Performance of 16S rRNA Gene Next-Generation Sequencing and the Culture Method in the Detection of Bacteria in Clinical Specimens

Author:

Botan Alexandru1ORCID,Campisciano Giuseppina23,Zerbato Verena4ORCID,Di Bella Stefano3,Simonetti Omar4,Busetti Marina5ORCID,Toc Dan Alexandru6ORCID,Luzzati Roberto3ORCID,Comar Manola23

Affiliation:

1. Faculty of Medicine, “Iuliu Hațieganu” University of Medicine and Pharmacy, 400012 Cluj-Napoca, Romania

2. Laboratory of Advanced Microbiology Diagnosis and Translational Research, Institute for Maternal and Child Health IRCCS Burlo Garofolo, 34137 Trieste, Italy

3. Clinical Department of Medical, Surgical and Health Sciences, Trieste University, 34129 Trieste, Italy

4. Infectious Diseases Unit, Trieste University Hospital, 34125 Trieste, Italy

5. Microbiology Unit, Trieste University Hospital (ASUGI), 34125 Trieste, Italy

6. Department of Microbiology, “Iuliu Hațieganu” University of Medicine and Pharmacy, 400012 Cluj-Napoca, Romania

Abstract

Effective treatment of infectious diseases requires prompt and accurate bacterial identification and tailored antimicrobial treatments. Traditional culture methods are considered the gold standard, but their effectiveness diminishes for fastidious and hard-to-grow microorganisms. In recent years, molecular diagnostic tools such as 16S rRNA gene next-generation sequencing (16S NGS) have gained popularity in the field. We analysed data from samples submitted for 16S NGS between July 2022 and July 2023 at the Department of Advanced Translational Microbiology in Trieste, Italy. The study included samples submitted for both culture-based identification and 16S NGS. Conventional media were used for culture, and bacterial identification was performed using MALDI-TOF mass spectrometry. The V3 region of the 16S rRNA gene was sequenced using the Ion PGM platform. Among the 123 samples submitted, drainage fluids (38%) and blood (23%) were the most common, with requests predominantly from the Infectious Diseases (31.7%) and Orthopedic (21.13%) Units. In samples collected from patients with confirmed infections, 16S NGS demonstrated diagnostic utility in over 60% of cases, either by confirming culture results in 21% or providing enhanced detection in 40% of instances. Among the 71 patients who had received antibiotic therapies before sampling (mean 2.3 prior antibiotic days), pre-sampling antibiotic consumption did not significantly affect the sensitivity of 16S NGS. In routine microbiology laboratories, combining 16S NGS with culture method enhances the sensitivity of microbiological diagnostics, even when sampling is conducted during antibiotic therapy.

Publisher

MDPI AG

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