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A new multiplex viral sequencing assay (mVseq) detects 20 transplant-relevant DNA viruses in clinical samples. This assay shows promise for identifying viral infections in transplant recipients, though clinical sensitivity varies.

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Area of Science:

  • Transplant infectious diseases
  • Molecular diagnostics
  • Virology

Background:

  • Viral infections pose significant risks, leading to complications and mortality in solid organ and hematopoietic cell transplant recipients.
  • Accurate and timely viral detection is crucial for managing transplant patients.

Purpose of the Study:

  • To develop and evaluate a novel multiplex viral sequencing assay (mVseq) for simultaneous detection of 20 transplant-relevant DNA viruses.
  • To assess the analytical and clinical performance of mVseq in various clinical specimens.

Main Methods:

  • Developed a single-tube multiplex PCR assay (mVseq) amplifying conserved viral regions from small clinical samples.
  • Sequenced amplified products using Illumina MiSeq and aligned reads to a target database.
  • Validated the assay using spiked samples and clinical specimens (plasma, BAL, CSF, urine, tissue) from transplant recipients.

Main Results:

  • mVseq demonstrated analytical sensitivity and dynamic range comparable to quantitative PCR (qPCR) in spike-in experiments.
  • Substantial agreement (92%) was observed between mVseq and single-target qPCR in clinical specimens, with an overall clinical sensitivity of 81%.
  • mVseq identified previously undetected BK virus, human herpesvirus-7, and Epstein-Barr virus infections in 12 patients.

Conclusions:

  • Factors like high host DNA and viral load differences in coinfections can impact mVseq's clinical sensitivity.
  • The mVseq assay is adaptable for detecting RNA viruses and other pathogens relevant to transplant recipients.
  • mVseq offers a flexible and scalable approach for comprehensive viral surveillance in immunocompromised individuals.