Monkeypox virus outbreak worldwide, what should we do?——Yeasen provides enzyme raw materials for virus detection

The World Health Organization says 92 confirmed and 28 suspected cases have been reported from 12 countries around the world infected with the Monkeypox virus since May 13. Available information suggests that human-to-human transmission is occurring among people who have had close physical contact with symptomatic cases. So what is monkeypox? How does it spread? What should we do about the spread of monkeypox?

 

1. What is monkeypox?
2. How is monkeypox spread and how can it be prevented?
3. Clinical diagnosis of monkeypox
4. Yeasen provides enzyme raw materials for virus detection

 

1. What is monkeypox?

Monkeypox is a viral zoonotic disease in which pathogens are transmitted from animals to humans. Early symptoms of monkeypox infection include fever, headache, muscle pain, swollen lymph nodes, chills, fatigue, etc., and a healing rash resembling blisters. Some patients may have a rash first or only have rash symptoms. The incubation period of monkeypox (the interval from infection to onset of symptoms) is usually 7 to 14 days and maybe 5 to 21 days. Monkeypox is usually a self-limiting disease, with symptoms usually lasting 2 to 4 weeks after onset. Severe cases of monkeypox are more common in children and are related to the patient's health, exposure to the virus, and complications. Underlying immunodeficiency conditions may lead to more severe outcomes. The complications of monkeypox may include secondary infection, bronchopneumonia, sepsis, encephalitis, and corneal infection. The case fatality rate of monkeypox in the general population is 0% to 11%, and the case fatality rate in children is higher, about 3% to 6%.
Its pathogen, monkeypox virus, is a DNA (deoxyribonucleic acid) virus, which belongs to the genus Orthopoxvirus of the family Poxviridae. It is a “close relative” to the smallpox virus that has ravaged thousands of years in human history. Genome sequencing identified two phylogenetically distinct monkeypox virus clades, the Congo Basin (Central Africa) clade, and the West African clade. The former causes more severe disease, higher mortality, and more frequent human-to-human transmission. Monkeypox virus has the typical form of orthodox virus, the shape is the rounded brick shape or oval shape, and the size is 200-400nm. The exterior is enveloped by a lipoprotein membrane, with two protein-containing side bodies in the middle, and a thick-membrane core containing the double-stranded DNA gene body. The monkeypox virus genome is double-stranded DNA, about 197 kb in length, and contains an inverted terminal repeat sequence in the same but opposite direction at the end of the genome. Monkeypox virus contains 190 open reading frames, 4 of which are located in inverted terminal repeats.
By May 21, three research groups from Portugal, Belgium, and the United States have released the preliminary results of the genome sequences of the viruses carried by infected persons in the recent Monkeypox outbreaks in many places around the world. Belgium has become the first country in the world to require Monkeypox cases to self-isolate for 21 days.
Monkeypox is a viral zoonotic disease that can be transmitted from animals to humans through close contact. Although it can be self-healing, it makes people "disabled". The premise of effective prevention and intervention is detection.

Figure 1. Monkeypox virus

Credit: UK Health Security Agency/Science Photo Library
Source: https://www.nature.com/articles/d41586-022-01421-8

 

2. How is monkeypox spread and how can it be prevented?

Monkeypox virus can be transmitted in certain rodents in Africa. Evidence of monkeypox virus infection has been found in many animals in Africa, and the natural host for monkeypox has not been identified, but rodents are the most likely. Eating undercooked meat and other animal products from infected animals is a possible risk factor. People living in or near forested areas may have indirect or direct contact with infected animals. Human-animal transmission routes include direct contact with body fluids or diseased parts of infected animals, eating meat from infected animals, being scratched and bitten by animals, and receiving contaminated objects. Human-to-human transmission is mainly through respiratory droplets or body fluids. Human-to-human transmission can be caused by close contact with respiratory secretions, skin lesions, or recently contaminated objects of an infected person. Transmission through respiratory droplets usually requires prolonged face-to-face contact,
The main prevention strategies for monkeypox are raising awareness of risk factors and educating people about what can be done to reduce exposure to the virus. First, surveillance and rapid detection of new cases are important to contain monkeypox transmission and reduce the risk of human-to-human transmission. Secondly, it is necessary to reduce the risk of human-to-animal transmission and avoid unprotected contact with wild animals, especially those that are sick or dead. Also, all foods containing animal meat or organs must be thoroughly cooked before consumption. Monkeypox can also be prevented by restricting trade in animals, and captive animals that may be infected with monkeypox should be separated from other animals and placed in quarantine. Any animals in contact with potentially infected animals should also be quarantined, given standard precautions, and observed for monkeypox symptoms within 30 days.

 

3. Clinical diagnosis of monkeypox

The best diagnostic samples for monkeypox are vesicles and pustular fluid in skin lesions, and dry crusts. At the same time, the diagnostic samples should be stored in dry and sterile test tubes and kept at a low temperature. Other rash diseases, such as chickenpox, syphilis, measles, etc., and other drug-related allergic rashes also need to be considered in the clinical diagnosis. However, premorbid lymphadenopathy is the only clinical feature that distinguishes monkeypox from varicella/smallpox.

3.1 Real-time fluorescent quantitative PCR

Monkeypox virus DNA can be collected by vigorously swabbing the lesion or by taking an oropharyngeal swab to ensure sufficient viral DNA is collected. The monkeypox virus detection is based on the nucleic acid amplification test (NAAT). Use real-time or conventional PCR to detect unique sequences of viral DNA. PCR can be used alone or in combination with sequencing.
For routine detection of monkeypox virus DNA in monkeypox virus-infected cell cultures, PCR or real-time fluorescent quantitative PCR is recommended in a biosafety level 3 facility. Real-time fluorescent quantitative PCR was used to detect the outer envelope protein gene, DNA polymerase gene, a conserved region of EOL, DNA-dependent RNA polymerase subunit 18, rpo18, and F3L gene.

3.2 Restriction endonuclease fragment length polymorphism (RFLP)

PCR-amplified gene or gene fragment RFLP can also be used for the detection of monkeypox virus DNA. However, RFLP requires virus culture, which takes a long time. The RFLP of the PCR product also needs to be digested and then subjected to gel electrophoresis. Therefore, it may not be the best detection method in the clinical environment with high requirements for rapidity, sensitivity, and specificity.

3.3 High-throughput sequencing technology (NGS)

Whole-genome sequencing using NGS technology remains the gold standard for the characterization of monkeypox virus from other orthopoxviruses. However, the cost of this technology is high, and the downstream processing of sequencing data requires large computing power. Therefore, in resource-poor countries, NGS may not be the appropriate method.

 

4. Yeasen provides enzyme raw materials for virus detection

Monkeypox virus is a DNA virus with higher stability in vitro, no need for reverse transcription, and lower requirements for the reaction system of the kit. RNA viruses are prone to mutation and spread faster. To avoid missed detection, the development of relevant detection reagents needs to do the following two points. The first is to look for relatively conserved regions of virus specificity that make them less susceptible to viral mutations. Secondly, the design of primers and probes for detection reagents needs to cover different subtypes, which is more difficult. Relatively speaking, as a DNA virus, the monkeypox virus does not mutate frequently, and its conserved regions are easy to find. And detection reagents need to cover fewer subtypes in primer and probe design, so the development of reagents is theoretically easier. The upstream of the IVD industry chain includes biological raw materials, chemical raw materials, packaging materials, electronic devices, software, and other fields. Yeasen provides biological raw materials for the IVD industry chain of monkeypox.
In China, many of Yeasen's PCR detection technology partners have designed PCR detection kits. At present, several listed companies have announced that they have stockpiled Monkeypox virus nucleic acid detection kits. Yeasen can provide core raw materials and overall solutions for virus detection. Yeasen provides a complete solution for DNA pathogen detection, including sample extraction, detection premix, and detection of raw materials. At the same time, Yeasen provides customized development services for reagent raw materials (IVD RDC), which can help customers quickly develop Monkeypox virus detection kits.
At the same time, Yeasen Biotechnology provides reagents and raw material custom development services (IVD RDC). According to customer goals, help customers customize enzymes, customize premix formulations (choose dyes, optimal concentrations of any component, freeze-drying system, rapid procedures), customize amplification primer probe pools, customized production, etc. Help customers market molecular diagnostic kits quickly and provide complete certification data support. The products provided by Yeasen are as follows.

Table 1. Related products

Product type	Product Name	SKU
Assay master mix	Hieff Unicon™ TaqMan Multiplex qPCR Master Mix (UDG plus)	13171ES
	Hieff Unicon™ Universal TaqMan multiplex qPCR master mix	11211ES
Single enzyme	Hieff UNICON™ HotStart Direct Taq DNA Polymerase	10717ES
	Hieff™ Double-Block anti-Taq DNA Polymerase Antibody	31303ES
	Hieff™ Bst plus DNA Polymerase	14402ES
	Uracil DNA Glycosylase (UDG), heat-labile	10303ES

 

Reference

[1] Yinka-Ogunleye A, Aruna O, Dalhat M, et al. Outbreak of human monkeypox in Nigeria in 2017-18: a clinical and epidemiological report. Lancet Infect Dis. 2019,19(8):872-879. (IF71.421)
[2] Erika Hammarlund, Anindya Dasgupta, Clemencia Pinilla, et al. Monkeypox virus evades antiviral CD4+and CD8+T cell responses by suppressing cognate T cell activation[J]. Proc Natl Acad Sci USA, 2008, 105(38): 14567-72.(IF12.779)
[3] Emmanuel Alakunle, Ugo Moens, Godwin Nchind, et al.Monkeypox Virus in Nigeria: Infection Biology, Epidemiology, and Evolution[J]. Viruses, 2020, 12(11):1257. (IF5.818)
[4] Sarah Keasey, Christine Pugh, Alexander Tikhonov, et al. Proteomic Basis of the Antibody Response to Monkeypox Virus Infection Examined in Cynomolgus Macaques and a Comparison to Human Smallpox Vaccination[J]. Plos one, 2010, 5(12): e15547.

 

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