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Especially,\n                      <jats:italic>Eubacterium limosum<\/jats:italic>\n                      and closely related strains attract attention not only for their capability to ferment C1 gases and liquids, but also due to their ability to produce butyrate. Apart from its well-elucidated metabolism,\n                      <jats:italic>E. limosum<\/jats:italic>\n                      is also genetically accessible, which makes it an interesting candidate to be an industrial biocatalyst.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      In this study, we examined genomic, phylogenetic, and physiologic features of\n                      <jats:italic>E. limosum<\/jats:italic>\n                      and the closest related species\n                      <jats:italic>E.\u00a0callanderi<\/jats:italic>\n                      as well as\n                      <jats:italic>E. maltosivorans<\/jats:italic>\n                      . We sequenced the genomes of the six\n                      <jats:italic>Eubacterium<\/jats:italic>\n                      strains \u2018FD\u2019 (DSM 3662\n                      <jats:sup>T<\/jats:sup>\n                      ), \u2018Marburg\u2019 (DSM 3468), \u20182A\u2019 (DSM 2593), \u201811A\u2019 (DSM 2594), \u2018G14\u2019 (DSM 107592), and \u201832\u2019 (DSM 20517) and subsequently compared these with previously available genomes of the\n                      <jats:italic>E. limosum<\/jats:italic>\n                      type strain (DSM 20543\n                      <jats:sup>T<\/jats:sup>\n                      ) as well as the strains \u2018B2\u2019, \u2018KIST612\u2019, \u2018YI\u2019 (DSM 105863\n                      <jats:sup>T<\/jats:sup>\n                      ), and \u2018SA11\u2019. This comparison revealed a close relationship between all eleven\n                      <jats:italic>Eubacterium<\/jats:italic>\n                      strains, forming three distinct clades:\n                      <jats:italic>E. limosum<\/jats:italic>\n                      ,\n                      <jats:italic>E.\u00a0callanderi<\/jats:italic>\n                      , and\n                      <jats:italic>E. maltosivorans<\/jats:italic>\n                      . Moreover, we identified the gene clusters responsible for methanol utilization as well as genes mediating chain elongation in all analyzed strains. Subsequent growth experiments revealed that strains of all three clades can convert methanol and produce acetate, butyrate, and hexanoate via reverse \u03b2-oxidation. Additionally, we used a harmonized electroporation protocol and successfully transformed eight of these\n                      <jats:italic>Eubacterium<\/jats:italic>\n                      strains to enable recombinant plasmid-based expression of the gene encoding the fluorescence-activating and absorption shifting tag (FAST). Engineered\n                      <jats:italic>Eubacterium<\/jats:italic>\n                      strains were verified regarding their FAST-mediated fluorescence at a single-cell level using a flow cytometry approach. Eventually, strains \u2018FD\u2019 (DSM 3662\n                      <jats:sup>T<\/jats:sup>\n                      ), \u20182A\u2019 (DSM 2593), \u201811A\u2019 (DSM 2594), and \u201832\u2019 (DSM 20517) were genetically engineered for the first time.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>\n                      Strains of\n                      <jats:italic>E. limosum<\/jats:italic>\n                      ,\n                      <jats:italic>E. callanderi<\/jats:italic>\n                      , and\n                      <jats:italic>E. maltosivorans<\/jats:italic>\n                      are outstanding candidates as biocatalysts for anaerobic C1-substrate conversion into valuable biocommodities. A large variety of strains is genetically accessible using a harmonized electroporation protocol, and FAST can serve as a reliable fluorescent reporter protein to characterize genetically engineered cells. In total eleven strains have been assigned to distinct clades, providing a clear and updated classification. Thus, the description of respective\n                      <jats:italic>Eubacterium<\/jats:italic>\n                      species has been emended, improved, aligned, and is requested to be implemented in respective databases.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1186\/s12934-024-02301-8","type":"journal-article","created":{"date-parts":[[2024,1,17]],"date-time":"2024-01-17T15:02:01Z","timestamp":1705503721000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":24,"title":["Refining and illuminating acetogenic Eubacterium strains for reclassification and metabolic engineering"],"prefix":"10.1186","volume":"23","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0238-4995","authenticated-orcid":false,"given":"Maximilian","family":"Flaiz","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2473-6202","authenticated-orcid":false,"given":"Anja","family":"Poehlein","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0007-8561-4341","authenticated-orcid":false,"given":"Wiebke","family":"Wilhelm","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0834-3780","authenticated-orcid":false,"given":"Alexander","family":"Mook","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8646-7925","authenticated-orcid":false,"given":"Rolf","family":"Daniel","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5586-221X","authenticated-orcid":false,"given":"Peter","family":"D\u00fcrre","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3802-9485","authenticated-orcid":false,"given":"Frank R.","family":"Bengelsdorf","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2024,1,17]]},"reference":[{"key":"2301_CR1","doi-asserted-by":"publisher","first-page":"168","DOI":"10.1016\/j.copbio.2019.10.002","volume":"62","author":"CA Cotton","year":"2020","unstructured":"Cotton CA, Claassens NJ, Benito-Vaquerizo S, Bar-Even A. 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