Multiplex Nucleic Acid Suspension Bead Array Detects Filoviruses

By LabMedica International staff writers
Posted on 29 Apr 2015

Filoviruses (FVs) are ribonucleic acid (RNA) viruses that belong to the family Filoviridae, which includes zoonotic pathogens of the three genera Ebolavirus (EV), Marburgvirus (MV), and Cuevavirus (CV).

Multiplex suspension bead array systems that allow fast and reliable detection of reverse transcriptase polymerase chain reaction (RT-PCR) amplified filovirus genomes and enabled subtyping of Ebolavirus species and Marburgvirus strains has been described.

Image: The SuperScript III One-Step reverse transcriptase polymerase chain reaction (RT-PCR) kit with Platinum Taq (Photo courtesy of Invitrogen).

Scientists at the Public Health Agency of Sweden (Solna, Sweden) and their colleagues set up a multiplex suspension bead array system, by designing two different RT-PCRs that could amplify all known EV and MV species, respectively, based on whole-genome sequences of FVs, including Reston ebolavirus (RESTV), and Bundibugyo ebolavirus (BDBV). The RT-PCRs were performed using a SuperScript III One-Step RT-PCR kit with Platinum Taq (Invitrogen, Carlsbad CA, USA).

The RT-PCR amplicons were also sequenced to confirm specificity. In parallel, different sets of magnetic Bio-Plex Pro COOH beads (Bio-Rad Laboratories, Hercules, CA, USA) were coupled with oligonucleotides specific for each EV species and MV strain. The suspension bead arrays were also tested on human sera spiked with Lassa virus, Dengue virus serotypes 1 to 4, Crimean-Congo hemorrhagic fever virus, Rift Valley fever virus, and 10 FV-negative human sera. The results did not reveal any cross-reactivity which demonstrated the utility of the microsphere-based assay as a rapid and accurate diagnostic tool for confirming suspected FV infections.

The EV system can distinguish between three EV species and the MV system can distinguish between four MV strains. Based on the sequence information from the current Ebola outbreak in West Africa, the EBOV probe described in the study will also detect that particular strain. The multiplex suspension bead array system developed can easily be adapted to emerging new strains by including additional beads. The study was published in the April 2015 issue of the Journal of Clinical Microbiology.

Related Links:
Public Health Agency of Sweden
Invitrogen
Bio-Rad Laboratories