{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T00:56:51Z","timestamp":1760057811094,"version":"build-2065373602"},"reference-count":58,"publisher":"MDPI AG","issue":"5","license":[{"start":{"date-parts":[[2025,2,25]],"date-time":"2025-02-25T00:00:00Z","timestamp":1740441600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Understanding the interactions between the cervico-vaginal microbiome, immune responses, and sexually transmitted infections (STIs) is crucial for developing targeted diagnostic and therapeutic strategies. Although microbiome analyses are not yet standard practice, integrating them into routine diagnostics could enhance personalized medicine and therapies. We investigated the extent to which partial 16S short-read amplicon microbiome analyses could inform on the presence of six commonly encountered STI-causing pathogens in a patient cohort referred for colposcopy, and whether relevant taxonomic or diagnostic discrepancies occur when using vaginal rather than cervical swabs. The study cohort included cervical and vaginal samples collected from women referred for colposcopy at the University Hospital Bonn between November 2021 and February 2022, due to an abnormal PAP smear or positive hrHPV results. 16S rRNA gene sequencing libraries were prepared targeting the V1\u2013V2 and V4 regions of the 16S RNA gene and sequenced on the Illumina MiSeq. PCR diagnostics for common STI-causing pathogens were conducted using the Allplex STI Essential Assay Kit (Seegene, Seoul, Republic of Korea). Concerning the bacterial microbiome, no significant differences were found between vaginal and cervical samples in terms of prevalence of taxa present or diversity. A total of 95 patients and 171 samples tested positive for at least one among Ureaplasma parvum, Ureaplasma urealyticum, Mycoplasma hominis, Mycoplasma genitalium, Chlamydophila trachomatis or Neisseria gonorrhoeae. Sequencing the V1\u2013V2 region enabled detection of one-third to half of the PCR-positive samples, with the detection likelihood increasing at lower cycle threshold (Ct) values. In contrast, sequencing the V4 region was less effective overall, yielding fewer species-level identifications and a higher proportion of undetermined taxa. We demonstrate that the vaginal microbiome closely mirrors the cervical microbiome, a relationship that has not been explored previously, but which broadens the possibilities for microbiome analysis and pathogen detection and establishes vaginal swabs as a reliable method for detecting the investigated pathogens, with sensitivities comparable with or superior to endocervical swabs. On the other hand, the sensitivity of partial 16S amplicon sequencing appears insufficient for effective STI diagnostics, as it fails to reliably identify or even detect pathogens at higher Ct values.<\/jats:p>","DOI":"10.3390\/ijms26051983","type":"journal-article","created":{"date-parts":[[2025,2,25]],"date-time":"2025-02-25T06:13:18Z","timestamp":1740463998000},"page":"1983","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Homogeneity Between Cervical and Vaginal Microbiomes and the Diagnostic Limitations of 16S Sequencing for STI Pathogens at Higher Ct Values"],"prefix":"10.3390","volume":"26","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2064-3885","authenticated-orcid":false,"given":"Claudio","family":"Neidh\u00f6fer","sequence":"first","affiliation":[{"name":"Institute of Experimental Haematology and Transfusion Medicine, University Hospital Bonn, 53127 Bonn, Germany"},{"name":"Institute of Medical Microbiology, Immunology and Parasitology, University Hospital Bonn, 53127 Bonn, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Mateja","family":"Condic","sequence":"additional","affiliation":[{"name":"Department of Gynecology and Gynecological Oncology, University Hospital Bonn, 53127 Bonn, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nathalie","family":"Hahn","sequence":"additional","affiliation":[{"name":"Institute of Medical Microbiology, Immunology and Parasitology, University Hospital Bonn, 53127 Bonn, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lucia A.","family":"Otten","sequence":"additional","affiliation":[{"name":"Department of Gynecology and Gynecological Oncology, University Hospital Bonn, 53127 Bonn, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8916-7605","authenticated-orcid":false,"given":"Damian J.","family":"Ralser","sequence":"additional","affiliation":[{"name":"Department of Gynecology and Gynecological Oncology, University Hospital Bonn, 53127 Bonn, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nina","family":"Wetzig","sequence":"additional","affiliation":[{"name":"Institute for Functional Gene Analytics, Bonn-Rhein-Sieg University of Applied Sciences, 53757 Sankt Augustin, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ralf","family":"Thiele","sequence":"additional","affiliation":[{"name":"Institute for Functional Gene Analytics, Bonn-Rhein-Sieg University of Applied Sciences, 53757 Sankt Augustin, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Achim","family":"Hoerauf","sequence":"additional","affiliation":[{"name":"Institute of Medical Microbiology, Immunology and Parasitology, University Hospital Bonn, 53127 Bonn, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Marijo","family":"Par\u010dina","sequence":"additional","affiliation":[{"name":"Institute of Medical Microbiology, Immunology and Parasitology, University Hospital Bonn, 53127 Bonn, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,2,25]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"967","DOI":"10.1016\/j.bbrc.2015.12.083","article-title":"Analysis of the microbiome: Advantages of whole genome shotgun versus 16S amplicon sequencing","volume":"469","author":"Ranjan","year":"2016","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Kosti, I., Lyalina, S., Pollard, K., Butte, A., and Sirota, M. (2020). Meta-Analysis of Vaginal Microbiome Data Provides New Insights Into Preterm Birth. Front. Microbiol., 11.","DOI":"10.3389\/fmicb.2020.00476"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1681","DOI":"10.1002\/jcp.25806","article-title":"Subclinical alteration of the cervical\u2013vaginal microbiome in women with idiopathic infertility","volume":"232","author":"Campisciano","year":"2017","journal-title":"J. Cell. Physiol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"199","DOI":"10.1007\/s13167-021-00244-3","article-title":"The interplay between the vaginal microbiome and innate immunity in the focus of predictive, preventive, and personalized medical approach to combat HPV-induced cervical cancer","volume":"12","author":"Kudela","year":"2021","journal-title":"EPMA J."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"61","DOI":"10.1515\/bmc-2022-0005","article-title":"Interaction of cervical microbiome with epigenome of epithelial cells: Significance of inflammation to primary healthcare","volume":"13","author":"Holubekova","year":"2022","journal-title":"Biomol. Concepts"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"341","DOI":"10.1038\/s41576-019-0113-7","article-title":"Clinical metagenomics","volume":"20","author":"Chiu","year":"2019","journal-title":"Nat. Rev. Genet."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Neidh\u00f6fer, C., Klein, N., Y\u00fcr\u00fckt\u00fcmen, A., Hattenhauer, T., Mispelbaum, R., Bode, C., Holderried, T.A., Hoerauf, A., and Par\u010dina, M. (2025). Retrospective analysis of 300 microbial cell-free DNA sequencing results in routine blood stream infection diagnostics. Front. Cell. Infect. Microbiol., 15.","DOI":"10.3389\/fcimb.2025.1504262"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"e52","DOI":"10.1016\/S2666-5247(23)00244-6","article-title":"Performance of clinical metagenomics in France: A prospective observational study","volume":"5","author":"Fourgeaud","year":"2024","journal-title":"Lancet Microbe"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"291","DOI":"10.1038\/s41579-023-00998-9","article-title":"Utilization of the microbiome in personalized medicine","volume":"22","author":"Ratiner","year":"2024","journal-title":"Nat. Rev. Microbiol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"12","DOI":"10.1016\/j.chom.2015.12.016","article-title":"Taking it personally: Personalized utilization of the human microbiome in health and disease","volume":"19","author":"Zmora","year":"2016","journal-title":"Cell Host Microbe"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"541","DOI":"10.1080\/10408398.2013.792772","article-title":"Impact of diet composition on blood glucose regulation","volume":"56","author":"Russell","year":"2016","journal-title":"Crit. Rev. Food Sci. Nutr."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1700","DOI":"10.1007\/s00125-018-4632-0","article-title":"Systematic review assessing the effectiveness of dietary intervention on gut microbiota in adults with type 2 diabetes","volume":"61","author":"Houghton","year":"2018","journal-title":"Diabetologia"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"723","DOI":"10.1007\/s10620-020-06112-w","article-title":"Diet and the Human Gut Microbiome: An International Review","volume":"65","author":"Wilson","year":"2020","journal-title":"Dig. Dis. Sci."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Auriemma, R.S., Scairati, R., Del Vecchio, G., Liccardi, A., Verde, N., Pirchio, R., Pivonello, R., Ercolini, D., and Colao, A. (2021). The Vaginal Microbiome: A Long Urogenital Colonization Throughout Woman Life. Front. Cell. Infect. Microbiol., 11.","DOI":"10.3389\/fcimb.2021.686167"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1989","DOI":"10.1093\/infdis\/jiu004","article-title":"Bacteria in the vaginal microbiome alter the innate immune response and barrier properties of the human vaginal epithelia in a species-specific manner","volume":"209","author":"Doerflinger","year":"2014","journal-title":"J. Infect. Dis."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"543","DOI":"10.1016\/S1286-4579(00)00313-0","article-title":"Role played by lactobacilli in controlling the population of vaginal pathogens","volume":"2","author":"Boris","year":"2000","journal-title":"Microbes Infect."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"221","DOI":"10.1111\/j.1574-6976.1993.tb00020.x","article-title":"Lactic acid bacteria, their metabolic products and interference with microbial growth","volume":"12","author":"Vandenbergh","year":"1993","journal-title":"FEMS Microbiol. Rev."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"153","DOI":"10.1089\/bfm.2010.0001","article-title":"Inhibition of human immunodeficiency virus type 1 by lactic acid bacteria from human breastmilk","volume":"5","author":"Maldonado","year":"2010","journal-title":"Breastfeed. Med."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"3793","DOI":"10.1007\/s00203-021-02414-3","article-title":"Vaginal microbiome: Normalcy vs dysbiosis","volume":"203","author":"Saraf","year":"2021","journal-title":"Arch. Microbiol."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Avitabile, E., Menotti, L., Giordani, B., Croatti, V., Parolin, C., and Vitali, B. (2024). Vaginal Lactobacilli Supernatants Protect from Herpes Simplex Virus Type 1 Infection in Cell Culture Models. Int. J. Mol. Sci., 25.","DOI":"10.3390\/ijms25052492"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Torcia, M. (2019). Interplay among Vaginal Microbiome, Immune Response and Sexually Transmitted Viral Infections. Int. J. Mol. Sci., 20.","DOI":"10.3390\/ijms20020266"},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Dong, M., Dong, Y., Bai, J., Li, H., Ma, X., Li, B., Wang, C., Li, H., Qi, W., and Wang, Y. (2023). Interactions between microbiota and cervical epithelial, immune, and mucus barrier. Front. Cell. Infect. Microbiol., 13.","DOI":"10.3389\/fcimb.2023.1124591"},{"key":"ref_23","unstructured":"Garcia, M.R., Leslie, S.W., and Wray, A.A. (2025). Sexually Transmitted Infections. StatPearls, StatPearls Publishing. Available online: https:\/\/www.ncbi.nlm.nih.gov\/books\/NBK560808\/."},{"key":"ref_24","first-page":"287","article-title":"Bacterial sexually transmitted infections","volume":"17","author":"Buder","year":"2019","journal-title":"JDDG J. Dtsch. Dermatol. Ges."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"6489","DOI":"10.1007\/s00432-023-04599-0","article-title":"Analysis of the cervical microbiome in women from the German national cervical cancer screening program","volume":"149","author":"Condic","year":"2023","journal-title":"J. Cancer Res. Clin. Oncol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e8013","DOI":"10.1136\/bmj.e8013","article-title":"Assessment of best single sample for finding chlamydia in women with and without symptoms: A diagnostic test study","volume":"345","author":"Schoeman","year":"2012","journal-title":"BMJ"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"408","DOI":"10.5811\/westjem.2022.3.53812","article-title":"Vaginal Swabs Are Non-Inferior to Endocervical Swabs for Sexually Transmitted Infection testing in the Emergency Department","volume":"23","author":"Krause","year":"2022","journal-title":"West. J. Emerg. Med."},{"key":"ref_28","first-page":"A101","article-title":"P051 Accuracy of sexually transmitted infections testing on self-collected vaginal samples versus cervical samples","volume":"95","author":"Cocuzza","year":"2019","journal-title":"Sex. Transm. Infect."},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Lunny, C., Taylor, D., Hoang, L., Wong, T., Gilbert, M., Lester, R., Krajden, M., and Ogilvie, G. (2015). Self-Collected versus Clinician-Collected Sampling for Chlamydia and Gonorrhea Screening: A Systemic Review and Meta-Analysis. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0132776"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1328","DOI":"10.1016\/j.cmi.2018.03.013","article-title":"Identifying a consensus sample type to test for Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, Trichomonas vaginalis and Human papillomavirus","volume":"24","author":"Coorevits","year":"2018","journal-title":"Clin. Microbiol. Infect."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"733","DOI":"10.3949\/ccjm.86a.18098","article-title":"STI update: Testing, treatment, and emerging threats","volume":"86","author":"Hlatshwayo","year":"2019","journal-title":"Cleve Clin. J. Med."},{"key":"ref_32","unstructured":"Centers for Disease Control and Prevention (2014). Sexually transmitted diseases treatment guidelines. MMWR, 63, 1\u2013137. Available online: https:\/\/www.cdc.gov\/mmwr\/preview\/mmwrhtml\/rr6302a1.htm."},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Zhou, L., Rodas, A., Llangar\u00ed, L., Sandoval, N., Cooper, P., and Sadiq, S. (2020). Single gene targeted nanopore sequencing enables simultaneous identification and antimicrobial resistance detection of sexually transmitted infections. PLoS ONE, 17.","DOI":"10.1101\/2020.11.06.371310"},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Neidh\u00f6fer, C., Bagniceva, M., Wetzig, N., Sieber, M.A., Thiele, R., and Par\u010dina, M. (2023). Pragmatic Considerations When Extracting DNA for Metagenomics Analyses of Clinical Samples. Int. J. Mol. Sci., 24.","DOI":"10.3390\/ijms241411262"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1845","DOI":"10.1111\/jdv.15146","article-title":"Should we be testing for urogenital Mycoplasma hominis, Ureaplasma parvum and Ureaplasma urealyticum in men and women?\u2014A position statement from the European STI Guidelines Editorial Board","volume":"32","author":"Horner","year":"2018","journal-title":"J. Eur. Acad. Dermatol. Venereol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"112","DOI":"10.1071\/MA24033","article-title":"Direct sequencing technologies for bacterial sexually transmitted infections","volume":"45","author":"Amy","year":"2024","journal-title":"Microbiol. Aust."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"604","DOI":"10.1097\/01.ede.0000173042.07579.17","article-title":"Evaluation of Nucleic Acid Amplification Tests in the Absence of a Perfect Gold-Standard Test: A Review of the Statistical and Epidemiologic Issues","volume":"16","author":"Hadgu","year":"2005","journal-title":"Epidemiology"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"hvad097-552","DOI":"10.1093\/clinchem\/hvad097.552","article-title":"B-226 Clinical Evaluation of ElsiQ\u2122 STI-12 Detection Kit 3\/4 for Detection of Human Sexual Transmitted Infection Pathogens in Human Vaginal Swab and\/or Urine Specimens","volume":"69","author":"Ahn","year":"2023","journal-title":"Clin. Chem."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"589","DOI":"10.1016\/j.trechm.2021.03.009","article-title":"Materials chemistry-enabled platforms in detecting sexually transmitted infections: Progress towards point-of-care tests","volume":"3","author":"Farokhzad","year":"2021","journal-title":"Trends Chem."},{"key":"ref_40","doi-asserted-by":"crossref","unstructured":"Christiansen, M.T., Brown, A.C., Kundu, S., Tutill, H.J., Williams, R., Brown, J.R., Holdstock, J., Holland, M.J., Stevenson, S., and Dave, J. (2014). Whole-genome enrichment and sequencing of Chlamydia trachomatis directly from clinical samples. BMC Infect. Dis., 14.","DOI":"10.1186\/s12879-014-0591-3"},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Plummer, E.L., Murray, G.L., Bodiyabadu, K., Su, J., Garland, S.M., Bradshaw, C.S., Read, T.R.H., Tabrizi, S.N., and Danielewski, J.A. (2020). A custom amplicon sequencing approach to detect resistance associated mutations and sequence types in Mycoplasma genitalium. J. Microbiol. Methods, 179.","DOI":"10.1016\/j.mimet.2020.106089"},{"key":"ref_42","doi-asserted-by":"crossref","unstructured":"Chiribau, C.B., Schmedes, S., Dong, Y., Tarigopula, N., Tekin, O., Cannons, A., Roberts, J., Haiduven, D., and Crowe, S.R. (2024). Detection of resistance to macrolides and fluoroquinolones in Mycoplasma genitalium by targeted next-generation sequencing. Microbiol. Spectr., 12.","DOI":"10.1128\/spectrum.03845-23"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"610","DOI":"10.1093\/clinchem\/hvaa306","article-title":"Multiplex PCR and Nanopore Sequencing of Genes Associated with Antimicrobial Resistance in Neisseria gonorrhoeae Directly from Clinical Samples","volume":"67","author":"Zhang","year":"2021","journal-title":"Clin. Chem."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"65","DOI":"10.1136\/sextrans-2015-052422","article-title":"Epidemiological typing of Neisseria gonorrhoeae and detection of markers associated with antimicrobial resistance directly from urine samples using next generation sequencing","volume":"93","author":"Graham","year":"2017","journal-title":"Sex. Transm. Infect."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"330","DOI":"10.1111\/jam.14380","article-title":"Generating amplicon reads for microbial community assessment with next-generation sequencing","volume":"128","author":"Tretyn","year":"2020","journal-title":"J. Appl. Microbiol."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"e37","DOI":"10.1093\/nar\/gku1341","article-title":"Insight into biases and sequencing errors for amplicon sequencing with the Illumina MiSeq platform","volume":"43","author":"Schirmer","year":"2015","journal-title":"Nucleic Acids Res."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"219","DOI":"10.1097\/PAT.0000000000000239","article-title":"Opportunities and pitfalls of molecular testing for detecting sexually transmitted pathogens","volume":"47","author":"Trembizki","year":"2015","journal-title":"Pathology"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"417","DOI":"10.1007\/s11908-010-0136-x","article-title":"Role of Mycoplasma and Ureaplasma Species in Female Lower Genital Tract Infections","volume":"12","author":"Patel","year":"2010","journal-title":"Curr. Infect. Dis. Rep."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"190","DOI":"10.1016\/j.siny.2008.11.009","article-title":"Congenital and opportunistic infections: Ureaplasma species and Mycoplasma hominis","volume":"14","author":"Waites","year":"2009","journal-title":"Semin. Fetal Neonatal. Med."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.ajog.2007.11.031","article-title":"Twenty percent of very preterm neonates (23\u201332 weeks of gestation) are born with bacteremia caused by genital Mycoplasmas","volume":"198","author":"Romero","year":"2008","journal-title":"Am. J. Obstet. Gynecol."},{"key":"ref_51","doi-asserted-by":"crossref","unstructured":"Zhang, N., Wang, R., Li, X., Liu, X., Tang, Z., and Liu, Y. (2014). Are Ureaplasma spp. a cause of nongonococcal urethritis? A systematic review and meta-analysis. PLoS ONE, 9.","DOI":"10.1371\/journal.pone.0113771"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"169","DOI":"10.1080\/1040841X.2020.1736986","article-title":"Ureaplasma species and preterm birth: Current perspectives","volume":"46","author":"Sprong","year":"2020","journal-title":"Crit. Rev. Microbiol."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"1650","DOI":"10.1111\/jdv.13849","article-title":"2016 European guideline on Mycoplasma genitalium infections","volume":"30","author":"Jensen","year":"2016","journal-title":"J. Eur. Acad. Dermatol. Venereol."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"852","DOI":"10.1038\/s41587-019-0209-9","article-title":"Reproducible, interactive, scalable and extensible microbiome data science using QIIME 2","volume":"37","author":"Bolyen","year":"2019","journal-title":"Nat. Biotechnol."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"581","DOI":"10.1038\/nmeth.3869","article-title":"DADA2: High-resolution sample inference from Illumina amplicon data","volume":"13","author":"Callahan","year":"2016","journal-title":"Nat. Methods"},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"D590","DOI":"10.1093\/nar\/gks1219","article-title":"The SILVA ribosomal RNA gene database project: Improved data processing and web-based tools","volume":"41","author":"Quast","year":"2013","journal-title":"Nucleic Acids Res."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1186\/s40168-018-0470-z","article-title":"Optimizing taxonomic classification of marker-gene amplicon sequences with QIIME 2\u2019s q2-feature-classifier plugin","volume":"6","author":"Bokulich","year":"2018","journal-title":"Microbiome"},{"key":"ref_58","doi-asserted-by":"crossref","unstructured":"Robeson, M.S., O\u2019Rourke, D.R., Kaehler, B.D., Ziemski, M., Dillon, M.R., Foster, J.T., and Bokulich, N.A. (2021). RESCRIPt: Reproducible sequence taxonomy reference database management. PLoS Comput. Biol., 17.","DOI":"10.1371\/journal.pcbi.1009581"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/5\/1983\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T16:42:01Z","timestamp":1760028121000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/5\/1983"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,2,25]]},"references-count":58,"journal-issue":{"issue":"5","published-online":{"date-parts":[[2025,3]]}},"alternative-id":["ijms26051983"],"URL":"https:\/\/doi.org\/10.3390\/ijms26051983","relation":{},"ISSN":["1422-0067"],"issn-type":[{"type":"electronic","value":"1422-0067"}],"subject":[],"published":{"date-parts":[[2025,2,25]]}}}