Application of FB23-2 in inhibition of PEDV or TGEV cell replication

Abstract

The invention discloses an application of FB23-2 in inhibition of PEDV (porcine epidemic diarrhea virus) or TGEV (transmissible gastroenteritis virus) cell replication, relates to the technical field of preventive veterinary medicine, and has the technical key points that the FB23-2 is used for inhibiting the replication of PEDV or TGEV (transmissible gastroenteritis virus) in cells; experiments show that the FB23-2 can efficiently inhibit the replication of PEDV and TGEV in cells, the concentration of the FB23-2 is controlled to be 2.5-25 [mu] M, the inhibition effect is optimal, and the FB23-2 can be used for preventing and controlling PEDV and TGEV viruses and has important significance for related scientific researches.

Description

Technical Field

[0001] The present invention relates to the technical field of preventive veterinary medicine, and in particular to application of FB23-2 in inhibiting PEDV or TGEV cell replication. Background Art

[0002] Porcine epidemic diarrhea virus (PEDV) and porcine transmissible gastroenteritis virus (TGEV) are both members of the alphacoronavirus genus of the Coronaviridae family. They are enveloped single-stranded positive-sense RNA viruses that are widely present in nature and artificial environments. Their diameter is about 80 to 150 nm, and the full genome length is about 28 kb. PEDV and TGEV mainly harm newborn piglets within 7 days of age. The clinical symptoms are very similar, mainly manifested as diarrhea, vomiting, and dehydration. The mortality rate of newborn piglets infected with the above two viruses can reach more than 95%, which is an important epidemic disease that hinders the healthy development of the pig industry. Vaccine immunization is currently the main means of preventing and controlling PEDV and TGEV in the pig farming industry.

[0003] At present, the PEDV and TGEV vaccines on the market are both live attenuated vaccines and inactivated vaccines, and no other vaccines such as subunit vaccines have been approved for marketing. However, the above viruses mainly harm newborn piglets, and newborn piglets do not have a sound immune system. Vaccination cannot stimulate humoral immunity and cellular immunity, so the effect is not good. In addition, maternal antibodies stimulated by immunizing sows can only be transferred to piglets through breast milk, with low transfer efficiency and poor protection effect.

[0004] To this end, the present invention aims to provide the use of FB23-2 in inhibiting PEDV or TGEV cell replication to solve the above problems. Summary of the invention

[0005] The purpose of the present invention is to solve the above problems and provide the use of FB23-2 in inhibiting the replication of PEDV or TGEV cells. FB23-2 can effectively inhibit the replication of PEDV or TGEV in cells.

[0006] In order to achieve the above object, the technical solution of the present invention is as follows:

[0007] The present invention provides the use of FB23-2 in inhibiting the replication of PEDV or TGEV cells. The FB23-2 can inhibit the replication of PEDV and TGEV in cells. When the concentration of the FB23-2 is 1 μM to <50 μM, the replication of PEDV in cells is inhibited. When the concentration of the FB23-2 is 1 μM to <50 μM, the replication of TGEV in cells is inhibited.

[0008] The chemical formula of FB23-2 is C 18 H 15 Cl2N3O3.

[0009] Compared with the prior art, this solution has the following beneficial effects:

[0010] The present invention finds that FB23-2 can be used to inhibit the replication of porcine epidemic diarrhea virus (PEDV) or porcine transmissible gastroenteritis virus (TGEV) in cells, and FB23-2 can be used to prevent and control PEDV and TGEV viruses, which is of great significance to related scientific research. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1 : is a bar graph of PEDV replication and a curve graph of cytotoxicity in an embodiment of the present invention, wherein A is a bar graph of different concentrations of FB23-2 inhibiting PEDV replication; B is a curve graph of different concentrations of FB23-2 having no significant cytotoxicity;

[0012] Figure 2 FB23-2 inhibits PEDV replication in an embodiment of the present invention, indicating that different concentrations of FB23-2 can inhibit PEDV replication and reduce the number of plaques;

[0013] Figure 3 : is a bar graph of TGEV replication and a curve graph of cytotoxicity in an embodiment of the present invention, wherein A is a bar graph of different concentrations of FB23-2 inhibiting TGEV replication; B is a curve graph result of different concentrations of FB23-2 having no significant cytotoxicity;

[0014] Figure 4 This is a graph showing the results of FB23-2 inhibiting TGEV replication in an embodiment of the present invention, indicating that different concentrations of FB23-2 can inhibit TGEV replication and reduce the number of plaques;

[0015] Figure 5 It is the structural formula of FB23-2 in the embodiment of the present invention. DETAILED DESCRIPTION

[0016] In order to enable those skilled in the art to better understand the solution of the present invention, the technical solution of the present invention will be further described in detail below in conjunction with the embodiments of the present invention and the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work should fall within the scope of protection of the present invention.

[0017] It should be noted that, in the absence of conflict, the embodiments and features of the embodiments of the present invention can be combined with each other. The present invention will be described in detail below in conjunction with the embodiments.

[0018] Embodiment 1:

[0019] The experiment showed that when the concentration was less than 50 μM, different concentrations of FB23-2 could inhibit the replication of PEDV, and there was no significant effect on cell activity after treatment with FB23-2.

[0020] PEDV was inoculated into Vero cells at an MOI of 0.01. After 1 hour of infection, the unadsorbed virus was washed away. Then FB23 or DMSO was added to the culture medium to make the final concentration of FB23 0μM, 1μM, 2.5μM, 10μM and 50μM, respectively. The culture medium was collected at 12 hours, 36 hours and 60 hours after infection, and the TCID 50 The method was used to determine the virus content. At the same time, control cells that were not infected with PEDV but added with FB23-2 were set up for the determination of cell activity.

[0021] The results showed that 12 hours and 36 hours after PEDV infection, the virus titers of the 2.5μM, 10μM and 50μM FB23-2 treatment groups were significantly lower than those of the 0μM FB23-2 control group, indicating that FB23-2 can inhibit PEDV replication by 5-10 times ( Figure 1 A). This indicates that FB23-2 inhibits PEDV replication ( Figure 1 A). Cell viability was detected in FB23-2 treated cells without PEDV infection. The CCK-8 test results showed that there was no significant difference in cell viability after cells were treated with 0.1-10 μM FB23-2, but the cell viability decreased significantly after treatment with FB23-2 at concentrations above 50 μM ( Figure 1 B).

[0022] Vero cells were infected with PEDV, and the unabsorbed virus was washed away 1 hour after infection. Then, 1% methylcellulose containing different concentrations of FB23-2 was added and cultured at 37°C for 48 hours. After removing the methylcellulose, the cells were stained with 1% crystal violet to observe the size and number of plaques.

[0023] The results showed that a small number of pinpoint plaques appeared in cells infected with PEDV without FB23-2 concentration; the number of plaques in PEDV infected cells treated with 2.5-10μM FB23-2 was reduced ( Figure 2 ). The above results fully demonstrate that FB23-2 significantly inhibits the infection and replication of PEDV, and the effect increases with the increase of FB23-2 concentration, showing a dose-dependent manner.

[0024] Embodiment 2:

[0025] Experiments showed that when the concentration was less than 100 μM, different concentrations of FB23-2 could inhibit the replication of TGEV, and there was no significant effect on cell activity after treatment with FB23-2.

[0026] PK-15 cells were inoculated with TGEV at an MOI of 0.01. The virus was washed away 1 hour after infection. Then FB23-2 or DMSO was added to the culture medium to make the final concentrations of FB23-2 1μM, 2.5μM, 10μM and 50μM, respectively. The culture medium was collected at 12 hours, 36 hours and 60 hours after infection and the TCID 50 The method was used to determine the virus content. At the same time, control cells that were not infected with TGEV but added with FB23-2 were set up for the determination of cell activity.

[0027] The results showed that at 12, 36 and 60 hours after TGEV infection, the virus titers of the 0.1μM, 2.5μM, 10μM and 50μM FB23-2 treatment groups were significantly lower than those of the 0μM FB23-2 control group, with the difference reaching 5-10 times, indicating that FB23-2 can inhibit TGEV replication ( Figure 3 A). Cell viability was detected in FB23-2 treated cells without TGEV infection. The CCK-8 test results showed that there was no significant difference in cell viability after cells were treated with 1-50μM concentration of FB23-2, but after treatment with 100μM concentration of FB23-2, cell viability decreased significantly ( Figure 3 B)

[0028] PK-15 cells were infected with TGEV, and the unadsorbed viruses were washed away 1 hour after infection. Then, 1% methylcellulose containing different concentrations of FB23-2 was added and cultured at 37°C for 48 hours. After removing the methylcellulose, the cells were stained with 1% crystal violet to observe the size and number of plaques.

[0029] The results showed that a large number of pinpoint plaques appeared in cells infected with TGEV without FB23-2 concentration; the number of plaques in TGEV infected cells treated with 2.5-10μM FB23-2 was reduced ( Figure 4 ). The above results fully demonstrate that FB23-2 significantly inhibits the infection and replication of TGEV, and the effect increases with the increase of FB23-2 concentration, showing a dose-dependent manner.

[0030] In summary, FB23-2 is effective in inhibiting the replication of PEDV and TGEV in cells and can be used as an antiviral drug.

[0031] The above specific embodiments are merely explanations of the present invention and are not limitations of the present invention. After reading this specification, those skilled in the art may make modifications to the embodiments without any creative contribution as needed. However, such modifications are protected by the patent law as long as they are within the scope of the claims of the present invention.

Claims

1. The use of FB23-2 in inhibiting PEDV or TGEV cell replication, characterized by: The FB23-2 can inhibit the replication of PEDV and TGEV in cells.

2. The use of FB23-2 as claimed in claim 1 in inhibiting PEDV or TGEV cell replication, characterized in that: The chemical formula of FB23-2 is C 18 H 15 Cl2N3O3.

3. The use of FB23-2 as claimed in claim 1 in inhibiting PEDV or TGEV cell replication, characterized in that: The FB23-2 inhibits the replication of PEDV in cells at a concentration of 1 μM to <50 μM.

4. The use of FB23-2 as claimed in claim 1 in inhibiting PEDV or TGEV cell replication, characterized in that: The FB23-2 inhibits the replication of TGEV in cells at a concentration of 1 μM to <50 μM.

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