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Cold-adapted equine influenza viruses

a technology for equine influenza and cold-adapted foals, which is applied in the field of experimentally generated cold-adapted equine influenza viruses, can solve the problems of undesirable side effects, minimal protection for horses, and current modalities that cannot be used in young foals, and achieve the effect of reducing nasal discharge and temperatur

Inactive Publication Date: 2006-06-08
DOWLING PATRICIA W +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

The present invention provides a new method for creating cold-adapted equine influenza viruses, which can be used as a therapeutic composition to protect animals from disease caused by influenza A viruses. These viruses have been genetically engineered to have specific phenotypes, such as cold-adaptation, temperature sensitivity, dominant interference, and attenuation. The invention also includes a therapeutic composition made from these viruses for the treatment of respiratory disease in animals. The cold-adapted viruses can be produced by passaging a wild-type equine influenza virus in embryonated chicken eggs and selecting viruses that stably grow and replicate at a reduced temperature. The invention also provides methods for identifying and isolating these viruses.

Problems solved by technology

This vaccine provides minimal, if any, protection for horses, and can produce undesirable side effects, for example, inflammatory reactions at the site of injection.
Furthermore, current modalities cannot be used in young foals, because they cannot overcome maternal immunity, and can induce tolerance in a younger animal.
Furthermore, these researchers noted that cold-adapted human influenza viruses, i.e., viruses that have been adapted to grow at lower than normal temperatures, tend to have a phenotype wherein the virus is temperature sensitive; that is, the virus does not grow well at certain higher, non-permissive temperatures at which the wild-type virus will grow and replicate.
Although these therapeutic compositions appear to be generally safe and effective in horses, they pose a significant danger of introducing into the environment a virus containing both human and equine influenza genes.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

example 1

[0062] This example discloses the production and phenotypic characterization of several cold-adapted equine influenza viruses of the present invention.

[0063] A. Parental equine influenza virus, A / equine / Kentucky / 1 / 91 (H3N8) (obtained from Tom Chambers, the University of Kentucky, Lexington, Ky.) was subjected to cold-adaptation in a foreign host species, i.e., embryonated chicken eggs, in the following manner. Embryonated, 10 or 11-day old chicken eggs, available, for example, from Truslow Farms, Chestertown, Md. or from HyVac, Adel, Iowa, were inoculated with the parental equine influenza virus by injecting about 0.1 milliliter (ml) undiluted AF containing approximately 106 plaque forming units (pfu) of virus into the allantoic cavity through a small hole punched in the shell of the egg. The holes in the eggs were sealed with nail polish and the eggs were incubated in a humidified incubator set at the appropriate temperature for three days. Following incubation, the eggs were cand...

example 2

[0072] Therapeutic compositions of the present invention were produced as follows.

[0073] A. A large stock of EIV-P821 was propagated in eggs as follows. About 60 specific pathogen-free embryonated chicken eggs were candled and non-viable eggs were discarded. Stock virus was diluted to about 1.0×105 pfu / ml in sterile PBS. Virus was inoculated into the allantoic cavity of the eggs as described in Example 1A. After a 3-day incubation in a humidified chamber at a temperature of about 34° C., AF was harvested from the eggs according to the method described in Example 1A. The harvested AF was mixed with a stabilizer solution, for example A1 / A2 stabilizer, available from Diamond Animal Health, Des Moines, Iowa, at 25% V / V (stabilizer / AF). The harvested AF was batched in a centrifuge tube and was clarified by centrifugation for 10 minutes at 1000 rpm in an IEC Centra-7R refrigerated table top centrifuge fitted with a swinging bucket rotor. The clarified fluid was distributed into 1-ml cryo...

example 3

[0076] A therapeutic composition comprising cold-adapted equine influenza virus EIV-P821 was tested for safety and its ability to replicate in three horses showing detectable prior immunity to equine influenza virus as follows. EIV-P821, produced as described in Example 1A, was grown in eggs as described in Example 2A and was formulated into a therapeutic composition comprising 107 pfu EIV-P821 / 2 ml BSA-MEM solution as described in Example 2C.

[0077] Three ponies having prior detectable hemagglutination inhibition (HAI) titers to equine influenza virus were inoculated with a therapeutic composition comprising EIV-P821 by the following method. Each pony was given a 2-ml dose of EIV-P821, administered intranasally using a syringe fitted with a blunt cannula long enough to reach past the false nostril, 1 ml per nostril.

[0078] The ponies were observed for approximately 30 minutes immediately following and at approximately four hours after vaccination for immediate type allergic reactio...

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Abstract

The present invention provides experimentally-generated cold-adapted equine influenza viruses, and reassortant influenza A viruses comprising at least one genome segment of such an equine influenza virus, wherein the equine influenza virus genome segment confers at least one identifying phenotype of the cold-adapted equine influenza virus, such as cold-adaptation, temperature sensitivity, dominant interference, or attenuation. Such viruses are formulated into therapeutic compositions to protect animals from diseases caused by influenza A viruses, and in particular, to protect horses from disease caused by equine influenza virus. The present invention also includes methods to protect animals from diseases caused by influenza A virus or other infectious agents utilizing the claimed therapeutic compositions. Such methods include using a therapeutic composition as a vaccine to generate a protective immune response in an animal prior to exposure to an infectious agent, as well as using a therapeutic composition as a treatment for an animal that has been recently infected with an infectious agent leading to respiratory disease, or is likely to be subsequently exposed to such an agent in a few days whereby the therapeutic composition reduces such respiratory disease, even in the absence of antibody-mediated immunity. The present invention also provides methods to produce cold-adapted equine influenza viruses, and reassortant influenza A viruses having at least one genome segment of an equine influenza virus generated by cold-adaptation.

Description

FIELD OF THE INVENTION [0001] The present invention relates to experimentally-generated cold-adapted equine influenza viruses, and particularly to cold-adapted equine influenza viruses having additional phenotypes, such as attenuation, dominant interference, or temperature sensitivity. The invention also includes reassortant influenza A viruses which contain at least one genome segment from such an equine influenza virus, such that the reassortant virus includes certain phenotypes of the donor equine influenza virus. The invention further includes genetically-engineered equine influenza viruses, produced through reverse genetics, which comprise certain identifying phenotypes of a cold-adapted equine influenza virus of the present invention. The present invention also relates to the use of these viruses in therapeutic compositions to protect animals from diseases caused by influenza viruses. BACKGROUND OF THE INVENTION [0002] Equine influenza virus has been recognized as a major resp...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): C12Q1/70C12Q1/68A01K13/00A61K39/145A61P11/00A61P31/04A61P31/12A61P31/16C12N7/08
CPCA61K39/145C12N7/00C12N2760/16134A61K2039/552A61K2039/5254A61K2039/543C12N2760/16164A61K39/12A61P11/00A61P31/04A61P31/12A61P31/16
Inventor DOWLING, PATRICIA W.YOUNGNER, JULIUS S.
Owner DOWLING PATRICIA W
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