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 R 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. Sequencing-based 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 : an official publication of the Infectious Diseases Society of America
Issue number6
StatePublished - Sep 15 2021


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

ASJC Scopus subject areas

  • Microbiology (medical)
  • Infectious Diseases


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