{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,22]],"date-time":"2026-10-22T03:45:35Z","timestamp":1792640735362,"version":"4.5.0"},"reference-count":59,"publisher":"Public Library of Science (PLoS)","issue":"3","license":[{"start":{"date-parts":[[2023,3,14]],"date-time":"2023-03-14T00:00:00Z","timestamp":1678752000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100015691","name":"Division of Microbiology and Infectious Diseases, National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01AI141958"],"award-info":[{"award-number":["R01AI141958"]}],"id":[{"id":"10.13039\/100015691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100015691","name":"Division of Microbiology and Infectious Diseases, National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01AI141958"],"award-info":[{"award-number":["R01AI141958"]}],"id":[{"id":"10.13039\/100015691","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS ONE"],"abstract":"<jats:p>The recent development of a system for long-term <jats:italic>in vitro<\/jats:italic> culture of the syphilis spirochete, <jats:italic>Treponema pallidum<\/jats:italic> subsp. <jats:italic>pallidum<\/jats:italic>, has introduced the possibility of detailed genetic analysis of this bacterium. In this study, the <jats:italic>in vitro<\/jats:italic> culture system was used to isolate and characterize clonal populations of <jats:italic>T<\/jats:italic>. <jats:italic>pallidum<\/jats:italic> subsp. <jats:italic>pallidum<\/jats:italic> Nichols, the most widely studied strain. In limiting dilutions experiments, it was possible to establish cultures with inocula as low as 0.5 <jats:italic>T<\/jats:italic>. <jats:italic>pallidum<\/jats:italic> per well despite the long generation time (~35 to 40 hours) of this organism. Six Nichols strain clones isolated by limiting dilution were characterized in detail. All clones exhibited indistinguishable morphology and motility, highly similar <jats:italic>in vitro<\/jats:italic> multiplication rates, and comparable infectivity in the rabbit model (ID50 \u2264 100 bacteria). Genomic sequencing revealed sequence heterogeneity in the form of insertions or deletions at 5 sites, single nucleotide variations at 20 sites, and polynucleotide (polyG\/C) tract length differences at 22 locations. Genomic sequences of the uncloned Nichols strain preparations propagated in rabbits or <jats:italic>in vitro<\/jats:italic> cultures exhibited substantial heterogeneity at these locations, indicating coexistence of many varied \u2018clonotypes\u2019 within these populations. Nearly all genetic variations were specific for the Nichols strain and were not detected in the &gt;280 <jats:italic>T<\/jats:italic>. <jats:italic>pallidum<\/jats:italic> genomic sequences that are currently available. We hypothesize that these Nichols strain-specific sequence variations arose independently either during human infection or within the 110 years since the strain\u2019s initial isolation, and thus represent examples of microevolution and divergence.<\/jats:p>","DOI":"10.1371\/journal.pone.0281187","type":"journal-article","created":{"date-parts":[[2023,3,14]],"date-time":"2023-03-14T17:28:18Z","timestamp":1678814898000},"page":"e0281187","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":17,"title":["Clonal isolates of Treponema pallidum subsp. pallidum Nichols provide evidence for the occurrence of microevolution during experimental rabbit infection and in vitro culture"],"prefix":"10.1371","volume":"18","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-3702-9715","authenticated-orcid":true,"given":"Diane 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