Multiplex screening for blood-borne viral, bacterial, and protozoan parasites using an OpenArray platform

Area of Science:

• Molecular biology
• Infectious disease diagnostics
• Biotechnology

Background:

• Nucleic acid tests enhance blood product safety but require simultaneous multi-pathogen detection due to emerging threats.
• Existing methods often lack the capacity for broad, simultaneous pathogen screening.

Purpose of the Study:

• To develop and validate a multiplex real-time PCR assay for simultaneous detection of nine blood-borne pathogens (BBPs).
• To evaluate the performance of the OpenArray nanofluidic platform combined with TaqMan chemistry for infectious disease testing.

Main Methods:

• Developed a panel of TaqMan assays targeting four viruses, two bacteria, and three protozoan parasites.
• Utilized the OpenArray nanofluidic real-time PCR platform for spatial multiplexing.
• Tested assay specificity and analytical sensitivity using purified pathogens and spiked human blood/plasma samples.

Main Results:

• Achieved limits of detection comparable to existing platforms (10 to 10,000 copies/mL) for most targets.
• Demonstrated no false-positive results in spiked human plasma or blood samples.
• Successfully identified pathogens at concentrations as low as 10 cells/mL (Trypanosoma cruzi) to 10,000 cells/mL (Escherichia coli).

Conclusions:

• The BBP OpenArray platform combined with TaqMan chemistry offers a viable multiplex real-time PCR method for pathogen detection.
• This approach represents a proof of concept for a next-generation platform for infectious disease screening in blood.
• The developed assay shows potential for improving the safety and efficiency of blood product screening.