Geneva/Bengaluru: The upsurge of mpox in the Democratic Republic of the Congo and its spread to neighbouring countries was first declared a public health emergency of international concern by the World Health Organization (WHO) Director-General on 14 August 2024.
While WHO D-G today again determined that the upsurge of mpox continues to constitute a public health emergency of international concern, scientists at Jawaharlal Nehru Centre For Advanced Scientific Research (JNCASR) in Bengaluru identified a new route to understand the virology of the mpox virus (MPV), develop diagnostic tools for the infection, and find a novel path with therapeutic implications.
MPV is a double-stranded DNA (dsDNA) virus. Polymerase chain reaction (PCR) detection of the extracellular viral protein gene is a widely established technique for identifying MPV in clinical specimens. Common detection approaches, including PCR, rely on the amplification of double-stranded DNA (dsDNA), which also employs fluorescent probes for quantifying amplification.
While these probes are sensitive to dsDNA concentration, they cannot differentiate between specific and non-specific amplification products. In contrast, specific sequences in DNA can fold into unique structures that deviate from the classical double helix, known as noncanonical nucleic acid conformations. Leveraging the potential of these unusual DNA structures as targets for small-molecule fluorescent probes may open new avenues for the development of highly reliable diagnostic assays.
G-quadruplex (GQ) is one such unusual noncanonical conformation observed in guanine (G)-rich nucleic acid sequences where four guanines interact through hydrogen bonding forming a planar G-tetrad plane, and the stacking of multiple G-tetrads results in GQ formation.
Scientists from JNCASR in Bengaluru claimed they have identified and characterised highly conserved GQ forming DNA sequences—a set of four, within the MPV genome, and specifically detected a specific GQ sequence using a tailored fluorescent small-molecule probe, enabling precise detection of MPV. The identification, characterization and targeting of unusual nucleic acid structures such as GQs have therapeutic implications.
These GQ sequences are stable under physiological conditions, highly conserved and are not present in other pox viruses, other pathogens, and the human genome. These characteristics make the GQ sequences valuable targets for the development of diagnostic tools and therapeutic intervention.
The fluorogenic molecular probe (BBJL) developed by Sumon Pratihar, Ramjayakumar Venkatesh, Mohamed Nabeel Mattath, and Thimmaiah Govindaraju also delivers more than 250-fold enhancement in fluorescence output upon binding with an MPV GQs (MP2). The ability of BBJL to selectively detect this highly conserved sequence in the MPV genome sets a precedent for the development of detection techniques targeting noncanonical nucleic acids.
The molecular probe is non-fluorescent in the absence of the target GQ-DNA and is the first practical demonstration of a GQ-targeted diagnostic strategy for the detection of Monkeypox virus (MPV). This represents the expansion of their modular diagnostic platform, GQ-targeted reliable conformation polymorphism (GQ-RCP), which was originally developed for the detection of SARS-CoV-2/COVID-19, by identifying MPV/Mpox-specific GQs and a novel fluorogenic probe which was earlier published in ACS Sensors
The sensitive probe can efficiently distinguish the target MPV-derived GQ from diverse other GQs, and other DNA conformations derived from the human genome. The GQs identified in the MPV genome can also serve as potential anti-viral targets.
Additional mapping of the MPV genome is being performed to identify potential GQ targets for future therapeutics. Consequently, this study augments the development of potential detection platforms based on GQ, and the GQs identified can be further investigated for their anti-viral properties.
Such molecular probes with superior conformation or sequences specific recognition of nucleic acids could alleviate the challenge in existing amplification-based techniques, in discriminating false-positive results arising from non-specific amplification. The identification and characterization of new GQ sequences in monkeypox virus (MPV) could help the broader scientific community seeking to understand the virology of MPV or develop diagnostic and therapeutic interventions.
