Mismatches in the primer and probe sequences of current Monkeypox virus diagnostic assays require rectification to improve detection accuracy UTHealthSPH Monkeypox MonkeypoxVirus Assay DiagnosticAssay
By Neha MathurOct 11 2022Reviewed by Aimee Molineux In a recent study posted to the medRxiv* preprint server, researchers analyzed the primer and probe sequences of the United States Centers for Disease Control and Prevention recommended monkeypox virus generic real-time polymerase chain reaction assay.
The issue is that the primers and probes used in these generic PCR assays are based on MPXV strains circulating almost a decade ago between 2002 and 2009. Given the rapid rate at which all DNA viruses evolve, the regions targeted by those oligos used for MPXV detection must have undergone substantial mutations.
Study findings They used 1,730 complete MPXV genomes retrieved during the 2022 worldwide outbreak. Aligning the three oligo sequences and their reverse complements from the MPXV generic assays yielded MPXV generic forward sequences with 100% identity to four genomes and generic reverse primer matching to 1.73% of the genomes. Further, the researchers found 31 sequence mismatches in 99.08% and 97.46% of MPXV generic forward sequences and 32 generic reverse primers , respectively.
Further, the study results showed that genetic variations in the primer-probe regions of MPXV genomes could indicate a temporal and spatial emergence pattern of monkeypox disease. Three assays, MPV_F3L, MPV_G2R_WA, 283, and OPV_F8L, showed the highest matching score of over >99% to the global MPXV genome database in 2022. However, the choice of the assay also varies with the sample type.
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