Deep sequencing approach for investigating infectious agents causing fever.

Area of Science:

• Medical Microbiology
• Genomics
• Infectious Diseases

Background:

• Acute undifferentiated fever (AUF) presents diagnostic challenges with diverse potential causes.
• While many AUF cases resolve spontaneously, some lead to severe illness, highlighting the need for improved diagnostic strategies.

Purpose of the Study:

• To evaluate the effectiveness of deep sequencing for identifying etiological agents in patients with AUF.
• To compare deep sequencing results with conventional diagnostic methods.

Main Methods:

• DNA and RNA were extracted from plasma/serum samples of AUF patients and controls.
• Deep sequencing was performed using an Illumina HiSeq 2000 platform after bulk amplification and cDNA synthesis.
• Bioinformatics analysis was conducted using Kraken and CLC assembly-alignment pipelines.

Main Results:

• Deep sequencing identified etiological agents in ten previously undiagnosed AUF patients, including Escherichia coli and scrub typhus.
• Conventional methods and deep sequencing results aligned in 87.5% of control samples for Kraken and 25% for CLC.
• Specific bacteria (Achromobacter xylosoxidans, Alteromonas macleodii) and Enterobacteria phage were common across samples.

Conclusions:

• Deep sequencing is a valuable approach for identifying causative agents of AUF and exploring blood microbial diversity.
• Current limitations for clinical application include high sequencing input needs and complex data analysis.