title: D-Alanine-Controlled Transient Intestinal Mono-Colonization with Non-Laboratory-Adapted Commensal E. coli Strain HS.
abstract: Soon after birth the mammalian gut microbiota forms a permanent and collectively highly resilient consortium. There is currently no robust method for re-deriving an already microbially colonized individual again-germ-free. We previously developed the in vivo growth-incompetent E. coli K-12 strain HA107 that is auxotrophic for the peptidoglycan components D-alanine (D-Ala) and meso-diaminopimelic acid (Dap) and can be used to transiently associate germ-free animals with live bacteria, without permanent loss of germ-free status. Here we describe the translation of this experimental model from the laboratory-adapted E. coli K-12 prototype to the better gut-adapted commensal strain E. coli HS. In this genetic background it was necessary to complete the D-Ala auxotrophy phenotype by additional knockout of the hypothetical third alanine racemase metC. Cells of the resulting fully auxotrophic strain assembled a peptidoglycan cell wall of normal composition, as long as provided with D-Ala and Dap in the medium, but could not proliferate a single time after D-Ala/Dap removal. Yet, unsupplemented bacteria remained active and were able to complete their cell cycle with fully sustained motility until immediately before autolytic death. Also in vivo, the transiently colonizing bacteria retained their ability to stimulate a live-bacteria-specific intestinal Immunoglobulin (Ig)A response. Full D-Ala auxotrophy enabled rapid recovery to again-germ-free status. E. coli HS has emerged from human studies and genomic analyses as a paradigm of benign intestinal commensal E. coli strains. Its reversibly colonizing derivative may provide a versatile research tool for mucosal bacterial conditioning or compound delivery without permanent colonization.
authors.affiliation: Institute for Infectious Diseases, University of Bern, Bern, Switzerland.|Graduate School GCB, University of Bern, Bern, Switzerland.
authors.forename: Miguelangel
authors.initials: M
authors.lastname: Cuenca
authors.affiliation: Institute for Infectious Diseases, University of Bern, Bern, Switzerland.|Graduate School GCB, University of Bern, Bern, Switzerland.
authors.forename: Simona P
authors.initials: SP
authors.lastname: Pfister
authors.affiliation: Institute for Infectious Diseases, University of Bern, Bern, Switzerland.|Graduate School GCB, University of Bern, Bern, Switzerland.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Buschor
authors.affiliation: Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
authors.forename: Firuza
authors.initials: F
authors.lastname: Bayramova
authors.affiliation: Laboratory for Molecular Infection Medicine Sweden, Department of Molecular Biology, Umeå Centre for Microbial Research, Umeå University, Umeå, Sweden.
authors.forename: Sara B
authors.initials: SB
authors.lastname: Hernandez
authors.affiliation: Laboratory for Molecular Infection Medicine Sweden, Department of Molecular Biology, Umeå Centre for Microbial Research, Umeå University, Umeå, Sweden.
authors.forename: Felipe
authors.initials: F
authors.lastname: Cava
authors.affiliation: Department of Chemistry, Indiana University, Bloomington, Indiana, United States of America.|Department of Biology, Indiana University, Bloomington, Indiana, United States of America.
authors.forename: Erkin
authors.initials: E
authors.lastname: Kuru
authors.affiliation: Department of Chemistry, Indiana University, Bloomington, Indiana, United States of America.
authors.forename: Michael S
authors.initials: MS
authors.lastname: Van Nieuwenhze
authors.affiliation: Department of Biology, Indiana University, Bloomington, Indiana, United States of America.
authors.forename: Yves V
authors.initials: YV
authors.lastname: Brun
authors.affiliation: Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
authors.forename: Fernanda M
authors.initials: FM
authors.lastname: Coelho
authors.affiliation: Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
authors.forename: Siegfried
authors.initials: S
authors.lastname: Hapfelmeier
mesh_terms: D000409:Alanine / Q000378:metabolism*; D000411:Alanine Racemase / Q000235:genetics; D000818:Animals; D001329:Autolysis / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism*; D003960:Diaminopimelic Acid / Q000378:metabolism*; D048168:Escherichia coli K12 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D000069196:Gastrointestinal Microbiome; D041981:Gastrointestinal Tract / Q000382:microbiology; D005856:Germ-Free Life; D006801:Humans; D007070:Immunoglobulin A / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D059013:Microbial Consortia; D023421:Models, Animal; D010457:Peptidoglycan / Q000378:metabolism; D013559:Symbiosis
chemical_list: D007070:Immunoglobulin A; D010457:Peptidoglycan; D003960:Diaminopimelic Acid; D000411:Alanine Racemase; D000409:Alanine