In groundbreaking research, scientists have found a new approach to understanding the virology of Monkeypox Virus (MPV) and developing advanced diagnostic tools, along with identifying novel therapeutic pathways. Recently renamed as Mpox virus, MPV has been declared a Public Health Emergency of International Concern (PHEIC) twice within the past three years due to its unexpected global spread. This has raised significant concerns, as the modes of transmission and symptoms remain poorly understood. A deeper comprehension of its virology, alongside the rapid advancement of effective diagnostic and therapeutic strategies, is critical.
MPV is a double-stranded DNA (dsDNA) virus, and the detection of its extracellular viral protein gene via polymerase chain reaction (PCR) is a well-established technique for identifying MPV in clinical specimens. Traditional PCR methods, which amplify dsDNA using fluorescent probes, are sensitive to DNA concentration but struggle to differentiate between specific and non-specific amplification products. In contrast, certain unique DNA structures known as noncanonical nucleic acid conformations deviate from the classic double-helix form. These unusual DNA structures, such as G-quadruplexes (GQs), hold potential as targets for small-molecule fluorescent probes, opening up possibilities for more reliable diagnostic assays.
G-quadruplexes (GQs) are distinctive noncanonical conformations found in guanine (G)-rich nucleic acid sequences, where four guanines interact through hydrogen bonding to form a planar G-tetrad. The stacking of multiple G-tetrads results in the formation of GQ structures. Researchers from the Jawaharlal Nehru Centre for Advanced Scientific Research (JNCASR), an autonomous institution under the Department of Science and Technology (DST), have identified and characterised four highly conserved GQ-forming DNA sequences within the MPV genome. Using a tailored fluorescent small-molecule probe, they successfully detected one specific GQ sequence, a breakthrough that enables precise detection of MPV. This research highlights the therapeutic potential of targeting such unusual nucleic acid structures.
The GQ sequences identified in the MPV genome are stable under physiological conditions, highly conserved, and absent in other pox viruses, pathogens, and the human genome. These attributes make GQ sequences valuable targets for developing diagnostic tools and therapeutic interventions. The fluorogenic molecular probe (BBJL), developed by Sumon Pratihar, Ramjayakumar Venkatesh, Mohamed Nabeel Mattath, and Thimmaiah Govindaraju, delivers over 250-fold enhancement in fluorescence output upon binding with MPV GQs (specifically MP2). This remarkable ability to selectively detect a conserved sequence in the MPV genome sets a new standard for diagnostic strategies targeting noncanonical nucleic acids.
The BBJL probe is non-fluorescent in the absence of its target GQ-DNA, marking the first practical demonstration of a GQ-targeted diagnostic approach for MPV detection. This innovation builds on their modular diagnostic platform, GQ-Reliable Conformation Polymorphism (GQ-RCP), originally developed for detecting SARS-CoV-2, and showcases its versatility by identifying MPV-specific GQs using a novel fluorogenic probe previously published in ACS Sensors. The sensitivity of this probe enables it to distinguish MPV-derived GQs from other GQs and DNA conformations found in the human genome, while the identified GQs can also serve as potential antiviral targets.
Further mapping of the MPV genome is ongoing to identify additional GQ targets for future therapeutic applications. This research significantly advances the development of GQ-based diagnostic platforms, and the identified GQs hold promise for broader exploration of their antiviral properties. Molecular probes like BBJL, with superior specificity for nucleic acid conformations, could address challenges in current amplification-based techniques by minimising false-positive results from non-specific amplifications. The discovery and characterisation of new GQ sequences in the MPV genome give the scientific community invaluable insights into MPV virology and avenues for novel diagnostic and therapeutic interventions.
Read More:
