Construction method and application of novel coronavirus-caused pneumonia model

Abstract

The invention provides a construction method and application of a novel coronavirus induced pneumonia model. According to the invention, new coronavirus-caused pneumonia models are constructed by using new coronavirus-caused mice of different pedigree in sequence. The pneumonia model caused by the novel coronavirus is high in pneumonia degree and good in simulation performance on clinical symptoms of mutant virus infection, can be used for screening new crown drugs, evaluating the effect of new crown treatment drugs, researching the pathogenesis of new crowns and the like, and has good application prospects.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine, and specifically relates to a method for constructing a pneumonia model caused by a new coronavirus and its application. Background Art

[0002] The new coronavirus (COVID-19) has been constantly mutating during its epidemiological evolution, deriving multiple variants. The World Health Organization (WHO) has listed several variants as variants of concern (VOCs), including Alpha, Beta, Delta, and Omicron. The presence of adaptive mutations in the genomes of these variants helps the virus evade the host's immune response, leading to an increase in the global reinfection rate. In 2022, the risk ratio (HR) of severe illness from reinfection with the COVID-19 variant in the Washington, D.C. population increased from 0.03% during the initial infection to 3.31%. However, whether the antibodies already present in the host's body can still provide protection during reinfection and whether repeated infection will increase the risk of disease after the first infection, especially the risk of pneumonia, requires further study.

[0003] With the emergence of new coronavirus mutants, the clinical symptoms of viral infection may change. The continuous emergence of mutants has brought new challenges to the research on the transmission, prevention and treatment of the new coronavirus, as well as new requirements for the construction of various animal models. It is necessary to prepare an animal model that can better simulate the clinical symptoms of mutant virus infection, so as to further develop vaccines and antiviral drugs for the prevention and treatment of pneumonia caused by the new coronavirus. Summary of the invention

[0004] In view of this, in order to make up for the deficiencies of the prior art, the present invention is proposed.

[0005] The first aspect of the present invention provides a method for constructing a non-human animal model of pneumonia caused by a new coronavirus, the method comprising infecting a non-human animal with a first new coronavirus and then infecting the non-human animal with a second new coronavirus.

[0006] Furthermore, the first novel coronavirus and the second novel coronavirus belong to different lineages or different variants of the same variant.

[0007] Furthermore, the first novel coronavirus and the second novel coronavirus belong to different lineages of the same variant strain.

[0008] In the present invention, mutation refers to the gene sequence mutation that occurs during the replication of the virus, which is mainly caused by the high error rate during the replication of RNA viruses. During the propagation and replication process, various mutants will be produced due to its lack of ability to correct replication errors. These mutations may cause changes in the infectivity, toxicity, immune escape characteristics, etc. of the virus. Up to now, a variety of variants of the new coronavirus have appeared, such as Alpha variants, Beta variants, Gamma variants, Delta variants, Omicron variants, and other variants. It should be understood that the virus may change over time and new classifications are envisioned. It should be understood that when new classifications appear, these new classifications are also within the scope of the present invention.

[0009] In the present invention, the Alpha variant (alias: B.1.1.7; VOC 202012 / 01; 201 / 501YV1) has a series of mutations in the S protein, namely: Δ69-70, A144, N501Y, A570D, D614G, P681H, T7161, S982A and D1118H. Compared with the original virus strain, the transmission ability of the Alpha variant has increased by 40% to 70%, which means that even if the severity of the disease has not changed, the virus will still cause higher incidence and more hospitalizations. The Beta variant (alias: VOC 202012 / 02; 20H / 501Y.V2) has 10 mutation sites in the S protein, namely D80A, D215G, A241-243, K417N, E484K, N501Y, D614G and A701V, of which the two mutation sites N501Y and D614G in the receptor binding region (RBD) of the new coronavirus S protein also appear in the Alpha variant. The Gamma variant (alias: R1; 20J / 501YV3) has 12 mutations in the S protein, namely L18F, T20N, P26S, D138Y, R190S, K417T, E484K, N501Y, D614G, H655Y, T1027 and V1176E. The Delta (B.1.617.2) variant belongs to a sub-branch of B.1.617, which also contains two branches, B.1.617.1 (Kappa) and B.1.617.3. These branches contain different variants. Among them, the S protein of the Delta variant has 10 amino acid mutations T19R, G142D, A156-157, R158G, L452R, E484Q, T478K, D614G, P681R, and D950N. In addition to the D614G mutation, the S protein of the Omicron variant also includes 8 mutations in NTD (A67V, Δ69-70, T951, G142D, A141-143, A211, L212, and Ins214EPE), and 15 mutations in RBD (G339D, S371L, S373P, S375F, K417N, N440K, G446S , S477N, T478K, E484A, Q493R, G496S, Q498R, N501Y and Y505H), T547K mutation, 3 mutations close to the Furin cleavage site (H655Y, N679K and P681H) and 6 mutations in the S2 region (N764K, D796Y, N856K, Q954H, N969 and L981F).

[0010] Furthermore, the variant strain is selected from the Omicron variant strain.

[0011] Furthermore, the Omicron variants include BA.1, BA.2, BA.3, BA.4, BA.5 or their progeny lineages.

[0012] In the present invention, examples in which the first novel coronavirus and the second novel coronavirus belong to different lineages of the same variant strain can be listed as the first novel coronavirus is selected from BA.1 or its descendant lineage, and the second novel coronavirus is selected from BA.2, BA.3, BA.4, BA.5 or their descendant lineages; the first novel coronavirus is selected from BA.2 or its descendant lineage, and the second novel coronavirus is selected from BA.1, BA.3, BA.4, BA.5 or their descendant lineages; the first novel coronavirus is selected from BA.3 or its descendant lineage, and the second novel coronavirus is selected from BA.1, BA.2, BA.4, BA.5 or their descendant lineages; the first novel coronavirus is selected from BA.4 or its descendant lineage, and the second novel coronavirus is selected from BA.1, BA.1, BA.3, BA.5 or its descendant lineage; the first novel coronavirus is selected from BA.5 or its descendant lineage, and the second novel coronavirus is selected from BA.1, BA.2, BA.3, BA.4 or its descendant lineage.

[0013] In the present invention, the progeny lineage refers to the different genetic lineages or branches formed within the virus population due to factors such as gene mutation and natural selection during the process of virus evolution.

[0014] In the present invention, the BA.2 progeny lineage includes but is not limited to XBB, which is derived from the recombination between two sub-lineages of BA.2 (BA.2.10.1 and BA.2.75).

[0015] Furthermore, the Omicron variant is selected from BA.5 and XBB.

[0016] Furthermore, the first novel coronavirus is selected from BA.5.

[0017] Furthermore, the second novel coronavirus is selected from XBB.

[0018] Further, the titer of the first novel coronavirus is 10 4 ~10 6 TCID 50 / ml.

[0019] Further, the titer of the first novel coronavirus is 10 5 TCID 50 / ml.

[0020] Furthermore, the usage volume of the first novel coronavirus is 10μl~1ml.

[0021] Furthermore, the usage volume of the first novel coronavirus is 50 μl.

[0022] Furthermore, the titer of the second novel coronavirus is 10 4 ~10 6 TCID 50 / ml.

[0023] Furthermore, the titer of the second novel coronavirus is 10 5 TCID 50 / ml.

[0024] Furthermore, the usage volume of the second new coronavirus is 10μl~1ml.

[0025] Furthermore, the usage volume of the second new coronavirus is 50 μl.

[0026] Furthermore, the second novel coronavirus is infected 2 to 50 weeks after the first novel coronavirus infection.

[0027] Furthermore, the second novel coronavirus is infected 2 to 5 weeks after the first novel coronavirus infection.

[0028] Furthermore, the second novel coronavirus was infected 4 weeks after the first novel coronavirus infection.

[0029] Furthermore, the non-human animal model of pneumonia caused by the new coronavirus is a moderate to severe pneumonia model.

[0030] Furthermore, the non-human animal model of pneumonia caused by the novel coronavirus exhibits moderate to severe pneumonia at least 0.5 days after the second novel coronavirus infection.

[0031] Furthermore, the non-human animal model of pneumonia caused by the novel coronavirus showed moderate to severe pneumonia 3 days after the second novel coronavirus infection.

[0032] Furthermore, the non-human animal model of pneumonia caused by the novel coronavirus showed moderate to severe pneumonia 3 days after the second novel coronavirus infection, and there was still a clear trend of worsening.

[0033] In the present invention, a non-human animal model refers to a non-human animal that has or exhibits characteristics of a disease or condition.

[0034] In the present invention, non-human animals include non-human vertebrates, more preferably mammals, which refer to all members of the class Mammalia, such as domesticated livestock (e.g., cattle, horses, pigs), pets (e.g., dogs, cats), or rodents. The term "rodent" refers to any and all members of the phylogenetic class Rodentia (e.g., mice, rats, squirrels, beavers, woodchucks, gophers, voles, groundhogs, hamsters, guinea pigs, and agouti), including any descendants derived therefrom.

[0035] Furthermore, the mammal is selected from mice.

[0036] Furthermore, the mouse is a nude mouse.

[0037] In the present invention, the scientific name of nude mouse is athymic nude mouse, which is a mouse with congenital thymus defect due to gene mutation.

[0038] Furthermore, the nude mice include but are not limited to BALB / c, NIH, NC, Swiss, 03H, and C57BL.

[0039] Furthermore, the nude mice are selected from BALB / c.

[0040] Further, the method comprises using a dose of 5×10 3 TCID 50 BALB / c mice were infected with BA.5 for 4 weeks and then treated with 5×10 3 TCID 50 XBB infection was used to obtain a non-human animal model of pneumonia caused by the new coronavirus.

[0041] A second aspect of the present invention provides any of the following methods:

[0042] (1) A method for screening drug candidates for treating pneumonia caused by a novel coronavirus, the method comprising:

[0043] a) administering the agent to be screened to a non-human animal with pneumonia caused by the novel coronavirus constructed by the method described in the first aspect of the present invention.

[0044] b) detecting the therapeutic effect of the agent to be screened on pneumonia caused by the new coronavirus;

[0045] (2) A method for evaluating the therapeutic effect of a drug for treating pneumonia caused by the new coronavirus, the method comprising:

[0046] a) administering the drug to a non-human animal with pneumonia caused by the novel coronavirus constructed by the method described in the first aspect of the present invention;

[0047] b) detecting the therapeutic effect of the drug on pneumonia caused by the novel coronavirus;

[0048] (3) A method for studying the pathogenesis of pneumonia caused by the novel coronavirus, the method comprising using a non-human animal with pneumonia caused by the novel coronavirus constructed by the construction method described in the first aspect of the present invention to study the pathogenesis of pneumonia caused by the novel coronavirus.

[0049] In the present invention, the method for screening drug candidates for treating pneumonia caused by the new coronavirus may involve administering different amounts of drug candidates (from no drug to an amount of drug close to the upper limit of the amount that can be successfully delivered to an animal, such as within the toxic limit), and may include delivering drugs in different formulations and routes. A single drug may be administered, or the drug may be combined in a combination of two or more drugs, especially in the case where the administration of a drug combination may result in a synergistic effect. The ability of a candidate agent to treat pneumonia caused by the new coronavirus may be evaluated by administering the candidate drug to a non-human animal model of pneumonia caused by the new coronavirus and evaluating the regulation of the pneumonia phenotype caused by the new coronavirus. The regulation of the pneumonia phenotype caused by the new coronavirus may be evaluated, for example, by evaluating the presence or absence of effects such as fever duration, viral nucleic acid negative conversion time, lung inflammation absorption, lung tissue viral load, lung tissue inflammatory cytokine or chemokine expression levels, etc. The candidate drug may be administered to the non-human animal model of pneumonia caused by the new coronavirus described in the first aspect of the present invention, and then the ability of the candidate drug to promote the prevention of pneumonia caused by the new coronavirus may be evaluated. Prevention of pneumonia caused by the new coronavirus refers to reducing the incidence and / or severity of pneumonia caused by the new coronavirus in animals relative to the expected incidence and / or severity of pneumonia caused by the new coronavirus in the absence of intervention (e.g., without administration of a drug with antiviral activity).

[0050] Drug candidates can be obtained from a wide variety of sources, including but not limited to synthetic, naturally occurring or recombinantly produced molecules, including small molecules, peptides, antibodies or other polypeptides. For example, a variety of organic compounds and biomolecules can be synthesized randomly or directed, or natural compound libraries in the form of bacterial, fungal, plant or animal extracts, or natural or synthetic libraries and compounds modified by conventional chemical, physical or biochemical means, or directed or random chemical modification of known pharmacological agents, such as acylation, alkylation, esterification, amidation, etc. to generate structural analogs, or polysaccharides, vaccines.

[0051] In the present invention, the term "treatment" refers to the administration of a compound or composition to control the progression of a disease. The control of disease progression should be understood as achieving a beneficial or desired clinical result, including but not limited to alleviating symptoms, reducing the duration of the disease, stabilizing the pathological state (especially avoiding additional exacerbations), delaying the progression of the disease, preventing, improving, alleviating (partially and completely) the pathological state.

[0052] Furthermore, the therapeutic effects include shortening the duration of fever, shortening the time for viral nucleic acid to turn negative, improving lung inflammation absorption, reducing the viral load in lung tissue, and reducing the expression level of inflammatory cytokines or chemokines in lung tissue.

[0053] The drug of the present invention is preferably administered in a pharmaceutically acceptable vehicle. Suitable pharmaceutical carriers are known to those skilled in the art. For parenteral administration, the compound is usually dissolved or suspended in sterile water or saline. For enteral administration, the compound is incorporated into an inert carrier in the form of tablets, liquids or capsules. Suitable carriers can be starch or sugar, and include lubricants, flavoring agents, adhesives and other materials of the same nature. The compound can also be applied topically by topical application of solutions, creams, gels or polymeric materials (e.g., PluronicTM, BASF). Alternatively, the compound can be administered in liposomes or microspheres (or microparticles). Methods for preparing liposomes and microspheres for administration to patients are known to those skilled in the art. In essence, the material is dissolved in an aqueous solution, appropriate phospholipids and lipids are added together with surfactants if necessary, and the material is dialyzed or sonicated as needed. Microspheres formed by polymers or proteins are well known to those skilled in the art and can be customized to enter the bloodstream directly through the gastrointestinal tract. Alternatively, the compound can be incorporated and the microspheres or microsphere complexes can be implanted to slowly release over a period of days to months.

[0054] The drug of the present invention can also be used in combination with other drugs for treating pneumonia caused by the new coronavirus, and other therapeutic compounds can be administered simultaneously with the main active ingredient, or even simultaneously in the same composition. Other therapeutic compounds can also be administered separately in a separate composition or in a dosage form different from the main active ingredient. Partial doses of the main ingredient can be administered simultaneously with other therapeutic compounds, while other doses can be administered alone. During the treatment process, the dosage of the drug of the present invention can be adjusted according to the severity of the symptoms, the frequency of recurrences, and the physiological response of the treatment regimen.

[0055] In the present invention, other drugs for treating pneumonia caused by the new coronavirus include but are not limited to namatevir, azithromycin, monoravir, and senotetravir.

[0056] The third aspect of the present invention provides any of the following applications:

[0057] (1) Use of the non-human animal model of pneumonia caused by the novel coronavirus constructed by the method described in the first aspect of the present invention in screening drug candidates for treating pneumonia caused by the novel coronavirus.

[0058] (2) Use of the non-human animal model of pneumonia caused by the novel coronavirus constructed by the method described in the first aspect of the present invention in evaluating the therapeutic effect of drugs for treating pneumonia caused by the novel coronavirus.

[0059] (3) Application of the non-human animal model of pneumonia caused by the novel coronavirus constructed by the method described in the first aspect of the present invention in studying the pathogenesis of pneumonia caused by the novel coronavirus.

[0060] Furthermore, the therapeutic effects include but are not limited to shortening the duration of fever, shortening the time for viral nucleic acid to turn negative, improving lung inflammation absorption, reducing lung tissue viral load, and reducing the expression level of inflammatory cytokines or chemokines in lung tissue.

[0061] The fourth aspect of the present invention provides the use of the new coronavirus Omicron BA.5 and XBB in constructing a non-human animal model of pneumonia caused by the new coronavirus.

[0062] The present invention has the following advantages and beneficial effects:

[0063] The present invention provides a method for constructing a pneumonia model caused by a novel coronavirus and its application. The present invention successively uses novel coronavirus-infected mice of different lineages to construct a pneumonia model caused by a novel coronavirus. The novel coronavirus-induced pneumonia model has a high degree of pneumonia and a good simulation of the clinical symptoms of mutant virus infection. It can be used for novel coronavirus drug screening, evaluation of the effects of novel coronavirus therapeutic drugs, and research on the pathogenesis of novel coronavirus, and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0064] Figure 1 This is a graph showing the weight of mice after reinfection.

[0065] Figure 2 This is a graph showing the viral load in the lung tissue of mice after reinfection.

[0066] Figure 3 This is a graph showing the severity of inflammation in the lung tissue of mice after reinfection.

[0067] Figure 4 This is a graph showing the expression of cytokines and chemokines in the lung tissue and serum of mice after reinfection. DETAILED DESCRIPTION

[0068] The present invention is further described below in conjunction with the embodiments. The following description is only a preferred embodiment of the present invention, and does not limit the present invention in other forms. Any technician familiar with the profession may use the above disclosed technical content to change it into an equivalent embodiment with equivalent changes. Any simple modification or equivalent change made to the following embodiments based on the technical essence of the present invention without departing from the content of the present invention falls within the protection scope of the present invention.

[0069] Example

[0070] 1. Experimental Materials

[0071] 1.1 Experimental viruses

[0072] Different subtypes of SARS-CoV-2Omicron: BA.5, XBB.

[0073] 1.2 Experimental animals and cells

[0074] Female BALB / c mice aged 6 to 8 weeks were purchased from Beijing Huafukang Biotechnology Co., Ltd.

[0075] 1.3 Experimental instruments

[0076] Electric grinder (Retsch MM400, Germany); PCR instrument (Bio-Rad, USA); real-time fluorescence quantitative PCR instrument (Applied Biosystems 7500, USA).

[0077] 1.4 Experimental reagents and consumables

[0078] RNA extraction kit: RNeasy Mini Kit (Qiagen);

[0079] Reverse transcription kit: PrimeScript TM RTreagent Kit with gDNA Eraser(Perfect RealTime)(TaKaRa RR047A);

[0080] Applied Biosystems TM PowerUp TM SYBR TM Green Master Mix(ThermoScientific).

[0081] 2. Experimental methods

[0082] Animal Experimentation:

[0083] BALB / c mice were randomly divided into 2 groups and infected with OmicronBA.5 strain, and then infected with BA.5 strain or XBB strain again 28 days after infection. A control group and a group infected with BA.5 or XBB alone were set up at the same time. The virus titer used was 10 5 TCID 50 / ml, the usage volume is 50μl.

[0084] The clinical symptoms of mice were observed after infection, and the lung tissues of mice were collected 5 days after reinfection for virology, pathology and other monitoring. All experimental operations related to SARS-CoV-2 virus infection were carried out in ABSL-3. Animal experiments passed animal ethics review with approval number: BLL23007.

[0085] 3. Experimental results

[0086] Primary infection with BA.5 followed by infection with XBB strain induced a more severe inflammatory response.

[0087] The weight of mice in all infection groups showed a downward trend. The weight loss rate after BA.5XBB infection was similar. The weight loss rate of the BA.5-BA.5 reinfection group was the slowest, and there was a subsequent upward trend at 3 dpi. The weight of mice in the BA.5-XBB reinfection group continued to decrease within 5 days after infection ( Figure 1 ). Compared with the BA.5-BA.5 group, the lung index of the reinfected group (BA.5-XBB group) was significantly increased 3 and 5 days after infection. Compared with the primary infection BA.5 / XBB group, the viral load in the lung tissue of mice in the BA.5-BA.5 and BA.5-XBB reinfected groups was cleared ( Figure 2 ). However, in terms of pathological manifestations, the degree of lung inflammation in mice in the BA.5-BA.5 group was significantly reduced, with 2 / 3 of the mice showing moderate pneumonia on the third day of infection and 1 / 3 showing mild pneumonia, and the degree of inflammation gradually decreased. Among the mice re-infected with the BA.5-XBB strain, 1 / 3 showed severe pneumonia, 2 / 3 showed moderate to severe pneumonia, and there was a large amount of interstitial inflammatory cell infiltration, showing a clear trend of worsening ( Figure 3 ).

[0088] At the same time, the expression of cytokines and chemokines in lung tissue and serum of the BA.5-XBB group was significantly increased ( Figure 4 ), which suggests that in the presence of primary infection with the Omicron BA.5 strain, reinfection with the XBB variant induced a more severe inflammatory response, leading to aggravated pathological damage to lung tissue.

[0089] The description of the above embodiments is only used to understand the method and core idea of ​​the present invention. It should be pointed out that, for those skilled in the art, several improvements and modifications can be made to the present invention without departing from the principle of the present invention, and these improvements and modifications will also fall within the scope of protection of the claims of the present invention.

Claims

1. A method for constructing a non-human animal model of pneumonia caused by a new coronavirus, characterized in that: The method includes infecting a non-human animal with a first novel coronavirus and then infecting the non-human animal with a second novel coronavirus.

2. The method according to claim 1, characterized in that The first novel coronavirus and the second novel coronavirus belong to different lineages or different variants of the same variant; Preferably, the first novel coronavirus and the second novel coronavirus belong to different lineages of the same variant strain; preferably, the variant strains include Alpha variant strains, Beta variant strains, Gamma variant strains, Delta variant strains, Omicron variant strains, and other variant strains; Preferably, the variant is selected from the Omicron variant; Preferably, the Omicron variants include BA.1, BA.2, BA.3, BA.4, BA.5 or their descendant lineages; Preferably, the BA.2 progeny pedigree comprises XBB; Preferably, the Omicron variant is selected from BA.5 and XBB; Preferably, the first novel coronavirus is selected from BA.5; Preferably, the second novel coronavirus is selected from XBB.

3. The method according to claim 1, characterized in that: The titer of the first novel coronavirus was 10 4 ~10 6 TCID 50 / ml; Preferably, the titer of the first novel coronavirus is 10 5 TCID 50 / ml; Preferably, the usage volume of the first novel coronavirus is 10 μl to 1 ml; Preferably, the first novel coronavirus is used in a volume of 50 μl.

4. The method according to claim 1, characterized in that: The titer of the second novel coronavirus was 10 4 ~10 6 TCID 50 / ml; Preferably, the titer of the second novel coronavirus is 10 5 TCID 50 / ml; Preferably, the usage volume of the second novel coronavirus is 10 μl to 1 ml; Preferably, the second new coronavirus is used in a volume of 50 μl.

5. The method according to claim 1, characterized in that The second novel coronavirus is infected 2 to 50 weeks after the first novel coronavirus infection; Preferably, the second novel coronavirus is infected 2 to 5 weeks after the first novel coronavirus infection; Preferably, the second novel coronavirus is infected 4 weeks after the first novel coronavirus infection; Preferably, the non-human animal model of pneumonia caused by the novel coronavirus is a moderate to severe pneumonia model; preferably, the non-human animal model of pneumonia caused by the novel coronavirus exhibits moderate to severe pneumonia at least 0.5 days after the second novel coronavirus infection; Preferably, the non-human animal model of pneumonia caused by the novel coronavirus exhibits moderate to severe pneumonia 3 days after the second novel coronavirus infection.

6. The method according to claim 1, characterized in that The non-human animal is a mammal; Preferably, the mammals include mice, rats, cattle, horses, pigs, dogs, and cats; Preferably, the mammal is selected from mice; Preferably, the mouse is a nude mouse; Preferably, the nude mice include BALB / c, NIH, NC, Swiss, 03H, and C57BL; Preferably, the nude mice are selected from BALB / c.

7. The method according to claim 6, characterized in that The method comprises using a dose of 5×10 3 TCID 50 BALB / c mice were infected with BA.5 for 4 weeks and then treated with 5×10 3 TCID 50 XBB infection was used to obtain a non-human animal model of pneumonia caused by the new coronavirus.

8. Any of the following methods: (1) A method for screening drug candidates for treating pneumonia caused by the new coronavirus, characterized in that: The method comprises: a) administering the agent to be screened to a non-human animal with pneumonia caused by the novel coronavirus constructed by the method described in any one of claims 1 to 7; b) detecting the therapeutic effect of the agent to be screened on pneumonia caused by the new coronavirus; (2) A method for evaluating the therapeutic effect of a drug for treating pneumonia caused by the new coronavirus, characterized in that the method comprises: a) administering the drug to a non-human animal with pneumonia caused by the novel coronavirus constructed by the method according to any one of claims 1 to 7; b) detecting the therapeutic effect of the drug on pneumonia caused by the novel coronavirus; (3) A method for studying the pathogenesis of pneumonia caused by the novel coronavirus, characterized in that the method comprises using a non-human animal with pneumonia caused by the novel coronavirus constructed by the construction method according to any one of claims 1 to 7 to study the pathogenesis of pneumonia caused by the novel coronavirus; Preferably, the agents to be screened include small molecule compounds, polypeptides, proteins, antibodies, vaccines, nucleic acid molecules, polysaccharides, and Chinese herbal medicine extracts; Preferably, the therapeutic effects include shortening the duration of fever, shortening the time for viral nucleic acid to turn negative, improving lung inflammation absorption, reducing the viral load in lung tissue, and reducing the expression level of inflammatory cytokines or chemokines in lung tissue.

9. Any of the following applications: (1) Use of a non-human animal model of pneumonia caused by the novel coronavirus constructed by the method described in any one of claims 1 to 7 in screening drug candidates for treating pneumonia caused by the novel coronavirus; (2) Use of the non-human animal model of pneumonia caused by the novel coronavirus constructed by the method described in any one of claims 1 to 7 in evaluating the therapeutic effect of drugs for treating pneumonia caused by the novel coronavirus; (3) Use of the non-human animal model of pneumonia caused by the novel coronavirus constructed by the method described in any one of claims 1 to 7 in studying the pathogenesis of pneumonia caused by the novel coronavirus; Preferably, the therapeutic effects include shortening the duration of fever, shortening the time for viral nucleic acid to turn negative, improving lung inflammation absorption, reducing the viral load in lung tissue, and reducing the expression level of inflammatory cytokines or chemokines in lung tissue.

10. Application of novel coronavirus Omicron BA.5 and XBB in constructing non-human animal models of pneumonia caused by novel coronavirus; Preferably, the non-human animal model refers to a non-human animal that has or exhibits characteristics of a disease or condition; preferably, the non-human animal is a mammal; Preferably, the mammals include mice, rats, cattle, horses, pigs, dogs, and cats; Preferably, the mammal is selected from mice.

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