Recent advances in molecular and serological assays for Monkeypox virus detection

This article offers a comprehensive review of the latest diagnostic tools developed for detecting Monkeypox virus (MPXV), especially in light of recent outbreaks. The authors focus on two major diagnostic categories: molecular assays, which detect viral genetic material, and serological assays, which identify antibodies or antigens associated with infection.

1. Molecular Diagnostics

The review begins with nucleic acid amplification tests (NAATs), which are central to accurate and early mpox diagnosis. Key methods include:

· Conventional PCR and real-time PCR (qPCR): These remain the gold standard due to high sensitivity and specificity. They can detect even low viral loads but require advanced lab infrastructure and skilled personnel.

· Isothermal amplification techniques: Methods such as LAMP (loop-mediated isothermal amplification) and RPA (recombinase polymerase amplification) offer rapid detection at constant temperatures, reducing the need for thermal cyclers and making them ideal for field or point-of-care settings.

· CRISPR-based diagnostics: Recently developed CRISPR-Cas systems (e.g., SHERLOCK, DETECTR) can detect MPXV with high specificity and speed. Though still in early stages, they hold promise for portable, low-cost diagnostics.

The article discusses the genomic targets used in these assays, like the F3L and B6R genes, and emphasizes the need for continued monitoring of viral mutations to ensure ongoing accuracy.

2. Serological Diagnostics

Serology plays a vital role in identifying past exposure to MPXV and in population surveillance. Covered techniques include:

· ELISA (Enzyme-Linked Immunosorbent Assay): Widely used to detect IgM and IgG antibodies. However, the article notes potential cross-reactivity with other orthopoxviruses (e.g., smallpox), which may limit specificity.

· Lateral Flow Assays (LFAs): These rapid, user-friendly tests can detect viral antigens or antibodies in minutes, but typically have lower sensitivity compared to ELISA or PCR.

· Neutralization assays: These provide functional data on immune protection but are labor-intensive and not widely used outside of research settings.

3. Emerging Technologies and Future Directions

The authors highlight the potential of:

·Synthetic biology approaches to design modular and programmable diagnostics.

·Multiplexed platforms that detect multiple pathogens simultaneously.

·Point-of-care integration, especially in resource-limited settings.

·Portable and smartphone-based readers to enhance accessibility and real-time data sharing.

They also stress the importance of assay validation, global standardization, and accessible testing infrastructure to manage future outbreaks effectively.

Conclusion

Overall, the article underscores that while PCR remains the most reliable method, rapid, decentralized diagnostics—especially isothermal and serological assays—are critical for outbreak control, particularly in underserved or remote areas. Ongoing innovation, data sharing, and investment in scalable diagnostic tools will be essential to strengthen global readiness against mpox and similar emerging infectious diseases.