A Study Project Headed by Fu Gao Sheds A Beam of Light on the Abysmal Darkness of the CCP‘s Ambitious Pursuit of Unrestricted Bioweapons

(a complete translation of the project plan is appended herein for reference)

In the wake of the emergence of the great work “Non-natural Origin of SARS and Genetic Weapons with Man-made New Species of Human Viruses (《非典非自然起源和人制人新种病毒基因武器》by Dezhou Xu, Professor and Director of Army Epidemiology Teaching and Research Office, Fourth Military Medical University, and Feng Li, Deputy Director of Epidemic Prevention Bureau, General Logistics Department, hereinafter referred to as the “Bioweapon Guide”) into the spotlight of the public, Lude introduced another enlightening document “Study of the Molecular Mechanisms for Cross-Species Infection, Transmission and Pathogenesis of Major Viruses” in his recent broadcasts.

News about this project can be found on the website of the Chinese Academy of Sciences

(Translation of the title of the news: Kickoff of the Project “Study of the Molecular Mechanisms for Cross-Species Infection, Transmission and Pathogenesis of Major Viruses” )

http://www.cas.cn/xw/yxdt/201104/t20110413_3114268.shtml

Please find the complete translation of this document attached as Appendix I to this article for details about this five-year study. However, I have to admit that I am not a professional in virology or any related fields, so errors and misunderstandings are unavoidable, for which I wish to have your tolerance and understanding.

This project document opens a window for us to peep into the CCP’s diabolic endeavour to develop the so-called “contemporary genetic weapons”, which, according to the Bioweapon Guide, are defined as “new species or pathogenic genes which are artificially created by modifying the genes of pathogens and their products and animals and plants in nature and even genes of humans by use of the ever-developing biological technology and laboratorial means in combination with animal group adaptation experiments and which are then weaponised to have the power to pose danger to the human race or biosphere or specific groups (individuals).“ (P99 of the Bioweapon Guide)

A most important characteristic of such weapons is “invisibility”, “which makes it extremely difficult to identify the methods, materials, locations, times, etc. of their release so that there is no way to uncover the discharge process, the reasons for the discharge, and who discharges, researches and develops, and orchestrates these weapons. Eventually, the incident will be smothered up” (P99 of the Bioweapon Guide)- and therefore, the culprit can get away with the secret attack unscathed.

No doubt, COVID-19 is a perfect contemporary genetic weapon in every sense. Do you feel a chill running down your spine?

Now let me give you some nuts and bolts of the study.

This 5-year study was launched in January 2011 under the banner of providing theoretical supports for the prevention and treatment of emerging human infectious diseases, with novel Coronavirus, nipah virus or similar paramyxoviruses, Hantaan virus (aka hantavirus), influenza virus (esp. highly pathogenic avian influenza virus), rabies virus, encephalitis B virus (aka Japanese encephalitis virus), and dengue virus singled out as the major objects of study, West Nile virus is also mentioned for development of human MHC transgenic mice.

In the 5-years period, oodles of samples of bats, rodents, insectivores, animals in the wild, zoos, and farms all over the country were collected for discovery and isolation of novel viruses. The subsequent study was particularly focused on cross-species infection and transmission of the viruses, understanding the molecular mechanisms for cross-species infection, pathogenesis, and neurotropism of the viruses.

The study was in part based on animal models with human HLA-A2/DR1 and HLA-A11/DR1 transgenic mice, which shows that the study aimed to understand the pathogenic effects of the modified viruses on human bodies.

The project aimed to build human coronavirus banks, virus strain banks, gene banks, and natural host animal virus resource banks.

Although the study was carried out under a noble pretext, there are quite a few suspicious things that demand questioning.

Firstly, why did the CCP squander so much money, manpower, and resources to scout every corner of the country for novel viruses, particularly, the eight types of viruses mentioned? If those viruses are left alone, the chance of them causing infection of human beings or even epidemic could be negligibly small while the efforts to find them, bring them to laboratories, study them, and store them in the virus banks substantially increase the relevant risks.

Secondly, why were the eight types of viruses (novel Coronavirus, nipah virus or similar paramyxoviruses, Hantaan virus, influenza virus, rabies virus, encephalitis B virus, dengue virus, and West Nile virus) particularly selected for the study, especially given the fact that the rabies vaccine was invented in 1885 and Japanese encephalitis vaccine was first available in China in the 1960s?

Thirdly, why did the researchers put so much emphasis on cross-species infection and transmission and show such great zeal and commitment in furthering the study to the molecular level? They seemed to be trying to crack all the genetic secrets about the underlying mechanisms. We cannot help but ask whether all these efforts were to prevent or facilitate the jumping of the animal viruses to humans.

Finally, why did the researchers show so much interest in the neurotropism (which means the quality or state of being neurotropic and “a neurotropic virus is a virus that is capable of infecting nerve cells”) of the viruses? We know that the blood–brain barrier helps protect our central nervous system against most pathogens. However, some viruses, such nipah virus, influenza virus, West Nile virus, rabies virus, and encephalitis B virus are all able to break through this barrier and cause severe damages to the brain. Is it a coincidence that a majority of the viruses involved in this study share this powerful trait?

By comparing the symptoms of SARS and COVID-19 (SARS-2), I notice that in the case of SARS, “rash and neurologic or gastrointestinal findings are absent, although a few patients have reported diarrhoea during the early febrile stage” whereas in the case of COVID-19, headache, loss of smell or taste, nausea or vomitting, diarrhea, sudden confusion, strokes, post-recovery cognitive impairment are reported, which show that SARS-2 is obviously upgraded with a new function: neurotropism. Is this another coincidence of nature or the fruit of many years of strenuous and persistent hard work of the numerous virological researchers under the leadership of the CCP?

If we believe that this study is part of the CCP’s ambitious pursuit of world hegemony through unrestricted bioweapons and that COVID-19 is a perfect bioweapon discharged by the CCP as a means to cope with the predicament caused by the political and economic pressure from the US, we are forced to grapple with quite a few disturbing problems.

Firstly, the 5-year study was finished more than 5 years ago, which begs the question of how much progress the CCP has made in its virological study and bioweapon project in the past 5 years?

Secondly, according to Dr. Limeng Yan’s report, the bat coronaviruses ZC45 and ZXC21 both are 89% identical to SARS-CoV-2 on the nucleotide level, which means 11% of the genome of either of these bat coronaviruses has be modified. Then how many of the CCP’s research results have been incorporated into the genome of SARS-CoV-2? How much do we need to know but will never be told by the CCP about the virus?

Thirdly, as a number of military laboratories, including quite a few BSL3 or even BSL4 labs, in China have been doing similar studies for years, what else do they have in their arsenals? If the CCP is cornered to the extent of desperation, will it threaten the world with these invisible weapons? As the Belt and Road Initiative has made inroads into so many countries and regions, is it possible the CCP has deployed its bioweapons in many of these countries? If yes, then the CCP will be able to hold not only the Chinese people but also the peoples of these countries’ hostage in their final confrontation with the alliance of the victimized countries.

Finally, now the CCP is busy churning out COVID-19 vaccines, exporting them to Europe, South and Southeast Asia, Africa, Middle East, Latin and South America for massive amounts of US dollars. Do you believe the devil determined to conquer the world with its secret weapon will provide the world with ironclad shields to ward off its attack? I tend to believe the CCP vaccines are bioweapons in disguise, which are not safe in themselves and may cause unexpected acute or even fatal reactions to the new mutant strains that are biding their times or other common viruses, such as the influenza virus. If this hypothetical catastrophe comes true, god knows how many lives will be harvested. This imagination calls up Lenin’s quote ‘the Capitalists will sell us the rope with which we will hang them.’ That is how preposterous the world has become.

(The ideas elaborated in this article are in large part based on Lude’s broadcast. I would like to express my thanks to Lude and other commentators.)

Appendix I. Translation of the study project document

URL for downloading the project document (in Chinese): https://max.book118.com/html/2017/0108/81228787.shtm

2011CB504700 – Study of the Molecular Mechanisms for Cross-Species Infection, Transmission and Pathogenesis of Major Viruses

Project Name: Study of the Molecular Mechanisms for Cross-Species Infection, Transmission and Pathogenesis of Major Viruses

Principal Scientist: Fu Gao          Institute of Microbiology, Chinese Academy of Sciences Time frame: from January 2011 to August 2015

Affiliated to: Chinese Academy of Sciences

II. Expected objectives

Overall goal

In response to the national strategic demands for prevention and treatment of emerging human infectious diseases and in view of the current occurrences and epidemics of major diseases in our country, this project focuses on the forefront of international virology research, chooses as its major study objects such typical viruses as novel Coronavirus, encephalitis B virus, dengue virus, rabies virus, influenza virus, etc. that pose severe danger or potential hazards to our country, uses scientific and technological means of virology, molecular biology, cellular biology, structural biology, and other modern life sciences to reveal the ecological distribution characteristics of major viruses, especially novel viruses, in their animal hosts; the project also analyses the genetic mutations and evolutionary laws of the viruses, elucidates the molecular basis of cross-species infection and pathogenesis of the viruses, makes breakthroughs in the basic theoretical research of cross-species transmission of the viruses, further brings our studies in this research field to the international advanced level, and provides scientific supports for the prevention and treatment of emerging infectious diseases in our country. Through implementation of this project, not only can we foster a batch of top-notch international innovative talents and academic mainstays in the field of virus research and form a team with international competitiveness but we can also establish research bases for molecular virology, molecular immunology, viral ecology, etc. and lay a solid scientific foundation for persistent advancement of the prevention and control of emerging human infectious diseases in our country.

Five-year expected objectives

  1. To establish a pathogenic ecology research platform for study of natural reservoir hosts of major viruses

To isolate novel Coronavirus, nipah virus or similar paramyxoviruses, Hantaan virus, and influenza virus from wild animals; to ascertain the direct animal origin and ecological transmission chain of SARS virus; to elucidate the types and distribution of several major known human viruses infected and carried by bats in our country and the molecular mechanism for storage and transmission of these viruses in bats and other major reservoir animals, and reveal the transmission mechanism and relations of encephalitis B virus between different animal hosts; to ascertain the ecological distribution, storage modes, survival, and law of growth and decline of the highly pathogenic avian influenza viruses in the major epidemic foci (Qinghai Lake basin and Dongting Lake) and the ecological distribution of influenza viruses in poultry and mammals, and build a natural host animal virus resource bank.

  • To illuminate the molecular mutations, evolutionary laws and biological characteristics of major viruses

To reveal the genetic variation characteristics of functional genomes through whole-genome sequencing of the novel Coronavirus, encephalitis B virus, nipah virus or similar paramyxoviruses, influenza virus, etc.; to reveal the genomic evolutionary laws of the viruses through genome informatics characteristic analysis. To determine the ‘UTR and 3’UTR CIS-acting elements required for replication of dengue virus in the cells of mosquitoes and identify the species specific CIS-acting elements and cytokines that play important roles in the replication of dengue virus. To illuminate the biological characteristics of the major viruses, especially novel viruses, including pathogenicity, transmissibility, and infectivity of the viruses, reveal the effects of genetic mutations on the pathogenicity and transmissibility of the viruses, and lay the molecular foundation for research into the cross-species transmission of viruses.

  • To reveal the interaction mechanism between the viral ligands and receptors in cross-species transmission

To construct a series of false viruses and chimeric viruses by means of modern molecular biological technology and reverse genetic technology in combination with the study of interactions between the protein biomacromolecules, elucidate the interaction mechanism between the viral ligands and host cell receptors of different species and the envelope protein-receptor binding and fusion mechanism for the virus’s entry into the host cells; to reveal the types and characteristics of the receptors of part of the novel Coronaviruses and lay the foundation for cross-species infection study of the novel Coronaviruses; to identify the receptor proteins of host cells through in vitro eukaryotic expression of the viruses and ascertain how changes of important structural loci influence the virus’s ability to bind to receptors and enter cells and relate to the high pathogenicity of the viruses; to search for flavivirus receptors by use of cellular transfection and other modern technology without working on the virus.

  • To analyse the replication mechanism of major viruses in different host cells

To analyse the mechanisms for genome replication, transcription, expression and assembly of novel Coronavirus, encephalitis B virus, etc., and ascertain the functional bases for viral proteins’ replication in the different host cells; to analyse the changes in the MicroRNA expression profiles of the viruses and the host cells in the viruses’ replication in different host cells and the effects of MicroRNA expression profiles on the replication and transcription of different viruses and the structural bases therefor, and explore the species for and tissue tropism of virus infection and the common and special features of the replication in different species and tissues; to identify the mosquito cellular protein factors for the dengue fever virus and determine their role in the RNA replication of the virus. To analyse the effects of mutation, insertion and loss of polymerase genes in influenza virus on replication and pathogenesis of the virus.

  • To reveal the functional mechanism of host restriction factors in cross-species transmission of viruses

To screen and identify the host cell restriction factors that interact with the proteins of influenza virus, Hantaan virus, rabies virus and retrovirus; to illuminate the molecular mechanism of host restriction factors in the infection, pathogenesis and cross-species infection of the viruses by studying the effects of host restriction factors of different species on the viral infectivity.  To screen intracellular target proteins that interact with the relevant proteins of the viruses through study of the interactions between the viral proteins and the host intracellular proteins, explore the effects of the interactions between the related new proteins and the target proteins on the functionalities of the viruses, and lay the theoretical foundation for research and development of small molecular medicine.

  • To develop animal models for novel virus infection

To develop animal models for infection with influenza virus, novel Coronavirus and other viruses at different physiological and immune states, reveal the infection, replication, transmissibility, change in immune response level, and pathological process of the viruses in the experimental animals, and in consideration of their pre-infection baseline physiological and immune status, reveal the relation between different immune response levels and the pathogenicity of the viruses, and analyse the molecular functional mechanism whereby different immune status cause different viral pathogenicity. To build human MHC transgenic mouse infection model; to reveal CD8+T and CD4+T lymphocyte ratio and distribution patterns, cytokine release status, and other patterns of changes in immune responses, and illuminate the role of MHC restriction in cross-species transmission of the viruses; to develop mouse models for real-time monitoring of the in vivo distribution of novel viruses.

  • To foster talents and build bases

To foster a batch of top-notch international innovative talents and academic mainstays in the field of virus research to form a team with international competitiveness and create a scientific research army that is interdisciplinary, interfield, sincerely cooperative, united and aspiring, establish scientific research bases for studies of molecular virology, molecular immunology, viral ecology, etc., make significant breakthroughs in the research of novel viral ecology, viral physiology and pathology, structural virology, viral genomics, and other research fields, publish more than one hundred scientific research papers of international frontier level, and provide theoretical and scientific supports for the prevention and treatment of major infectious diseases in our country.

III. Research programme

Academic guideline:

            The project is based on the status quo of occurrence and cross-species transmission of major viral infectious diseases in our country and the great demands for prevention and treatment of infectious diseases, synthesises modern viral ecology, molecular virology, molecular biology, bioinformatics, cellular biology, and biochemistry for the purpose of analysing the cross-species transmission mechanism of viruses with novel Coronavirus, flavivirus, rabies virus, influenza virus, etc. chosen as the major objects of study, uses interdisciplinary efforts to bring out the best for revealing the molecular mechanisms for cross-species transmission and pathogenesis of the major viruses from the perspectives of ecological distribution, genetic variation and evolution of the viruses and the interactions between viruses and host cells. The implementation of the project will bring about an excellent team with international competitiveness in the research field of virus cross-species transmission, and significant breakthroughs in the study of the occurrence mechanisms of emerging infectious diseases, so as to provide theoretical bases and technical supports for the national responses to public health emergencies, the construction of public security system, and the safeguard of public health.

Technological route:

            Refer to the flowchart (figure 1) for details. This project will systematically study the ecological distribution, laws of variation and evolution, identification of viruses and receptors, intracellular replication mechanisms, effects of host restriction factors on virus replication, etc. with respect to novel Coronavirus, encephalitis B virus, rabies virus, influenza virus, and other major viruses, with the research work revolving around the molecular mechanisms for cross-species infection and transmission and pathogenesis of the viruses.

Mechanisms for cross-species Infection, transmission and pathogenesis of major viruses

IV. Yearly plans

 Contents of researchExpected objectives
First yearTo collect in quantity the serum, throat swap and annal swap samples of bats, rodents, and insectivores from all over the country; to collect cotton swap, tissue and blood samples of different animals, including wild animals, zoo animals, and specially bred economic animals, and chickens, pigs, dogs, and other livestock and poultry, at different regions. To isolate and conduct viral epidemiological investigation and study of influenza virus, novel Coronavirus, encephalitis B virus, etc., study the host characteristics of the viruses, and perform whole-genome sequencing and analysis of the viruses isolated for identification; to observe the biological characteristics of the novel viruses with the focus on their pathogenicity for animals, and the replication and cross-species transmission mechanisms of transmissible influenza virus in different hosts.To use prokaryotic or eukaryotic cells to express the envelope proteins of encephalitis B virus, dengue virus, influenza virus; to use molecular sieve chromatography, ion exchange, and other technology to isolate and purify proteins; to screen the crystal structures of envelope proteins; to develop cellular models for study of the autophagy pathways. To study the mechanisms of replication and cross-species transmission of Coronavirus in different hosts; to study the mechanisms of replication and cross-species transmission of dengue virus in different hosts; to study the molecular mechanism whereby the miRNA expression profiles replicated in different tissues regulate the virus replication. To search for host restriction factors for influenza virus based on the platform of interactions between mature viral proteins and host factors; to screen host cytokines that interact with the main proteins (mainly nucleoproteins) of Hantaan virus; to identify the host proteins related to differences between virus strains in different species by means of protein interactions, and obtain immune related cells and central nervous tissues at the early incubation and middle and late stages of infection by rabies viruses of different virulence. To develop human MHC transgenic mice; to identify the neurotropic biological characteristics of highly pathogenic avian influenza virus, West Nile virus, and other novel viruses.To collect 500 bat specimens and 500 livestock specimens.To obtain the isolated strains and genome sequences of influenza virus, novel Coronavirus, encephalitis B virus, etc.; to ascertain the pathogenicity and transmissibility of the isolated strains.  To obtain the envelope proteins of different genotypes of encephalitis B virus, dengue virus, influenza virus; to obtain the crystal structures of part of the envelope proteins.To introduce site-directed mutagenesis to the target genes of the viruses and observe the differences in the replication capacities of mutant strains and wild strains in cells from different hosts and different tissues, and search for the key sites that determine the virus’s replication in host cells of different species. To screen for 5-10 types of host cytokines that interact with the proteins of influenza virus, Hantaan virus, and rabies virus, respectively. To obtain human HLA-A2/DR1 and HLA-A11/DR1 transgenic mice. To complete the identification of the biological characteristics of 5-10 strains of highly pathogenic avian influenza virus, rabies virus, and other novel viruses, and obtain at least 4 strains of novel viruses with different neurotropism. To publish 10 or more research papers.
Second yearTo monitor the infection status of novel Coronavirus, novel Hantaan virus, novel paramyxovirus, and novel influenza virus in animals; to perform virus culture and isolation on specimens tested positive for the viruses; to analyse the differences and evolutionary relations between the new viral sequences and the genome sequenses of similar known human viruses. To continue with the isolation and investigation of viruses; to study features and patterns of changes in the biological characteristics of the viruses after transmission among different hosts; to perform whole genome sequencing on the viruses isolated for identification, use genome informatics to study the laws and features of genetic variation and molecular evolution of the viruses, and develop a reverse genetic technology system. To isolate the membrane proteins of virus-susceptible cells, use GST pull-down approach to screen for receptor candidates; to use mass spectrometry to identify receptor candidates; to study the interaction characteristics of the receptors with the HA proteins of different influenza viruses; to study on the cellular models the molecular mechanism whereby autophagy influences the entry of influenza virus. To study the mechanisms of replication and cross-species transmission of influenza virus in different hosts; to study the mechanisms of replication and cross-species transmission of coronavirus in different hosts; to study the mechanisms of replication and cross-species transmission of dengue virus in different hosts; to study the molecular mechanism whereby the miRNA expression profiles replicated in different tissues regulate the virus replication.To use protein-protein interactions and virology approaches to explore the regulatory mechanism of the host restriction factors in the replication process of influenza virus; to track the effects of the interactions between the host restriction factors and the virus nucleoproteins on the replication, assembly, maturing and release of the viruses and the relevant biological significance; to perform proteomics analysis of peripheral immune cells and central nervous tissues by chromatography and mass spectrometry technology to discover the differences manifested in the proteome level of the host before and after infection of rabies virus. To develop animal models for real-time monitoring of the in vivo distribution of the viruses and perform preliminary study of the infection dynamics of the highly pathogenic avian influenza virus and other viruses.To obtain part of the genome sequences of 10 or more virus strains; to determine the differences in the serology and genetic sequences of novel viruses and human viruses. To obtain the isolated strains and whole genome sequences of influenza virus, novel Coronavirus, encephalitis B virus, etc.; to ascertain the genetic variation patterns and biological characteristics of the prevalent strains; to develop 1 virus reverse genetic technology system. To obtain the cellular receptor candidates of encephalitis B virus and dengue virus; to obtain some important protein residues with effects on the HA proteins and sialic acid receptors of influenza virus; to acquire preliminary understanding of how influenza virus influences the signal pathway of cellular autophagy. To obtain the target recombinant viruses; to build human coronavirus banks; to reveal the effects of different mutations on the RNA translation and replication of the viruses; to obtain the host cell miRNA and virus miRNA expression profiles. To screen for 2-6 host restriction factors that interact with influenza virus and Hantaan virus, respectively; to illuminate part of the host restriction factors; to elucidate the functional mechanisms whereby part of the host restriction factors influence the replication of influenza virus and Hantaan virus; to obtain the differences in the protein levels of the hosts before and after infection of rabies virus. To ascertain the in vivo infection characteristics of highly pathogenic avian influenza virus in HLA-A2/DR1 and HLA-A11/DR1 transgenic mice, and preliminarily reveal the role of MHC restriction in the cross-species infection of highly pathogenic avian influenza virus; to complete genome sequencing of the novel viruses and discover the neurotropic-related virulence sites by bioinformatics approaches and verify the findings by reverse genetic means. To publish 20 or more research papers..
Third yearTo perform virus isolation and culturing on specimens tested positive; to carry out cell and animal sensitivity tests on the newly classified viruses and evaluate the scopes of hosts for the viruses and the pathogenicity of the viruses for experimental animals. To continue with the molecular epidemiological investigation and study of the viruses and analyse the laws of evolution and mutation of the viruses; to use the newly built reverse genetic technology platform to study the effects of gene site mutation, genetic mutation, and genetic reassortment on the pathogenicity and transmissibility of the viruses with the emphasis on exploring the key factors and elements with effects on the transmission characteristics, particularly cross-species transmission characteristics, of the viruses. To further verify the selected receptor candidates for encephalitis B virus and dengue virus by means of the cellular models; to study the expression and purification of influenza virus mutants and the crystal structures of the sialic acid receptor complexes; to study, based on the cellular models, the molecular mechanism whereby autophagy influences the entry of influenza virus. To study the mechanisms of replication and cross-species transmission of influenza virus in different hosts; to study the mechanisms of replication and cross-species transmission of coronavirus in different hosts; to study the mechanisms of replication and cross-species transmission of dengue virus in different hosts; to study the molecular mechanism whereby the miRNA expression profiles replicated in different tissues regulate the virus replication.To further explore the regulatory mechanism of host restriction factors in the replication of influenza virus and Hantaan virus; to explore the essence of how weak virulence induces early anti-infective factors (host restriction factors) and the effects that early pathogenic factors induced by strong virulence have on the host restriction factors.  To build animal models for real-time monitoring of the body distribution of the viruses; to study the functions of the neurotropic-related virulence sites of novel viruses.To obtain the whole genome sequences of 3-5 strains of the newly isolated viruses; to reveal the genetic variations of coronaviruses in wild animals or livestock. To build virus strain banks and gene banks; to determine the key genetic loci with effects on the pathogenicity and transmissibility of the viruses. To obtain 1-2 receptors or auxiliary receptors of encephalitis B virus and dengue virus; to reveal at molecular structure level the mechanism whereby the interactions between the HA proteins of influenza virus and the sialic acid receptors influence the cross-species transmission of influenza virus; to elucidate how the expression proteins of influenza virus influence the cellular autophagy level. To reveal the dynamic changes in the growth ability of the viruses in different hosts; to obtain the infectious clones of human coronavirus; to obtain the cellular protein factors on the RNAs of the four types of viruses.  To illuminate in depth the regulatory mechanism of the host restriction factors in the replication of influenza virus and Hantaan virus; to screen for 1-2 host restriction factors that interact with the rabies virus. To obtain by transgenic technology the animal models for real-time monitoring of the in vivo distribution of influenza virus; to preliminarily illuminate the functional mechanisms of the neurotropic-related virulence sites of novel viruses through cytological and zoological experiments. 7.          To publish 20 or more research papers..
Fourth year To study the receptors and the molecular mechanisms of cross-species infection of novel viruses; to analyse the relations between the prevalence of Hantavirus renal hemorrhagic disease in the major epidemic areas and the genetic variations of the virus; to monitor the epidemic status of canine influenza virus. To continue with the isolation of viruses, closely monitor the emergence of novel viruses in animal clinical situation and in human and hospital clinical situation, and study the genomic characteristics of these novel viruses; to use the newly built reverse genetic technology platform to study the roles and functions of the noncoding regions of virus genes and explore how the changes of nucleotides in the noncoding regions of virus genes influence the pathogenicity and transmissibility, particularly the cross-species transmission characteristics, of the viruses. In view of the findings from the previous three years, if the original plan fails to obtain the receptors or auxiliary receptors for the viruses, other methods will be used for further screening; to perform in vitro expression of the receptors selected and study the interactions between the virus envelope proteins and the receptors. To study the mechanisms of replication and cross-species transmission of influenza virus in different hosts; to study the mechanisms of replication and cross-species transmission of coronavirus in different hosts; to study the mechanisms of replication and cross-species transmission of dengue virus in different hosts; to study the molecular mechanism whereby the miRNA expression profiles replicated in different tissues regulate different replication.To compare effects of the host restriction factors of different species on the infection and pathogenesis of influenza virus; to compare the interactions of different types of host cells with the Hantaan viruses from different hosts and of different types that cause HFRS and HPS and severe and mild HFRS to identify the similarities and differences, and explore the molecular bases and regulatory mechanisms of host cell restriction factors of different species for the infection and pathogenesis of viruses; to further analyse the structural proteins of rabies virus to find out which elements play the major roles in influencing the host restriction factors. To carry out in vivo infection dynamics study of the novel viruses; to screen for and identify the the host proteins/MicroRNA related to neurotropism of the novel viruses.To reveal the key gene sequences of bats, rodents and insectivores related to cross-species transmission of viruses; to preliminarily ascertain the potential sources of canine influenza virus in our country. To enrich the virus strain banks and gene banks; to illuminate the genes and biological characteristics of novel viruses; to identify the functional sites in the noncoding regions of viruses. To obtain the recombinant expression produced receptor proteins; to illuminate the direct interactability of virus envelope proteins and receptors. To reveal the differences in replication capacities of the viruses in different body parts; to reveal differences in the infection and replication capacities of mutant strains and wild strains of the viruses in the cells of different species; to analyse the structural bases of the viruses’ capacities to replicate in the hosts. To elucidate the molecular bases and regulatory mechanisms for the effects of the host restriction factors of different species on the infection and pathogenesis of influenza virus and Hantaan virus; to find out which elements in the structural proteins of rabies virus play the major roles in influencing the host restriction factors.To complete the in vivo mouse infection dynamics study of the highly pathogenic avian influenza viruses and reveal the tissue tropism and infection process of highly pathogenic avian influenza virus in vivo mammals; to discover the host proteins/MicroRNA related to neurotropism of the novel viruses by proteomics, biochips, and other technology. To publish 25 research papers..
Fifth yearTo carry out in vivo infection dynamics study of the novel viruses; to screen for and identify the the host proteins/MicroRNA related to neurotropism of the novel viruses.To continue with the isolation, genome informatics study and biological characteristics study of the viruses; to synthesize the findings achieved by the research group in the previous years and analyse the role genomic mutations play in the cross-species transmission of the virus; To screen the crystal structures of the complexes of viral envelope proteins and receptors; To study the functions of host proteins/MicroRNA related to neurotropism of the novel viruses. To further supplement and complete the research results from the previous 4 years and perform summary and acceptance of the project.To illuminate the relations between the novel Coronavirus, Hantaan virus, influenza virus, etc. carried by animals and the relevant human diseases. To enrich the virus strain banks and gene banks; to summarise the epidemic patterns of the viruses and the functional sites with effects on the biological characteristics of the viruses.To obtain the crystal structures of the complexes of viral envelope proteins and receptors, illuminate the interaction mechanisms between viruses and receptors, and reveal the mechanisms for cross-species transmission of the viruses.To illuminate the roles and molecular mechanisms of host restriction factors in the infection, pathogenesis and cross-species infection of influenza virus; to illuminate the molecular mechanism for and immunopathogenetic meaning of the formation of numerous inclusion bodies after infection of Hantaan virus; to illuminate the innate immune mechanism of the in vivo central nervous system against the rabies virus and the pathogenetic mechanism of the rabies virus on the central nervous system. To publish 25 or more papers.To complete acceptance of the project.
  • Contents of research
  • Ecological distributions of major viruses in natural hosts

On the basis of the investigation and studies on such animal viruses as novel Coronavirus, encephalitis B virus, and Hantaan virus from different regions, different times, and different hosts, we will further expand the investigation to more regions and hosts, ascertain the types and distributions of several major know human viruses infected and carried by bats and the molecular mechanisms for the storage and transmission of the viruses in bats and other major reservoir animals, and study the mechanisms and relations for transmission of encephalitis B virus among different animal hosts; we will study the effects of ecological environmental factors on the scopes of hosts infectable by the viruses; we will evaluate the potential dangers the novel viruses pose to humans and animals; we will investigate the ecological distributions of influenza virus among wildfowl, poultry, and mammals, and study the relations between the migration of migratory birds and the epidemics of diseases to provide theoretical bases for formulation of epidemic prevention and control policies; we will build a database about the ecological distributions of viruses among natural hosts to lay the phisical foundation for study and analysis of the cross-species transmission of the major viruses.

  • Molecular mutations, evolutionary laws, biological characteristics of major viruses

On the basis of our ecological and epidemiological investigation and study of the major viruses, we will choose novel Coronavirus, encephalitis B virus, rabies virus, influenza virus, and other major viruses as our objects of study, perform whole genome sequencing on these viruses and study the variations and evolutionary characteristics of the functional genomes; in the meantime, we will study the biological characteristics of the major viruses, especially the novel viruses, including the pathogenicity, transmissibility, and infectivity of the viruses, and study the effects of genetic variations on the cross-species transmission of the viruses. Since studies in the recent years show that the nonstructural proteins and noncoding regions of the viruses play a very important role in the pathogenicity and transmissibility of the viruses, we will put a premium on studying the variations in and evolutionary characteristics of the noncoding regions of the viruses in our whole gemone sequencing study and observe how the variations in the noncoding regions influence the biological characteristics of the viruses.

  • Interactions between the viruses and the receptors in cross-species transmission

We will seek breakthroughs in the study of encephalitis B vius, influenza virus, etc. and study their affinity with the receptors and the interaction mechanisms between the cellular receptors and ligands; we will use modern molecular biological technology and reverse genetic technology to construct a series of false viruses and chimeric viruses. We will study the interactions between viruses and host cells of different species in combination with the study of the interactions between protein biomacromolecules; in the meantime, we will use insect cells to express the surface proteins and various mutant proteins of the viruses, study the roles of receptors and ligands at the structural level and on this basis, use physical method (SPR) and biological test methods to study the effects of mutations in the viral surface proteins on the binding ability of the receptors. We will use reverse genetics technology to package the viruses and evaluate at cellular and animal levels the effects of virus mutations on host tropism and pathogenesis of the viruses so as to reveal the mechanisms for cross-species transmission of the viruses.

  • Replication mechanism of major viruses in different hosts

We will seek breakthroughs in the study of novel human Coronavirus, dengue virus, and influenza virus, use the reverse genetic platform and adopt molecular biological, cellular biological and other means to study the functional bases for viral proteins to regulate, mutate, reassort, develop tropism in the process of replication in the cells of different hosts,  study the changes in the MicroRNA expression profiles of the virus and the host cells in the course of virus replication in different host cells and the effects of MicroRNA expression profiles on the replication and transcription of different viruses and the structural bases therefor, and explore the species for and tissue tropism of virus infection and the common and special features of the replication in different species and tissues; we will study the CIS-acting elements involved in the replication of dengue fever virus in the mosquito cells  and determine their role in the RNA replication of the virus.

  • Functions of host restriction factors in cross-species transmission of viruses

We will use the yeast two-hybrid technique and virology research technology to explore the regulatory mechanism of the host restriction factors in the replication of influenza virus; we will compare the effects of the host restriction factors of different species on the infection and pathogenesis of influenza virus, and study and analyse the role and molecular mechanism of host restriction factors in the infection, pathogenesis, and cross-species infection of influenza virus. We will screen for the host cytokines that interact with the critical proteins of Hantaan virus and rabies virus, track how the host restriction factors, after interacting with the viral nucleoproteins, influence the replication, assembly, maturing, and release of the viruses and the relevant biological meaning, and analyse the molecular mechanism for and immunopathogenetic meaning of the formation of inclusion bodies after the viruses infect the cells, and explore the molecular bases and regulatory mechanisms of host cell restriction factors of different species for the infection and pathogenesis of viruses.

  • Animal models for cross-species infection of novel viruses

We will use mice of different ages (adult and old mice) as experimental objects, describe the differences in their physiological status and baseline immune levels, choose influenza virus or novel Coronavirus to infect the mice to build infection models for animals of different physiological and immune status; we will study the infection, replication, transmissibility, changes in the immune response levels, pathological process, etc., analyse the relations between different physiological and immune status, different immune response levels and the pathogenicity of viruses in consideration of the pre-infection baseline physiological and immune status, and illuminate the molecular mechanism whereby different immune status result in different pathogenicity of the viruses. We will build human MHC transgenic mouse infection models. We will perform dynamic monitoring of the CD8+T and CD4+T lymphocyte ratio and distribution pattern, cytokine release status, and other patterns of changes in immune responses, and analyse the role of MHC restriction in cross-species transmissions of the viruses.

Appendix II. Some information about the viruses involved in the study

Now let me provide some basic knowledge about these viruses here.

Nipah virus infection is a zoonotic illness that is transmitted to people from animals, and can also be transmitted through contaminated food or directly from person-to-person. In infected people, it causes a range of illnesses from asymptomatic (subclinical) infection to acute respiratory illness and fatal encephalitis. The virus can also cause severe disease in animals such as pigs, resulting in significant economic losses for farmers.

Although Nipah virus has caused only a few known outbreaks in Asia, it infects a wide range of animals and causes severe disease and death in people.

https://www.who.int/health-topics/nipah-virus-infection#tab=tab_1

Hantaviruses (Hantaan Virus) are a family of viruses spread mainly by rodents and can cause varied disease syndromes in people worldwide… Hantaviruses in the Americas are known as “New World” hantaviruses and may cause hantavirus pulmonary syndrome (HPS). Other hantaviruses, known as “Old World” hantaviruses, are found mostly in Europe and Asia and may cause hemorrhagic fever with renal syndrome (HFRS).

Each hantavirus serotype has a specific rodent host species and is spread to people via aerosolized virus that is shed in urine, feces, and saliva, and less frequently by a bite from an infected host.

https://www.cdc.gov/hantavirus/index.html

  • Humans can be infected with avian, swine and other zoonotic influenza viruses
  • Human infections are primarily acquired through direct contact with infected animals or contaminated environments, these viruses have not acquired the ability of sustained transmission among humans.
  • Avian, swine and other zoonotic influenza virus infections in humans may cause disease ranging from mild upper respiratory tract infection (fever and cough), early sputum production and rapid progression to severe pneumonia, sepsis with shock, acute respiratory distress syndrome and even death. Conjunctivitis, gastrointestinal symptoms, encephalitis and encephalopathy have also been reported to varying degrees depending on subtype.

Influenza viruses, with the vast silent reservoir in aquatic birds, are impossible to eradicate. Zoonotic influenza infection in humans will continue to occur…

https://www.who.int/news-room/fact-sheets/detail/influenza-(avian-and-other-zoonotic)

  • Rabies is a vaccine-preventable viral disease which occurs in more than 150 countries and territories.
  • Dogs are the main source of human rabies deaths, contributing up to 99% of all rabies transmissions to humans.

The first symptoms of rabies may be very similar to those of the flu including general weakness or discomfort, fever, or headache. These symptoms may last for days.

There may be also discomfort or a prickling or itching sensation at the site of the bite, progressing within days to acute symptoms of cerebral dysfunction, anxiety, confusion, and agitation. As the disease progresses, the person may experience delirium, abnormal behavior, hallucinations, hydrophobia (fear of water), and insomnia. The acute period of disease typically ends after 2 to 10 days. Once clinical signs of rabies appear, the disease is nearly always fatal, and treatment is typically supportive. To date less than 20 cases of human survival from clinical rabies have been documented, and only a few survivors had no history of pre- or postexposure prophylaxis.

https://www.who.int/news-room/fact-sheets/detail/rabies

https://www.cdc.gov/rabies/symptoms/index.html

  • West Nile virus can cause a fatal neurological disease in humans.
  • However, approximately 80% of people who are infected will not show any symptoms.
  • West Nile virus is mainly transmitted to people through the bites of infected mosquitoes.
  • Vaccines are available for use in horses but not yet available for people.
  • Birds are the natural hosts of West Nile virus.

https://www.who.int/news-room/fact-sheets/detail/west-nile-virus

  • Japanese encephalitis virus (JEV) is a flavivirus related to dengue, yellow fever and West Nile viruses, and is spread by mosquitoes.
  • JEV is the main cause of viral encephalitis in many countries of Asia with an estimated 68 000 clinical cases every year.
  • Although symptomatic Japanese encephalitis (JE) is rare, the case-fatality rate among those with encephalitis can be as high as 30%. Permanent neurologic or psychiatric sequelae can occur in 30%–50% of those with encephalitis.
  • There is no cure for the disease. Treatment is focused on relieving severe clinical signs and supporting the patient to overcome the infection.
  • Safe and effective vaccines are available to prevent

https://www.who.int/news-room/fact-sheets/detail/japanese-encephalitis

Dengue viruses are spread to people through the bite of an infected Aedes species (Ae. aegypti or Ae. albopictus) mosquito. These mosquitoes also spread Zika, chikungunya, and other viruses.

The most common symptom of dengue is fever with any of the following:

  • Nausea, vomiting
  • Rash
  • Aches and pains (eye pain, typically behind the eyes, muscle, joint, or bone pain)
  • Any warning sign

Symptoms of dengue typically last 2–7 days. Most people will recover after about a week.

There is no specific medicine to treat dengue.

https://www.cdc.gov/dengue/symptoms/index.html

Now here comes the coronaviruses. However, I would like to show you both the symptoms of SARS and COVID-19.

Severe acute respiratory syndrome (SARS) is a viral respiratory disease caused by a SARS-associated coronavirus.

The incubation period of SARS is usually 2-7 days but may be as long as 10 days.

The first symptom of the illness is generally fever (>38°C), which is often high, and sometimes associated with chills and rigors. It may also be accompanied by other symptoms including headache, malaise, and muscle pain. At the onset of illness, some cases have mild respiratory symptoms. Typically, rash and neurologic or gastrointestinal findings are absent, although a few patients have reported diarrhoea during the early febrile stage.

After 3-7 days, a lower respiratory phase begins with the onset of a dry, non-productive cough or dyspnoea (shortness of breath) that may be accompanied by, or progress to, hypoxemia (low blood oxygen levels). In 10–20% of cases, the respiratory illness is severe enough to require intubation and mechanical ventilation. Chest radiographs may be normal throughout the course of illness, though not for all patients. The white blood cell count is often decreased early in the disease, and many people have low platelet counts at the peak of the disease.

https://www.who.int/health-topics/severe-acute-respiratory-syndrome#tab=tab_1

https://www.who.int/health-topics/severe-acute-respiratory-syndrome#tab=tab_2

The symptoms of COVID-19 are as follows:

Common Symptoms

Researchers in China found that the most common symptoms among people who were hospitalized with COVID-19 include:

Fever: 99%

Fatigue:70%

A dry cough: 59%

Loss of appetite: 40%

Body aches: 35%

Shortness of breath: 31%

Mucus or phlegm: 27%

Symptoms usually begin 2 to 14 days after you come into contact with the virus.

Other symptoms may include:

Sore throat

Headache

Chills, sometimes with shaking

Loss of smell or taste

Congestion or runny nose

Nausea or vomiting

Diarrhea

Emergency Symptoms

Trouble breathing

Constant pain or pressure in your chest

Bluish lips or face

Sudden confusion

Strokes have also been reported in some people who have COVID-19.

In peer-reviewed literature and public discussion, persistent symptoms are being reported among COVID-19 survivors, including individuals who initially experience a mild acute illness…

While older patients may have an increased risk for severe disease, young survivors, including those physically-fit prior to SARS-CoV-2 infection, have also reported symptoms months after acute illness…

Though there is limited information on late sequelae of COVID-19, reports of persistent symptoms in persons who recovered from acute COVID-19 illness have emerged. . The most commonly reported symptoms include fatigue, dyspnea, cough, arthralgia, and chest pain. Other reported symptoms include cognitive impairment, depression, myalgia, headache, fever, and palpitations. More serious complications appear to be less common but have been reported. These complications include:

  • Cardiovascular: myocardial inflammation, ventricular dysfunction
  • Respiratory: pulmonary function abnormalities
  • Renal: acute kidney injury
  • Dermatologic: rash, alopecia
  • Neurological: olfactory and gustatory dysfunction, sleep dysregulation, altered cognition, memory impairment
  • Psychiatric: depression, anxiety, changes in mood

https://www.webmd.com/lung/covid-19-symptoms#1

https://www.cdc.gov/coronavirus/2019-ncov/hcp/clinical-care/late-sequelae.html

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