Negative-control-anchored urinary microbiome profiling with absolute 16S quantification: a pilot study in newly diagnosed, treatment-naive bladder cancer and healthy individuals.
| Title: | Negative-control-anchored urinary microbiome profiling with absolute 16S quantification: a pilot study in newly diagnosed, treatment-naive bladder cancer and healthy individuals. |
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| Authors: | Accetto T; Department of Microbiology, Biotechnical Faculty, University of Ljubljana, SI-1000 Ljubljana, Slovenia.; Strašek Smrdel K; Institute of Microbiology and Immunology, Faculty of Medicine, University of Ljubljana, SI-1000 Ljubljana, Slovenia.; Taskovska M; Clinical department of Urology, University Medical Centre Ljubljana, SI-1000 Ljubljana, Slovenia.; Chair of Surgery, Faculty of Medicine, University of Ljubljana, SI-1000 Ljubljana, Slovenia.; Starčič Erjavec M; Department of Microbiology, Biotechnical Faculty, University of Ljubljana, SI-1000 Ljubljana, Slovenia.; Smrkolj T; Clinical department of Urology, University Medical Centre Ljubljana, SI-1000 Ljubljana, Slovenia.; Chair of Surgery, Faculty of Medicine, University of Ljubljana, SI-1000 Ljubljana, Slovenia.; Seme K; Institute of Microbiology and Immunology, Faculty of Medicine, University of Ljubljana, SI-1000 Ljubljana, Slovenia.; Kreft ME; Institute of Cell Biology, Faculty of Medicine, University of Ljubljana, SI-1000 Ljubljana, Slovenia. |
| Source: | FEMS microbiology letters [FEMS Microbiol Lett] 2026 Jan 07; Vol. 373. |
| Publication Type: | Journal Article |
| Language: | English |
| Journal Info: | Publisher: Oxford University Press Country of Publication: England NLM ID: 7705721 Publication Model: Print Cited Medium: Internet ISSN: 1574-6968 (Electronic) Linking ISSN: 03781097 NLM ISO Abbreviation: FEMS Microbiol Lett Subsets: MEDLINE |
| Imprint Name(s): | Publication: 2015- : Oxford Oxford University Press; Original Publication: Amsterdam, Published by Elsevier/North Holland on behalf of the Federation of European Microbiological Societies. |
| MeSH Terms: | Urinary Bladder Neoplasms*/microbiology ; Urinary Bladder Neoplasms*/diagnosis ; Urinary Bladder Neoplasms*/urine ; Microbiota*/genetics ; RNA, Ribosomal, 16S*/genetics ; Urine*/microbiology ; Bacteria*/genetics ; Bacteria*/classification ; Bacteria*/isolation & purification; DNA, Bacterial/genetics ; Humans ; Pilot Projects ; Male ; Middle Aged ; Female ; Aged ; Bacterial Load ; Healthy Volunteers |
| Abstract: | Recent studies utilizing 16S rRNA amplicon sequencing have challenged the notion of urine sterility, yet urine is a low-biomass specimen in which apparent community profiles can be strongly influenced by background signal from reagents and processing. To address this interpretability gap, we integrate culture-independent absolute 16S rRNA gene quantification with urinary 16S amplicon sequencing in a negative-control-anchored workflow. Bacterial load provides a biomass-aware quality control gate that defines interpretable low-biomass thresholds and objective exclusion criteria. As a pilot application, we compared midstream urine collected prior to instrumentation from healthy volunteers and newly diagnosed bladder cancer (BC) patients, quality filtering retained 29 controls and 5 BC cases. Samples > 106 copies/ml typically produced > 10 000 reads; near 105 copies/ml read counts dropped sharply yet remained distinguishable from background. Thirteen negative controls (V3-V4 PCR and stabilization buffer; median 90, mean 124 reads) supported excluding samples with |
| Grant Information: | J7-2594 The Slovenian Research and Innovation Agency |
| Contributed Indexing: | Keywords: absolute quantification; bladder cancer; low biomass; microbiota; midstream urine; negative controls; next-generation sequencing; nonmetric multidimensional scaling; urinary tract infection |
| Substance Nomenclature: | 0 (RNA, Ribosomal, 16S); 0 (DNA, Bacterial) |
| Entry Date(s): | Date Created: 20260217 Date Completed: 20260306 Latest Revision: 20260516 |
| Update Code: | 20260517 |
| PubMed Central ID: | PMC13017691 |
| DOI: | 10.1093/femsle/fnag020 |
| PMID: | 41700712 |
| Database: | MEDLINE |
Journal Article