Mpox disease, diagnosis, and point of care platforms

Abstract

Human Mpox disease (MPX) is an endemic zoonotic disease that develops when patients are infected with the Mpox virus (MPXV). MPXV shares a high level of genetic similarity to other poxviruses and the clinical presentation of MPX is similar to other poxvirus infections which can result in a delay in diagnosis. In addition, the MPXV virus is phylogenetically divided into two different clades which affects the severity of disease. In recent years, there has been an unusual worldwide spread of MPXV, leading to a global public health problem. The most important step in the fight against MPX is rapid, highly specific, and accurate diagnosis. Following the rapid spread of disease in recent years, efforts to develop diagnostic tests have gained momentum. Here, MPX, MPX epidemiology, and MPX diagnostic tests are discussed. Furthermore, biochemical diagnostic tests, molecular diagnostic tests and their development, and point-of-care (PoC) diagnostic applications are reviewed. Molecular diagnostic technologies such as polymerase chain reaction, recombinase polymerase amplification, and loop-mediated isothermal amplification methods that detect MPX are evaluated. Additionally, next-generation combined molecular techniques and their importance in PoC transition are explored.

Keywords: Mpox (MPX); loop‐mediated isothermal amplification (LAMP); point‐of‐care (PoC) applications; polymerase chain reaction (PCR); recombinase polymerase amplification (RPA).

FIGURE 1

Chart representation of World Health Organization (WHO)‐verified Mpox data from September 2022 to January 2024. Data presented in the WHO January 2024 report were obtained from the CSV. (a) According to January 2024 data, the total number of cases reported in WHO regions are expressed on the graph. (b) Total number of confirmed deaths across WHO regions, based on January 2024 data. (c) Case numbers of the top 10 countries reporting the most cases to WHO and the other 107 countries from September 2022 to January 2024.

FIGURE 2

Schematic representation of Mpox transmission pattern. Transmission is divided into three: animal to animal, animal to human, and human to human. Human‐to‐human transmission is divided into two: vertical (transmission from infected mother to baby) and horizontal transmission. Transmission from animal to human occurs through direct contact with infected rodents, or through animal bites and scratches. Horizontal transmission from person to person occurs through sexual contact, close contact with respiratory secretions of the infected individual, contact with contaminated objects, and consumption of infected animal meat.

FIGURE 3

(a) Schematic representation of VF‐RPA MPXV detection platform illustrating the (i) RPA reaction; (ii) VF‐probe hybridization, and (iii) result output which occurs as a result of color development following the detection of nucleic acids. Adapted from Reference [63] under CC BY‐NC‐ND 4.0 license. (b) 1‐h workflow for the MPXV loop‐mediated isothermal amplification lateral flow biosensor (MPXV‐LAMP‐LFB) including the nucleic acid extraction, amplification, and result output steps. (c) Schematic representation of the MPXV‐LAMP‐LFB platform featuring the working principles of LAMP reactions (Clade I MPXV and Clade II MPXV) which leads to the formation of digoxin/biotin and FAM/biotin‐labeled amplicons post‐amplification and (d) the LFB and result output in which Clades I and II MPXV strains resulting in the T1 and T2 lines, respectively. CL acts as a control and appears if the result is negative and if one or both strains are present. Images (b–d) have been adapted from Reference [66] under Creative Commons CC‐BY license. LAMP, loop‐mediated isothermal amplification; LFB, lateral flow biosensor; MPXV, Mpox virus; VF‐RPA, vertical flow strip recombinase polymerase amplification.

FIGURE 4

Schematic representation of next‐generation CRISPR combined detection technology. CRISPR‐based detection applications are basically divided into two; amplification‐free CRISPR and CRISPR with pre‐amplification. Traditional amplification methods such as PCR, isothermal amplification techniques such as RPA, and LAMP increase the assay's sensitivity by amplifying the viral product. Afterwards, viral nucleic acid is detected using CRISPR‐Cas systems 12 or 13. Finally, fluorescence‐readout and lateral‐flow test methods are used to observe and read the detection results. CRISPR, clustered regularly interspaced short palindromic repeats; LAMP, loop‐mediated isothermal amplification; PCR, polymerase chain reaction; RPA, recombinase polymerase amplification.

FIGURE 5

Portable CRISPR plasmon resonance‐based fiber type (CRISPR‐SPR‐FT) biosensing platform. (a) Schematic representation of the design. The portable device is composed of a biosensor. AuNPs with partial complementary DNAs and ssDNA reporters were used to detect MPXV. Cas12a‐crRNA is activated when target DNA is encountered, which leads to the trans‐cleavage of reporters at key sites, thereby leading to spectral signal transmission. A fiber‐based system is used to record signals in real‐time. (b) Photograph of the 125‐μm fiber tip of disposable SPR‐based biosensor. (c) SPR stimulation is depicted. (d) MMF and CF structures are illustrated. Adapted from Reference [70] under CC‐BY‐NC‐ND 4.0 license. CF, coreless fiber; CRISPR, clustered regularly interspaced short palindromic repeats; MMF, multimodal fiber; MPXV, Mpox virus; SPR, surface plasmon resonance.