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Upon entry into human lungs, they may infect lung macrophages, causing Legionnaires' disease (LD), an atypical pneumonia, using similar mechanisms as in their protozoan hosts, despite the 2 hosts being separated by a billion years of evolution. In this study, we used experimental evolution to identify genes conferring host specificity to L. pneumophila. To this end, we passaged L. pneumophila in 2 different hosts\u2014Acanthamoeba castellanii and the human macrophage-like cells U937\u2014separately and by switching between the hosts twice a week for a year. In total, we identified 1,518 mutations present in at least 5% of the population at the time of sampling. Forty-nine mutations were fixed in the 18 populations at the end of the experiment. Two interesting groups of mutations included (i) mutations in 4 different strain-specific genes involved in lipopolysaccharide (LPS) synthesis, found only in the lineages passaged with A. castellanii and (ii) mutations in the gene coding for LerC, a key regulator of protein effector expression, which was independently mutated in 6 lineages grown in presence of the macrophage cells. We propose that the mutations degrading the function of the regulator LerC improve the fitness of L. pneumophila in human-derived cells and that modifications in the LPS are beneficial for growth in A. castellanii. This study is a first step in further investigating determinants of host specificity in L. pneumophila.<\/jats:p>","DOI":"10.1093\/molbev\/msaf161","type":"journal-article","created":{"date-parts":[[2025,7,2]],"date-time":"2025-07-02T08:02:53Z","timestamp":1751443373000},"source":"Crossref","is-referenced-by-count":4,"title":["Host-Specific Adaptation of\n                    <i>Legionella pneumophila<\/i>\n                    to Single and Multiple Hosts"],"prefix":"10.1093","volume":"42","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3229-4029","authenticated-orcid":false,"given":"Ana\u00edsa B","family":"Moreno","sequence":"first","affiliation":[{"name":"Department of Medical Biochemistry and Microbiology, Science for Life Laboratories, Uppsala University , Uppsala 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, Legionella Pathogenesis, Lyon ,","place":["France"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christian","family":"Dobre-Lereanu","sequence":"additional","affiliation":[{"name":"Department of Medical Biochemistry and Microbiology, Science for Life Laboratories, Uppsala University , Uppsala ,","place":["Sweden"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6640-2174","authenticated-orcid":false,"given":"Dan I","family":"Andersson","sequence":"additional","affiliation":[{"name":"Department of Medical Biochemistry and Microbiology, Science for Life Laboratories, Uppsala University , Uppsala ,","place":["Sweden"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8354-2398","authenticated-orcid":false,"given":"Lionel","family":"Guy","sequence":"additional","affiliation":[{"name":"Department of Medical Biochemistry and Microbiology, Science for Life Laboratories, Uppsala 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