Evaluation of analytical performance of the STANDARD^TM M10 MPX/OPX assay for the simultaneous DNA detection and clade attribution of Monkeypox virus

ABSTRACT

Monkeypox virus (MPXV) infection confirmation needs reliable polymerase chain reaction (PCR) assays; in addition, viral clade attribution is a key factor in containment measures, considering a more severe syndrome in clade I and the possibility of simultaneous circulation. This study evaluates the performance of all-in-one STANDARD M10 MPX/OPX (SD BIOSENSOR, South Korea – M10). Frozen samples from 205 subjects were selected and stratified according to routine test results (RealStar® Orthopoxvirus PCR Kit 1.0, Altona DIAGNOTICS, Germany – RS; RS-1): in detail, 100 negative skin lesions (SL) and 200 positive samples at the variable stage of infection were analysed. Positive samples were retested with RS (RS-2). Positive and Negative Percent Agreements (PPA, NPA) were calculated. The median (IQR) C_t values of RS and M10 (OPXV target) assays were highly similar. The PPA of M10 compared to RS-1 was 89.5% considering system interpretation, and 96.0% when the operator classified results as positive if any target was detected; NPA was 100%. Comparing the RS-2 run and M10, an overall concordance of 95.3% between assays was found; however, considering operator interpretation, M10 returned more positive results than RS-2. The occurrence of False-Negative results was likely associated with the influence of thawing on low viral concentration; no False-Positive tests were observed. All samples collected at the time of Mpox diagnosis were positive and M10 correctly attributed the clade (West-Africa/II). The M10 MPX/OPX assay demonstrated high reliability in confirming MPXV infection and clade attribution.

KEYWORDS:

• Monkeypox virus
• clade determination
• methods comparison
• all-in-one RealTime-PCR
• Orthopoxvirus

Acknowledgements

The “PerBiocid” project of the Italian National Research Council (progetto@CNR2020: SAC.AD002.173.032) is gratefully acknowledged. We thank Doctor Francesca Garbarino from SD Biosensor Inc. (Republic of Korea), Roberta Gullotta and Davide Antoniani from Relab S.r.l. (Italy) for their technical support and test provision.

Disclosure statement

DM had sponsorship for attending the 50th AMCLI Congress from Relab S.r.l. (Italy) and the 33rd European Congress of Clinical Microbiology and Infectious Diseases from SD Biosensor Inc. (Republic of Korea). The other authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Ethics

The study was conducted in accordance with the Declaration of Helsinki, using appropriately frozen samples in the framework of a larger protocol approved by the institutional Ethics Committee “Comitato Etico Milano Area 1” (protocol number n. 2022/ST/124).

Authors’ contribution

AR, DMo, AG, SA, GA, GR, AC, and SN collected samples and clinical data, in the context of patient management. GG, CC, and FB performed assays. AR and FS managed the database. MRG managed the study funding. AM and DMi planned the study, interpreted the results, and wrote the paper draft and final version. All the authors reviewed and agreed on the final manuscript version.

Additional information

Funding

This research was funded by SD Biosensor Inc. (Republic of Korea). The sponsor had no role in study design, data analysis, or manuscript preparation.