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Farnleitner, A.

Publications and source records attributed to Farnleitner, A..

2 recordsLinked to original sources
bioRxiv2025

A rapid ionic liquid-based DNA extraction method for molecular diagnostics of urinary tract infections

Rapid and reliable DNA extraction from urine is a critical bottleneck in advancing molecular diagnostics for urinary tract infections (UTIs) in both centralized and decentralized settings. Here, we present an ionic liquid-based DNA extraction method (IL-DEx) that enables recovery of bacterial DNA from urine samples in under 30 minutes using minimal equipment and no hazardous chemicals. IL-DEx was benchmarked against a widely used commercial kit (QIAamp DNA Mini Kit, QIAGEN) using reference strains, clinical isolates, and spiked urine samples. For gram-negative bacteria, IL-DEx achieved comparable DNA yields (47-102% relative efficiency), while recoveries from gram-positive bacteria were lower (0.7-8%) but sufficient for downstream detection. Quantitative PCR (qPCR) revealed linear DNA recovery across five to six orders of magnitude (108-102 CFU/ml, R2 >0.99), with detection limits of [~]102-103 CFU/ml for gram-negatives and [~]103-104 CFU/ml for gram-positives using 1 ml urine. Clinical evaluation with 13 patient urine samples (ten culture-positive, three culture-negative) demonstrated that IL-DEx reliably enabled pathogen detection by qPCR and full-length 16S rRNA gene sequencing (Oxford Nanopore). Performance was comparable to three other extraction methods tested head-to-head, including the QIAamp DNA Mini Kit (QIAGEN), the MagaZorb DNA Mini-Prep Kit (Promega), and a phenol-chloroform extraction method. These findings establish IL-DEx as the first ionic liquid-based approach evaluated for DNA recovery from clinical urine samples, providing a fast, simple, and low-cost method suitable for integration into molecular workflows for UTI diagnostics across diverse laboratory and clinical settings. ImportanceUrinary tract infections (UTIs) are among the most common infections worldwide and a major driver of antibiotic use. Rapid and accurate diagnosis is critical to guide therapy, reduce inappropriate antibiotic prescriptions, and improve patient outcomes. While molecular diagnostics can drastically reduce time to identify uropathogens, their implementation remains constrained by upstream DNA extraction - a step that is often laborious, cost-intensive, or incompatible with rapid diagnostic workflows. We developed a fast, simple, and low-cost DNA extraction method (IL-DEx) that uses an ionic liquid and magnetic beads to recover bacterial DNA directly from urine. IL-DEx eliminates hazardous reagents and complex equipment while delivering performance comparable to established extraction kits. By streamlining this critical pre-analytical step, IL-DEx enables faster molecular diagnostics and broadens access to modern UTI testing. Its simplicity and robustness position it as a valuable tool for improving diagnostic speed, antimicrobial stewardship, and patient care across healthcare settings.

molecular biology↗
bioRxiv2020

Large scale metagenome assembly reveals novel animal-associated microbial genomes, biosynthetic gene clusters, and other genetic diversity

Large-scale metagenome assemblies of human microbiomes have produced a vast catalogue of previously unseen microbial genomes; however, comparatively few microbial genomes derive from other vertebrates. Here, we generated 5596 metagenome-assembled genomes from the gut metagenomes of 180 predominantly wild animal species representing 5 classes, in addition to 14 existing animal gut metagenome datasets. The MAGs comprised 1522 species-level genome bins (SGBs); most of which were novel at the species, genus, or family levels, and the majority were enriched in host versus environment metagenomes. Many traits distinguished SGBs enriched in host or environmental biomes, including the number of antimicrobial resistance genes. We identified 1986 diverse biosynthetic gene clusters; only 23 clustered with any MIBiG database references. Gene-based assembly revealed tremendous gene diversity, much of it host- or environment-specific. Our MAG and gene datasets greatly expand the microbial genome repertoire and provide a broad view of microbial adaptations to the vertebrate gut. ImportanceMicrobiome studies on a select few mammalian species (e.g., humans, mice, and cattle) have revealed a great deal of novel genomic diversity in the gut microbiome. However, little is known of the microbial diversity in the gut of other vertebrates. We studied the gut microbiome of a large set of mostly wild animal species consisting of mammals, birds, reptiles, amphibians, and fish. Unfortunately, we found that existing reference databases commonly used for metagenomic analyses failed to capture the microbiome diversity among vertebrates. To increase database representation, we applied advanced metagenome assembly methods to our animal gut data and to many public gut metagenome datasets that had not been used to obtain microbial genomes. Our resulting genome and gene cluster collections comprised a great deal of novel taxonomic and genomic diversity, which we extensively characterized. Our findings substantially expand what is known of microbial genomic diversity in the vertebrate gut.

microbiology↗