Pharmaceutical use of favipiravir in the treatment of type midgut spore disease in bombyx mori

Abstract

The pharmaceutical use of the drug of the application is the pharmaceutical use of the broad-spectrum RNA virus drug of the application, i.e. the pharmaceutical use of the drug of the application in the treatment of the silkworm midgut type pyosis BmCPV. The drug of the application is dissolved in DMSO, and the concentration is 17.5 mg / mL. The drug of the application can inhibit the expression of the BmCPV S1 gene, and the treatment effect of the silkworm midgut type pyosis can be achieved. And the drug of the application can be continuously administered according to the dosage of the application, and the silkworm will not be negatively affected.

Description

Technical Field

[0001] This invention belongs to the fields of molecular biology and virology, specifically relating to the pharmaceutical use of the broad-spectrum anti-RNA virus drug favipiravir in the treatment of midgut septicemia (BmCPV) in silkworms. Background Technology

[0002] Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) can infect the midgut cells of silkworms, causing midgut septicemia and resulting in low silkworm yields. It poses a serious threat to sericulture production, and current prevention and control measures are limited to disinfection.

[0003] Silkworm cytoplasmic polyhedrovirus (SPCV) is an RNA virus composed of 10 independent dsRNA segments (S1–S10), encoding 6 structural (capsid) proteins (VP1, VP2, VP3, VP4, VP6, VP7) and 3 non-structural proteins. It infects the midgut cells of silkworms, causing midgut septicemia. Midgut septicemia, commonly known as "dry white belly," is primarily transmitted through ingestion. Symptoms include: silkworms consuming mulberry leaves, lethargy, often lying still on the silkworm bed, slow growth and development, significant population disparities, and in the later stages, diseased silkworms exhibit a translucent, "hollow head" appearance at the thorax, accompanied by shrinkage, diarrhea, and in severe cases, milky white mucus in the excrement. Disease progression: Infection in the 1st instar typically manifests in the 2nd–3rd instar; in the 2nd instar, in the 3rd–4th instar; in the 3rd instar, in the 4th–5th instar; and in the 4th instar, in the 5th instar. Infection in the mid-to-late 5th instar generally results in cocoon spinning, but some cocoons may be dead. The disease is characterized by its slow progression and long course, thus classifying it as a chronic infectious disease. In production, it mostly occurs in the 3rd and 4th instars, gradually spreading to large numbers after the 5th instars are fed. The lesions are mainly in the midgut; when the dorsal body wall of the silkworm is torn, milky-white folds can be seen on the midgut, and in severe cases, the entire midgut turns milky-white.

[0004] After BmCPV virus invades cylindrical cells, the pathological changes in the cells cannot be observed in the early stages of proliferation and replication. However, as the virus continues to replicate and cycle, a series of pathological changes in the infected cylindrical tissue cells can be clearly observed, such as: shedding of the peritrophic membrane, cell deformation, nuclear position shift, vacuolation of cytoplasm, cytoplasm filled with polyhedra, cell collapse, and shedding.

[0005] Currently, there are no conventional drugs for silkworm viral diseases in sericulture, making silkworm viral diseases incurable. Summary of the Invention

[0006] Technical problem to be solved: This invention provides a pharmaceutical use of favipiravir in the treatment of midgut septicemia in silkworm cytoplasmic polyhedrosis virus.

[0007] Technical solution: Application of favipiravir in the preparation of drugs for treating BmCPV infection in silkworms.

[0008] Application of favipiravir in the preparation of drugs for the prevention of BmCPV infection in silkworms.

[0009] The above-mentioned favipiravir was dissolved in DMSO at a concentration of 17.5 mg / mL.

[0010] Favipiravir treats BmCPV-infected silkworms by inhibiting the expression of the BmCPV S1 gene.

[0011] This drug is used to prevent and treat BmCPV infection in silkworms. The active ingredient is favipiravir.

[0012] Beneficial effects: This invention achieves a therapeutic effect on midgut septicemia in silkworms by inhibiting the expression of the BmCPV S1 gene with favipiravir. Furthermore, continuous administration of the drug according to the dosage specified in this invention will not have any negative effects on the silkworms. Attached Figure Description

[0013] Figure 1 This diagram illustrates the S1 gene expression levels during and after drug administration, where A represents the S1 gene expression level during drug administration and B represents the S1 gene expression level after drug administration. The results show that viral replication was significantly inhibited during drug administration after infection; however, after drug administration was stopped, viral replication not only increased but also rebounded. Therefore, it is speculated that continuous drug administration is necessary to inhibit viral replication in silkworms infected with the virus.

[0014] Figure 2 The diagram shows the detection of detoxification enzyme genes after drug administration to uninfected silkworms. The results show that after drug administration, the transcription level of only one detoxification enzyme gene, CarE8v1, increased, while the transcription levels of the other two detoxification enzyme genes decreased, but the changes were not significant, indicating that the drug had no toxic side effects on silkworms.

[0015] Figure 3 This is a schematic diagram of the detection of detoxification enzyme genes in silkworms infected with the virus but not treated with the drug. The results show that the virus infection of silkworms activated the silkworm's immune system, and therefore the transcription levels of the three detoxification enzyme genes increased significantly.

[0016] Figure 4 A schematic diagram of detoxification enzyme gene detection after drug administration to a virus-infected organism; the results show that after drug administration, due to the drug's inhibitory effect on viral proliferation, the relative transcription level of the detoxification enzyme gene in the silkworm's immune system decreased (relative to...). Figure 3 (To be continued).

[0017] Figure 5The experimental groups were compared among fifth-instar, sixth-day silkworms, with 1: virus-infected group treated with medication; 2: non-virus-infected group treated with medication; 3: virus-infected group; and 4: blank control group. The results showed that: (1) the drug had no effect on the growth and development of silkworms; and (2) after administration, compared with the untreated silkworms, the growth and development of the silkworms were significantly less affected by the virus infection. Detailed Implementation

[0018] Example 1

[0019] 1. Silkworm hatching and breeding

[0020] The silkworm variety selected is the Jingsong × Haoyue hybrid. The silkworm eggs are incubated in an incubation box at 25℃ and 85% relative humidity. After the silkworm eggs turn green, they are treated with light and then collected. After that, they are raised on mulberry leaves normally.

[0021] 2. Infection with BmCPV virus and addition of antiviral drugs

[0022] Once the silkworms reach the fourth instar, they are grouped and reared. The purified BmCPV virus particles are diluted to 1×10⁻⁶. 6 / mL, mulberry leaves were cut into identical cubes and soaked in the virus solution. The control group was soaked in water. After air-drying, each group was given the same number of leaves. 12 hours after infection, the animals were fed mulberry leaves containing antiviral drugs (the drug was dissolved and diluted in DMSO to 2.5mg / mL, evenly applied to the surface of the mulberry leaves, and then dried). Sand was removed before feeding the medicated mulberry leaves. (Antiviral drugs were given during the fourth instar period).

[0023] 3. Dissection of the midgut tissue

[0024] Midgut tissue was harvested on the third day of the fourth year and the third day of the fifth year.

[0025] 4. RNA extraction from tissues and quantitative PCR analysis

[0026] Total RNA was extracted from the midgut and reverse transcribed into cDNA. Primers for the S1 gene were designed based on the coding region sequence of BmCPV virus in GenBank, and gene expression levels were detected by quantitative PCR.

[0027] 5. Macroeconomic Data Collection and Processing

[0028] During the silkworm rearing process, the weight of silkworms before and after drug administration (i.e., the weight of silkworms starting from the fourth instar and the weight of silkworms in the fifth instar), the cocoon layer ratio of each group, the mortality rate, and the size of the cocoons were recorded.

[0029] The therapeutic effects of the drugs on each experimental group:

[0030] Table 1. Therapeutic effects of the drug of the present invention against BmCPV virus.

[0031]

[0032] Note: Blank group – not infected with the virus and not given any medication; Control group – infected with the virus but not given any medication; Experimental group – both infected with the virus and given medication; Mortality rate – number of deaths before the end of age five / (total number - number of midguts removed) × 100%

[0033] Although embodiments of the present invention have been disclosed above, they are not limited to the applications listed in the specification and embodiments. They can be applied to various fields suitable for the present invention. For those skilled in the art, other modifications can be easily made. Therefore, without departing from the general concept defined by the claims and their equivalents, the present invention is not limited to the specific details and illustrations shown and described herein.

Claims

1. Use of favipiravir in the preparation of a medicament for treating BmCPV infected silkworms.

2. Use of favipiravir in the preparation of a medicament for preventing BmCPV infected silkworms.

3. Use according to claim 1 or 2, characterized in that, The favipiravir is dissolved in DMSO at a concentration of 17.5 mg / mL.

Images