{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,4,12]],"date-time":"2026-04-12T04:43:43Z","timestamp":1775969023585,"version":"3.50.1"},"posted":{"date-parts":[[2025,5,7]]},"group-title":"Microbiology","reference-count":38,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,5,7]],"date-time":"2025-05-07T00:00:00Z","timestamp":1746576000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000057","name":"National Institute of General Medical Sciences","doi-asserted-by":"crossref","award":["GM140886"],"award-info":[{"award-number":["GM140886"]}],"id":[{"id":"10.13039\/100000057","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100000057","name":"National Institute of General Medical Sciences","doi-asserted-by":"crossref","award":["P20GM103432"],"award-info":[{"award-number":["P20GM103432"]}],"id":[{"id":"10.13039\/100000057","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,5,5]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  <jats:italic>Streptococcus mutans<\/jats:italic>\n                  is a major cause of dental caries worldwide. Targeted therapeutic strategies to eradicate\n                  <jats:italic>S. mutans<\/jats:italic>\n                  include oral phage rinses. In this study, we investigated how phage resistance develops in\n                  <jats:italic>S. mutans<\/jats:italic>\n                  . As a model phage, we used \u0278APCM01, which is known to infect a serotype e strain. We isolated and sequenced the genomes of 15 spontaneous resistant mutants and found that 10 had acquired novel CRISPR spacers targeting the phage, with a total of 18 new spacers identified. Additionally, eight strains contained mutations in rhamnose-glucose polysaccharide (RGP) biosynthetic genes, three of which also acquired spacers. Only the\n                  <jats:italic>rgp<\/jats:italic>\n                  mutants exhibited defects in phage absorption, supporting the role of these cell surface glycans as the phage receptor. Mutations in\n                  <jats:italic>rgpF<\/jats:italic>\n                  and the newly identified gene\n                  <jats:italic>rgpX<\/jats:italic>\n                  led to severe cell division defects and impaired biofilm formation, the latter of which shared by the\n                  <jats:italic>rgpD<\/jats:italic>\n                  mutant. Thus,\n                  <jats:italic>rgp<\/jats:italic>\n                  mutations confer phage resistance but impose severe fitness costs, limiting pathogenic potential. Surprisingly, we found that \u0278APCM01 was capable of binding to and injecting its genome into UA159, a model serotype c strain. However, UA159 was resistant to infection due to an unknown post-entry defense mechanism. Consequently, \u0278APCM01 has the potential to infect both major serotypes associated with dental caries.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Repositories<\/jats:title>\n                  <jats:p>\n                    The genome sequence of\n                    <jats:italic>Streptococcus mutans<\/jats:italic>\n                    DPC6143 was deposited at NCBI with the accession number NZ_CP172847.1.\n                  <\/jats:p>\n                <\/jats:sec>","DOI":"10.1101\/2025.05.05.652303","type":"posted-content","created":{"date-parts":[[2025,5,7]],"date-time":"2025-05-07T04:40:12Z","timestamp":1746592812000},"source":"Crossref","is-referenced-by-count":0,"title":["Acquired CRISPR spacers and rhamnose-glucose polysaccharide defects confer resistance to\n                  <i>Streptococcus mutans<\/i>\n                  phage \u0278APCM01"],"prefix":"10.64898","author":[{"given":"Lucas A.","family":"Wall","sequence":"first","affiliation":[{"name":"Department of Molecular Biology, University of Wyoming, Laramie, WY 82071, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0273-1371","authenticated-orcid":false,"given":"Daniel","family":"Wall","sequence":"additional","affiliation":[{"name":"Department of Molecular Biology, University of Wyoming, Laramie, WY 82071, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2026041120552862000_2025.05.05.652303v1.1","doi-asserted-by":"publisher","DOI":"10.1177\/0022034517750572"},{"key":"2026041120552862000_2025.05.05.652303v1.2","doi-asserted-by":"crossref","unstructured":"Abranches J , Zeng L , Kajfasz JK , Palmer SR , Chakraborty B , Wen ZT , et al. Biology of oral Streptococci. Microbiol Spectr. 2018;6(5).","DOI":"10.1128\/microbiolspec.GPP3-0042-2018"},{"key":"2026041120552862000_2025.05.05.652303v1.3","doi-asserted-by":"crossref","unstructured":"Hooshiar MH , Salari S , Nasiri K , Salim US , Saeed LM , Yasamineh S , et al. The potential use of bacteriophages as antibacterial agents in dental infection. Virol J. 2024;21(1).","DOI":"10.1186\/s12985-024-02510-y"},{"key":"2026041120552862000_2025.05.05.652303v1.4","doi-asserted-by":"crossref","unstructured":"Dalmasso M , de Haas E , Neve H , Strain R , Cousin FJ , Stockdale SR , et al. Isolation of a novel phage with activity against biofilms. Plos One. 2015;10(9).","DOI":"10.1371\/journal.pone.0138651"},{"key":"2026041120552862000_2025.05.05.652303v1.5","doi-asserted-by":"publisher","DOI":"10.1111\/j.1574-6968.2007.00873.x"},{"key":"2026041120552862000_2025.05.05.652303v1.6","doi-asserted-by":"crossref","unstructured":"Ben-Zaken H , Kraitman R , Coppenhagen-Glazer S , Khalifa L , Alkalay-Oren S , Gelman D , et al. Isolation and characterization of phage as a possible treatment agent for caries. Viruses. 2021;13(5).","DOI":"10.3390\/v13050825"},{"key":"2026041120552862000_2025.05.05.652303v1.7","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.07726-11"},{"key":"2026041120552862000_2025.05.05.652303v1.8","doi-asserted-by":"publisher","DOI":"10.1016\/j.isci.2023.108465"},{"key":"2026041120552862000_2025.05.05.652303v1.9","doi-asserted-by":"publisher","DOI":"10.1007\/BF01576015"},{"key":"2026041120552862000_2025.05.05.652303v1.10","doi-asserted-by":"publisher","DOI":"10.7717\/peerj.4057"},{"key":"2026041120552862000_2025.05.05.652303v1.11","doi-asserted-by":"crossref","unstructured":"Leprince A , Mahillon J . Phage adsorption to Gram-positive bacteria. Viruses. 2023;15(1).","DOI":"10.3390\/v15010196"},{"key":"2026041120552862000_2025.05.05.652303v1.12","doi-asserted-by":"publisher","DOI":"10.1126\/science.1138140"},{"key":"2026041120552862000_2025.05.05.652303v1.13","doi-asserted-by":"crossref","unstructured":"Mosterd C , Moineau S . Characterization of a type II-A CRISPR-Cas system in Streptococcus mutans . Msphere. 2020;5(3).","DOI":"10.1128\/mSphere.00235-20"},{"key":"2026041120552862000_2025.05.05.652303v1.14","doi-asserted-by":"publisher","DOI":"10.1128\/JB.02333-14"},{"key":"2026041120552862000_2025.05.05.652303v1.15","doi-asserted-by":"publisher","DOI":"10.1099\/mic.0.027508-0"},{"key":"2026041120552862000_2025.05.05.652303v1.16","doi-asserted-by":"publisher","DOI":"10.1111\/j.1574-6968.2009.01546.x"},{"key":"2026041120552862000_2025.05.05.652303v1.17","doi-asserted-by":"crossref","unstructured":"Silva JB , Storms Z , Sauvageau D . Host receptors for bacteriophage adsorption. FEMS Microbiol Lett. 2016;363(4).","DOI":"10.1093\/femsle\/fnw002"},{"key":"2026041120552862000_2025.05.05.652303v1.18","doi-asserted-by":"publisher","DOI":"10.1016\/j.csbj.2021.07.011"},{"key":"2026041120552862000_2025.05.05.652303v1.19","doi-asserted-by":"crossref","unstructured":"Labrie SJ , Mosterd C , Loignon S , Dupuis M\u00c9 , Desjardins P , Rousseau GM , et al. A mutation in the methionine aminopeptidase gene provides phage resistance in. Sci Rep-Uk. 2019;9.","DOI":"10.1038\/s41598-019-49975-4"},{"key":"2026041120552862000_2025.05.05.652303v1.20","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.29397"},{"key":"2026041120552862000_2025.05.05.652303v1.21","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkac976"},{"key":"2026041120552862000_2025.05.05.652303v1.22","doi-asserted-by":"crossref","unstructured":"Mosterd C , Moineau S . Primed CRISPR-Cas adaptation and impaired phage adsorption in Streptococcus mutans. Msphere. 2021;6(3).","DOI":"10.1128\/mSphere.00185-21"},{"key":"2026041120552862000_2025.05.05.652303v1.23","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbc.2022.102488"},{"key":"2026041120552862000_2025.05.05.652303v1.24","doi-asserted-by":"publisher","DOI":"10.1038\/S41586-024-07487-W"},{"key":"2026041120552862000_2025.05.05.652303v1.25","doi-asserted-by":"crossref","unstructured":"Kovacs CJ , Faustoferri RC , Quivey RG, Jr . RgpF Is required for maintenance of stress tolerance and virulence in Streptococcus mutans. J Bacteriol. 2017;199(24).","DOI":"10.1128\/JB.00497-17"},{"key":"2026041120552862000_2025.05.05.652303v1.26","doi-asserted-by":"crossref","unstructured":"Bischer AP , Kovacs CJ , Faustoferri RC , Quivey RG, Jr . Disruption of L-rhamnose biosynthesis results in severe growth defects in Streptococcus mutans. J Bacteriol. 2020;202(6).","DOI":"10.1128\/JB.00728-19"},{"key":"2026041120552862000_2025.05.05.652303v1.27","doi-asserted-by":"publisher","DOI":"10.1128\/JB.180.21.5803-5807.1998"},{"key":"2026041120552862000_2025.05.05.652303v1.28","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.70.6.2891-2898.2002"},{"key":"2026041120552862000_2025.05.05.652303v1.29","doi-asserted-by":"publisher","DOI":"10.1101\/2022.04.08.487609"},{"key":"2026041120552862000_2025.05.05.652303v1.30","doi-asserted-by":"publisher","DOI":"10.1002\/pro.4519"},{"key":"2026041120552862000_2025.05.05.652303v1.31","doi-asserted-by":"publisher","DOI":"10.1111\/omi.2015.30.issue-6"},{"key":"2026041120552862000_2025.05.05.652303v1.32","doi-asserted-by":"publisher","DOI":"10.1038\/s41589-021-00803-9"},{"key":"2026041120552862000_2025.05.05.652303v1.33","doi-asserted-by":"publisher","DOI":"10.1111\/j.1399-302X.2007.00412.x"},{"key":"2026041120552862000_2025.05.05.652303v1.34","doi-asserted-by":"crossref","unstructured":"Cai JN , Jung JE , Lee MH , Choi HM , Jeon JG . Sucrose challenges to Streptococcus mutans biofilms and the curve fitting for the biofilm changes. FEMS Microbiol Ecol. 2018;94(7).","DOI":"10.1093\/femsec\/fiy091"},{"key":"2026041120552862000_2025.05.05.652303v1.35","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-025-56205-1"},{"key":"2026041120552862000_2025.05.05.652303v1.36","doi-asserted-by":"publisher","DOI":"10.1111\/mmi.14330"},{"key":"2026041120552862000_2025.05.05.652303v1.37","doi-asserted-by":"publisher","DOI":"10.1021\/jacs.4c06658"},{"key":"2026041120552862000_2025.05.05.652303v1.38","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2019.09.012"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.1101\/2025.05.05.652303","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,4,12]],"date-time":"2026-04-12T03:55:33Z","timestamp":1775966133000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.1101\/2025.05.05.652303"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,5,7]]},"references-count":38,"URL":"https:\/\/doi.org\/10.1101\/2025.05.05.652303","relation":{},"subject":[],"published":{"date-parts":[[2025,5,7]]},"subtype":"preprint"}}