Diagnostic Value of 16S Ribosomal RNA Gene Polymerase Chain Reaction/Sanger Sequencing in Clinical Practice

Madiha Fida, Sarwat Khalil, Omar Abu Saleh, Douglas W. Challener, Muhammad Rizwan Sohail, Joshua N. Yang, Bobbi S. Pritt, Audrey N. Schuetz, Robin Patel

Research output: Contribution to journalArticlepeer-review


Background. Accurate microbiologic diagnosis is important for appropriate management of infectious diseases. Sequencingbased molecular diagnostics are increasingly used for precision diagnosis of infections. However, their clinical utility is unclear. Methods. We conducted a retrospective analysis of specimens that underwent 16S ribosomal RNA (rRNA) gene polymerase chain reaction (PCR) followed by Sanger sequencing at our institution from April 2017 through March 2019. Results. A total of 566 specimens obtained from 460 patients were studied. Patients were considered clinically infected or noninfected based on final diagnosis and management. In 17% of patients, 16S rRNA PCR/sequencing was positive and in 5% of patients, this test led to an impact on clinical care. In comparison, bacterial cultures were positive in 21% of patients. Specimens with a positive Gram stain had 12 times greater odds of having a positive molecular result than those with a negative Gram stain (95% confidence interval for odds ratio, 5.2-31.4). Overall, PCR positivity was higher in cardiovascular specimens (37%) obtained from clinically infected patients, with bacterial cultures being more likely to be positive for musculoskeletal specimens (P < .001). 16S rRNA PCR/sequencing identified a probable pathogen in 10% culture-negative specimens. Conclusion. 16S rRNA PCR/sequencing can play a role in the diagnostic evaluation of patients with culture-negative infections, especially those with cardiovascular infections.

Original languageEnglish (US)
Pages (from-to)961-968
Number of pages8
JournalClinical Infectious Diseases
Issue number6
StatePublished - Sep 15 2021


  • 16S rRNA gene PCR
  • Sanger sequencing
  • bacterial infections
  • broad range bacterial PCR
  • molecular diagnostics

ASJC Scopus subject areas

  • Microbiology (medical)
  • Infectious Diseases


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