Abstract
BackgroundReliable high-throughput microbial pathogen identification in human urine samples is crucial for patients with cystitis symptoms. Currently employed methods are time-consuming and could lead to unnecessary or inadequate antibiotic treatment. Purpose of this study was to assess the potential of mass spectrometry for uropathogen identification from a native urine sample.MethodsIn total, 16 urine samples having more than 105 CFU/mL were collected from clinical outpatients. These samples were analysed using standard urine culture methods, followed by 16S rRNA gene sequencing serving as control and here described culture-independent MALDI-TOF/TOF MS method being tested.ResultsHere we present advantages and disadvantages of bottom-up proteomics, using MALDI-TOF/TOF tandem mass spectrometry, for culture-independent identification of uropathogens (e.g. directly from urine samples). The direct approach provided reliable identification of bacteria at the genus level in monobacterial samples. Taxonomic identifications obtained by proteomics were compared both to standard urine culture test used in clinics and genomic test based on 16S rRNA sequencing.ConclusionsOur findings indicate that mass spectrometry has great potential as a reliable high-throughput tool for microbial pathogen identification in human urine samples. In this case, the MALDI-TOF/TOF, was used as an analytical tool for the determination of bacteria in urine samples, and the results obtained emphasize high importance of storage conditions and sample preparation method impacting reliability of MS2 data analysis. The proposed method is simple enough to be utilized in existing clinical settings and is highly suitable for suspected single organism infectious etiologies. Further research is required in order to identify pathogens in polymicrobial urine samples.
Highlights
Reliable high-throughput microbial pathogen identification in human urine samples is crucial for patients with cystitis symptoms
60–80% of all uncomplicated bacterial Urinary tract infection (UTI) are caused by E. coli
Urine culture test The microorganisms were identified by routine microbiology methods [42]
Summary
Reliable high-throughput microbial pathogen identification in human urine samples is crucial for patients with cystitis symptoms. Methods: In total, 16 urine samples having more than 105 CFU/mL were collected from clinical outpatients. In order to identify microorganisms in clinical microbiology laboratories, most used methods are microbiological techniques which are still based on cultivation on different culture media [9]. Despite advances in genomics and proteomics, urine culture method is still the golden standard for the diagnosis of UTIs. Urine samples containing more than 105 CFU/mL of a single microbial species usually indicate clinical relevance. The first limitation is the time required for the cultivation of microorganisms and subsequent identification [10]. The second limitation is the requirement for fresh urine samples. Some of these limitations may result in overall negative urine cultures in up to 80% of cases, in many microbiology laboratories [12]. A wide variety of sampling methods and inappropriate specimen transport are major cause of preanalytical errors [13]
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