Metagenomic sequencing enables accurate pathogen and antimicrobial susceptibility profiling in complicated UTIs in approximately four hours

Sep 30, 2026

Uploaded by Rowan

Urinary tract infections affect 405 million people worldwide, and conventional culture-based diagnostics can take 2 to 4 days. This study tests eleven culture-independent sample-preparation approaches across 78 complicated UTI patients, followed by real-time nanopore metagenomic sequencing and analysis. Results strongly match culture-based routines, with an optimized workflow achieving 99% accuracy for pathogen identification and 90% accuracy for antimicrobial susceptibility profiling, plus robust detection at low bacterial loads, enabling informed decisions in ~4 hours.

LanguageEnglish

CategoryHealthcare

FormatPDF

Pages16

Reading Time40 min

AccessFree to download

Background

Burden and limitations of current UTI diagnostics

Antimicrobial resistance and need for multiplexed diagnostics

Study approach

Culture-independent sample preparation and real-time nanopore mNGS

Comparison with culture-based clinical routines

Results and performance

Pathogen identification accuracy

Antimicrobial susceptibility profiling accuracy

Robustness at low bacterial loads and added detections

Clinical impact

Turnaround time and cost considerations

DNA yield and pre-screening to reduce costs

Antimicrobial stewardship implications

| | | | | --- | --- | --- | | Article [https://doi.org/10.1038/s41467-025-66865-8](https://doi.org/10.1038/s41467-025-66865-8) | | | | Metagenomic sequencing enables accurate pathogen and antimicrobial susceptibility proﬁling in complicated UTIs in approximately four hours | | | | Received: 19 March 2025
Accepted: 14 November 2025 Check for updates | Anurag Basavaraj Bellankimath1,8, Sverre Branders 1,8, Isabell Kegel2,3, Jawad Ali 1, Fatemeh Asadi1, Truls E. Bjerklund Johansen 4,5, Can Imirzalioglu2,3, Torsten Hain2,3, Florian Wagenlehner 3,6 &
Raﬁ Ahmad 1,7
Urinary tract infections (UTIs) affect 405 million people worldwide. Current diagnostics rely on cultures, which can take 2 to 4 days. This study evaluates eleven culture-independent methods for sample preparation from 78 complicated UTI patients, followed by real-time nanopore sequencing and data analysis. The metagenomic results are highly consistent with culture-based clinical routines (MALDI-TOF/VITEK-2). The optimized method demonstrated an accuracy score of 99%(100/101) for pathogen identiﬁcation and 90%(589/ 653) for antimicrobial susceptibility proﬁling with 95% speciﬁcity. The method’s robustness is highlighted by its ability to accurately identify pathogens with as few as 32 bacterial cells/µL and a low bacterial-to-host cell ratio limit of 0.5. Additionally, mNGS identiﬁed 13 pathogens that routine diagnostics missed, which were subsequently conﬁrmed by Vivalytic or PCR. This method is up to 30% more economical than published studies and commercial kits. DNA yield and ﬂow cytometry can be used for pre-screening to reduce costs, which is crucial for clinical adoption. This research highlights the rapid diagnosis of clinical UTIs using a cost-effective and scalable method that requires around four hours from sample collection to informed decision-making. Furthermore, it aims to improve antimicrobial and diagnostic stewardship by reducing empirical treatment and ensuring more judicious antibiotic use. | | | Urinary Tract Infections (UTIs) are the second most prevalent infec- Untreated UTIs can progress to pyelonephritis2 and urosepsis, which tions worldwide, affecting 405 million people annually and causing account for approximately one-quarter of all sepsis cases3,4. Cathetersigniﬁcant morbidity and economic costs. Over the past three decades, associated UTIs account for 75–95% of hospital-acquired UTIs5. If not global deaths due to UTIs have increased dramatically by 140%1. promptly treated, a secondary blood infection, particularly among | | | 1Department of Biotechnology, University of Inland Norway, Hamar, Norway. 2Institute of Medical Microbiology, Medical Microbiome-Metagenome Unit (M3U), Justus Liebig University Giessen, Giessen, Germany. 3German Center for Infection Research (DZIF), Partner Site Giessen-Marburg-Langen, Giessen, Germany. 4Institute of Clinical Medicine, University of Oslo, Oslo, Norway. 5Institute of Clinical Medicine, University of Aarhus, Aarhus, Denmark. 6Clinic for Urology, Pediatric Urology and Andrology, Justus Liebig University Giessen, Giessen, Germany. 7Institute of Clinical Medicine, Faculty of Health Sciences, UiT—The Arctic University of Norway, Tromsø, Norway. 8These authors contributed equally: Anurag Basavaraj Bellankimath, Sverre Branders. e-mail: raﬁ.[ahmad@inn.no](ahmad@inn.no) catheter-associated UTIs, is known to increase mortality rates to 30% posing a signiﬁcant risk6. In severe UTIs, administering effective antibiotics as soon as possible is paramount. A diverse range of virulence factors and pathogenic strains resistant to multiple drugs contribute to the complexity of UTIs7,8. Rapid and accurate identiﬁcation of pathogens and determination of antimicrobial susceptibility are central to managing UTIs and improving antimicrobial stewardship in an era of increasing antimicrobial resistance (AMR). Furthermore, AMR may be attributed to polygenic traits, which cannot be adequately identiﬁed through

FAQs

Why are current diagnostics for complicated UTIs insufficient?

Culture-based diagnostics rely on growth, which typically takes 2 to 4 days and delays susceptibility results. This makes timely antibiotic selection difficult in severe infections.

What workflow does the study evaluate for UTI diagnosis?

The study assesses eleven culture-independent sample-preparation methods, then applies real-time nanopore sequencing and computational data analysis to characterize pathogens and antimicrobial resistance.

How accurate is the optimized metagenomic approach?

The optimized method reports 99% accuracy for pathogen identification (100/101) and 90% accuracy for antimicrobial susceptibility profiling (589/653), with 95% specificity.

Sep 30, 2026

Uploaded by Rowan