made microbial ecosystems dominated by lactic acid bacteria. Appl. Environ. Microbiol. 84, e00134–e00118 (2018).

    Google Scholar 

  35. Alimolaei, M. & Golchin, M. A comparison of methods for extracting plasmids from a difficult to lyse bacterium: Lactobacillus casei. Biologicals 45, 47–51 (2017).

    Google Scholar 

  36. Wuyts, S. et al. Comparative genome analysis of lactobacillus mudanjiangensis, an understudied member of the lactobacillus plantarum group. Microb. Genomics 5, (2019).

  37. Parks, D. H., Imelfort, M., Skennerton, C. T., Hugenholtz, P. & Tyson, G. W. CheckM: Assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes. Genome Res. 25, 1043–1055 (2015).

    Google Scholar 

  38. Orakov, A. et al. GUNC: detection of chimerism and contamination in prokaryotic genomes. Genome Biol. 22, 1–19 (2021).

    Google Scholar 

  39. Kankainen, M. et al. Comparative genomic analysis of Lactobacillus rhamnosus GG reveals pili containing a human- mucus binding protein. Proc. Natl. Acad. Sci. 106, 17193–17198 (2009).

    Google Scholar 

  40. Petrova, M. I. et al. Comparative genomic and phenotypic analysis of the vaginal probiotic lactobacillus rhamnosus GR-1. Front. Microbiol 9, (2018).

  41. Kleerebezem, M. et al. Complete genome sequence of Lactobacillus plantarum WCFS1. Proc. Natl. Acad. Sci. 100, 1990–1995 (2003).

    Google Scholar 

  42. Malik, S. et al. High mannose-specific lectin Msl mediates key interactions of the vaginal Lactobacillus plantarum isolate CMPG5300. Sci. Rep. 2016 61 (6), 1–16 (2016).

    Google Scholar 

  43. Danielsen, M. Characterization of the tetracycline resistance plasmid pMD5057 from Lactobacillus plantarum 5057 reveals a composite structure. Plasmid 48, 98–103 (2002).

    Google Scholar 

  44. Leer, R. J., van Luijk, N., Posno, M. & Pouwels, P. H. Structural and functional analysis of two cryptic plasmids from Lactobacillus pentosus MD353 and Lactobacillus plantarum ATCC 8014. MGG Mol. Gen. Genet. 234, 265–274 (1992).

    Google Scholar 

  45. Allonsius, C. N. et al. Inhibition of Candida albicans morphogenesis by chitinase from Lactobacillus rhamnosus GG. Sci. Rep. 9, 2900 (2019).

    Google Scholar 

  46. Reid, G., Cook, R. L. & Bruce, A. W. Examination of strains of lactobacilli for properties that may influence bacterial interference in the urinary tract. J. Urol. 138, 330–335 (1987).

    Google Scholar 

  47. Sriramulu, D. D. et al. Lactobacillus reuteri DSM 20016 produces cobalamin-dependent diol dehydratase in metabolosomes and metabolizes 1,2-propanediol by disproportionation. J. Bacteriol. 190, 4559–4567 (2008).

    Google Scholar 

  48. Jörissen, J. et al. Case-Control Microbiome Study of Chronic Otitis Media with Effusion in Children Points at Streptococcus salivarius as a Pathobiont-Inhibiting Species. mSystems 6, (2021).

  49. Mignolet, J., Fontaine, L., Kleerebezem, M. & Hols, P. Complete genome sequence of Streptococcus salivarius HSISS4, a human commensal bacterium highly prevalent in the digestive tract. Genome Announc 4, (2016).

  50. Butler, R. R., Soomer-James, J. T. A., Frenette, M. & Pombert, J. F. Complete genome sequences of two human oral microbiome commensals, Streptococcus salivarius ATCC 25975 and S. salivarius ATCC 27945. Genome Announc 5, (2017).

  51. Parks, D. H. et al. GTDB: an ongoing census of bacterial and archaeal diversity through a phylogenetically consistent, rank normalized and complete genome-based taxonomy. Nucleic Acids Res. 50, D785–D794 (2022).

    Google Scholar 

  52. Hyatt, D. et al. Prodigal: Prokaryotic gene recognition and translation initiation site identification. BMC Bioinform. 11, 1–11 (2010).

    Google Scholar 

  53. Wittouck, S., Eilers, T., van Noort, V. & Lebeer, S. SCARAP: scalable cross-species comparative genomics of prokaryotes. Bioinformatics btae735 https://doi.org/10.1093/bioinformatics/btae735 (2024).

  54. Jain, C., Rodriguez-R, L. M., Phillippy, A. M., Konstantinidis, K. T. & Aluru, S. High throughput ANI analysis of 90K prokaryotic genomes reveals clear species boundaries. Nat. Commun. 2018. 91 9, 1–8 (2018).

    Google Scholar 

  55. Zhang, Z., Schwartz, S., Wagner, L. & Miller, W. A greedy algorithm for aligning DNA sequences. J. Comput. Biol. 7, 203–214 (2000).

    Google Scholar 

  56. Bustin, S. A. et al. The MIQE guidelines: Minimum information for publication of quantitative real-time PCR experiments. Clin. Chem. 55, 611–622 (2009).

    Google Scholar 

  57. Lebeer, S. et al. Selective targeting of skin pathobionts and inflammation with topically applied lactobacilli. Cell Rep. Med 3, (2022).

  58. Wuyts, S. et al. Large-Scale Phylogenomics of the Lactobacillus casei Group Highlights Taxonomic Inconsistencies and Reveals Novel Clade-Associated Features. mSystems 2, (2017).

  59. Vander Donck, L. et al. Host-independent synergism between Lactobacillus crispatus and other vaginal lactobacilli. Cell Rep 44, (2025).

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