Usefulness of next-generation DNA sequencing for the diagnosis of urinary tract infection

Abstract

Acute urinary tract infection (UTI) is a highly common clinical condition. Although bacterial culture is the gold standard diagnostic test, false negative results may be possible, leading to the pathogen being unidentified. In recent years, bacterial DNA sequencing analysis has garnered much attention, but clinical studies are rare in Japan. In this study, we assessed the usefulness of next-generation DNA sequencing (NGS) analysis for acute UTI patients. We thus performed an observational, retrospective case series study. Urine and blood samples were collected from ten acute UTI patients, of whom four had also been diagnosed with urosepsis. Seven variable regions of bacterial 16S rRNA genes were amplified by PCR and then sequenced by IonPGM. The identified bacterial species were compared with those identified using the culture tests and the clinical parameters were analyzed. As a result, the NGS method effectively identified predominant culture-positive bacteria in urine samples. The urine NGS also detected several culture-negative species, which have been reported to be potentially pathogenic. Out of four urosepsis cases, three were pathogen-positive in blood NGS results, while two were pathogen-negative in blood culture. In one sepsis case, although blood culture was negative for Escherichia coli, this species was detected by blood NGS. For nonsepsis cases, however, blood NGS, as well as blood culture, was less effective in detecting bacterial signals. In conclusion, NGS is potentially useful for identifying pathogenic bacteria in urine from acute UTI patients but is less applicable in patients who do not meet clinical criteria for sepsis.

Urinary Microbiome: Yin and Yang of the Urinary Tract

“…The metagenomic analysis using Next Generation Sequencing (NGS) has facilitated the quantitative characterization of microbiomes, providing information on microbial populations and helping to discover uncultured microbes ( Wolfe and Brubaker, 2015 ; Ishihara et al., 2020 ). Two approaches could be taken as far as NGS techniques are concerned: amplicon sequencing and shotgun sequencing ( Figure 1 ).…”

“…Microbial identification based on molecular techniques has allowed a more accurate diagnosis of UTI, detecting bacteria and other uncultured uropathogenic microorganisms and avoiding false negative results. In this regard, a better treatment choice is achieved in patients with urinary tract infection ( Ishihara et al., 2020 ).…”

A New Gold Rush: A Review of Current and Developing Diagnostic Tools for Urinary Tract Infections

“…Finally, in an observational, retrospective case series by Ishihara et al, traditional urine/blood cultures and NGS were used study the urine and blood of 10 patients presenting to the emergency department with acute urinary tract infection, 4 of which were classified as having urosepsis [62]. Overall, 90% of patients were pathogen-positive on urine culture compared to 100% of patients identified as pathogen-positive by NGS.…”

Urinary microbiota; Which non‐ınvasive urine collection method should we use?

“…Next-generation sequencing in UTI diagnosis is a new and highly sensitive method that helps to detect clinically important species. 10,13 Even replacing urine culture with microbiota analysis in UTI diagnosis may be considered in the future. Traditionally, it was thought that all of the identified bacteria in the urinary tract are pathological.…”

“…[3][4][5][6][7][8][9] Recently, it was shown that 16S ribosomal RNA sequencing can detect bacteria residing in the urinary tract from the urinary samples with a high sensitivity and specifity, even if the patient has UTI symptoms or not. 2,10 Therefore, we have decided to investigate the reliability of a two-step urine collection method and the validity of the hypothesis which argues that the first voided urine reflects the urethra and the midstream urine reflects the bladder. Our goal was to establish the optimal noninvasive urine sample collection method for the microbiota studies, preferably without collecting two different samples.…”