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In this study, we examined the role of 6S RNA in murine infectivity and tick persistence of the Lyme disease spirochete <jats:italic>Borrelia<\/jats:italic> (<jats:italic>Borreliella<\/jats:italic>) <jats:italic>burgdorferi<\/jats:italic>. <jats:italic>B. burgdorferi<\/jats:italic> 6S RNA (Bb6S RNA) binds to RNA polymerase, is expressed independent of growth phase or nutrient stress in culture, and is processed by RNase Y. We found that <jats:italic>rny<\/jats:italic> (<jats:italic>bb0504<\/jats:italic>), the gene encoding RNase Y, is essential for <jats:italic>B. burgdorferi<\/jats:italic> growth, while <jats:italic>ssrS<\/jats:italic>, the gene encoding 6S RNA, is not essential, indicating a broader role for RNase Y activity in the spirochete. Bb6S RNA regulates expression of the <jats:italic>ospC<\/jats:italic> and <jats:italic>dbpA<\/jats:italic> genes encoding outer surface protein C and decorin binding protein A, respectively, which are lipoproteins important for host infection. The highest levels of Bb6S RNA are found when the spirochete resides in unfed nymphs. <jats:italic>ssrS<\/jats:italic> mutants lacking Bb6S RNA were compromised for infectivity by needle inoculation, but injected mice seroconverted, indicating an ability to activate the adaptive immune response. <jats:italic>ssrS<\/jats:italic> mutants were successfully acquired by larval ticks and persisted through fed nymphs. Bb6S RNA is one of the first regulatory RNAs identified in <jats:italic>B. burgdorferi<\/jats:italic> that controls the expression of lipoproteins involved in host infectivity.<\/jats:p>","DOI":"10.1111\/mmi.14427","type":"journal-article","created":{"date-parts":[[2019,11,19]],"date-time":"2019-11-19T18:45:20Z","timestamp":1574189120000},"page":"399-417","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":8,"title":["Characterization of 6S RNA in the Lyme disease spirochete"],"prefix":"10.1111","volume":"113","author":[{"given":"Dan","family":"Drecktrah","sequence":"first","affiliation":[{"name":"Division of Biological Sciences University of Montana Missoula MT USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Laura S.","family":"Hall","sequence":"additional","affiliation":[{"name":"Division of Biological Sciences University of Montana Missoula MT USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Amanda J.","family":"Brinkworth","sequence":"additional","affiliation":[{"name":"Division of Biological Sciences University of Montana Missoula MT USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jeanette R.","family":"Comstock","sequence":"additional","affiliation":[{"name":"Division of Biological Sciences University of Montana Missoula MT USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Karen M.","family":"Wassarman","sequence":"additional","affiliation":[{"name":"Department of Bacteriology University of Wisconsin\u2010Madison Madison WI USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8352-7593","authenticated-orcid":false,"given":"D. 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