RU2252253C2 - Thermosensitive strain of horse abortion herpes virus env-1, pharmaceutical composition, vaccine and method for animal immunization - Google Patents

Abstract

FIELD: virology, veterinary.

SUBSTANCE: invention relates to thermosensitive strain of horse abortion herpes virus ENV-1, pharmaceutical composition, vaccine and method for animal immunization based on the same. Strain replication is limited by upper respiratory tract and doesn't cause of viremia. Furthermore strain has insufficient growth at temperature of wild-type virus normal growth. Strain of present invention, as well as compositions and vaccines based on the same are useful in clinical and viral horse protection against ENV-1 infections, viral induced abortion and paresis.

EFFECT: new pharmaceutical composition, vaccine and method for horse immunization.

4 cl, 10 tbl, 7 ex

Description

DESCRIPTION

The present invention relates to a mutant horse abortion virus, to a method for producing said mutant, to using said mutant, and to live vaccines derived from this mutant.

Horse Abortion Virus (EHV-1), herpes virus, is the main horse pathogen responsible for virus-induced abortion, a neurological disease such as paresis, upper respiratory tract infections, and newborn foal diseases (BNF). BNGs are caused by a transplacental infection of the fetus that occurs in the near term of delivery, while the foal is born weak with a severe respiratory disease, and sometimes with jaundice, caused by infection of the liver with the EHV-1 virus. These animals usually die a few days after birth. Horse Rhinopneumonitis Virus (EHV-4) is the leading cause of acute respiratory infections (“rhinopneumonitis”) and causes infections in most horses during the first two years of their life. Rhinopneumonitis is characterized by a febrile condition, anorexia and abundant discharge from the nose, which later turn into purulent discharge. In rare cases, EHV-4 infection causes abortion in mares. In addition, EHV1 and EHV4 lead to the establishment of persistent long-lived latent infections. After reactivation, these viruses cause a recurrent disease, accompanied by the isolation of the virus and its transfer to other animals.

The fight against herpes virus infection in horses and their diseases, in particular, with EHV-1-mediated abortion of horses, paresis and diseases of newborn foals caused by transplacental infection of the fetus that occurs in the near term, does not give satisfactory results. Although inactivated as well as modified live vaccines are available, it is obvious that none of these vaccines blocks the infection sufficiently and does not prevent the occurrence of latent infection caused by the wild-type virus. Therefore, there is an urgent need for safe vaccines with improved protective functions against field infections caused by these viruses, and in particular against infections caused by EHV1.

The present invention relates to such vaccines.

In its first aspect, the present invention relates to the mutant Ts EHV-1, deposited in the European collection of animal cell cultures (ECACC), Salisbury, Wiltshire SP4 ОJG, UK, June 10, 1999 under accession number V99061001, and to his offspring.

Mutants Ts EHV-1 of the present invention, in addition, are phenotypically characterized in that:

they form small plaques when they grow on several horse cell lines,

they lose their ability to grow on rabbit kidney cells, and in particular on RK13 cells and

they have a limited ability to cause viremia.

Mutants Ts EHV-1 of the present invention have the advantage that their replication is limited to the upper respiratory tract of an ordinary horse, without causing viremia or causing limited viremia. Ts mutants are safe for mares, but at the same time they provide significant immune stimulation after breeding in the upper respiratory tract. Ts mutants do not easily pass from animal to animal, which limits the possibility of their transmission and reversal.

In accordance with the present invention, the term “offspring” also applies to all strains obtained by subsequent serial passage of the deposited mutant Ts EHV-1.

According to the present invention, a temperature-sensitive mutant is defined as a mutant virus that detects insufficient growth at a temperature close to or near it, at which the wild-type virus detects normal growth. Mutants Ts EHV-1 of the present invention are characterized in that they are temperature sensitive at the body temperature of the animal. Mutants Ts EHV-1 of the present invention are not replicated at temperatures above 38.5-39.0 ° C. Mutants Ts EHV-1 of the present invention are preferably not replicated at a temperature of 38.5 ° C.

In accordance with the present invention, the term “small plaques” means plaques that are half or one third the size of the plaques formed by the parent wild-type strain in horse cells.

In accordance with the present invention, the expression “limited ability to cause viremia" means the ability not to induce or induce in some animals a low (that is, only detectable) viremia level for 1-3 or 4 days compared with the ability of the parent strain to cause viremia.

The temperature-sensitive EHV-1 mutants of the present invention can be obtained by treating cow, horse or other permissive cell cultures at 34 ° C. with non-toxic concentrations of mutagens, such as 5-bromo-2-deoxyuridine, azacytidine and the like, during virus replication in vitro followed by biological cloning of the offspring of the virus from these treated cultures in bovine, equine or other permissive cell lines.

The favorable properties of the Ts mutants of the present invention make them highly suitable for the preparation of a vaccine. Thus, in a second aspect, the present invention relates to a composition, and in particular, to a vaccine composition comprising the EHV-1 Ts mutant of the present invention and a pharmaceutically acceptable carrier or excipient. More specifically, the (vaccine) composition of the present invention includes the Ts mutant EHV1 deposited in (ECACC), Salisbury, UK, under accession number V99061001, and / or its progeny. Pharmaceutically acceptable carriers or excipients suitable for use in the vaccine of the present invention are sterilized water, saline, aqueous buffers such as PBS and the like. In addition, the vaccine of the present invention may contain other additives, such as adjuvants, stabilizers, antioxidants, and the like.

The vaccine compositions of the present invention are safe and can be used for clinical and virological protection of horses against EHV-1 infections and for protection against virus-induced abortion and paresis. In addition, it was found that the vaccine of the present invention kills transplacental infection and thus protects newborn foals from disease. The vaccine composition of the present invention can be administered not only to horses, but also to other animals susceptible to EHV-1 infection, such as donkeys, zebras, and the like. It is contemplated that cattle that are susceptible to EHV-1 and EHV-4 infections can be treated with the vaccine of the present invention.

In addition, it was unexpectedly discovered that vaccines containing the Ts-mutant EHV-1 of the present invention not only protect against EHV-1 infections, but also against diseases and their associated virus isolation after infection with the EHV-4 virus. Thus, said vaccine can be used to cross-protect vaccinated horses. These vaccines provide improved protection by effectively blocking infection with wild-type viruses.

Vaccine compositions of the present invention can be obtained by standard methods. A preferred vaccine of the present invention is a live vaccine. To obtain a live vaccine, the source EHV Ts mutant virus can be cultured in a cell culture, such as primary or secondary cow kidney cells or horse cells. Viruses cultured in this way can be harvested by collecting liquid cultures of tissue cells and / or cells. During the collection process, the output of these viruses can be, but not necessarily, stimulated by methods that facilitate the release of infectious particles from the growth substrate, for example, by sonication. The specified live vaccine can be obtained in the form of a suspension, or it can be lyophilized.

Pharmaceutically acceptable carriers that can be used in the vaccine of the present invention are sterilized water, saline, aqueous buffers such as PBS and the like. In addition, the vaccine of the present invention may contain other additives, such as adjuvants, stabilizers, antioxidants, and the like.

Suitable stabilizers are, for example, carbohydrates, including sorbitol, mannitol, starch, sucrose, dextran and glucose, proteins and their decomposition products, including, but not limited to, albumin and casein, protein-containing agents such as bovine serum or separated milk, and buffers, including, but not limited to, alkali metal phosphates. In lyophilized vaccine compositions, it is preferable to add one or more stabilizers.

Suitable adjuvants include, but are not limited to, hydroxide, phosphate or alumina, amphigen, tocopherols, monophosphenyl lipid A, muramyl dipeptide, oil emulsions, glucans, carbomers, block copolymers, cytokines and saponins such as Quil A. Amount the adjuvant to be added depends on the nature of this adjuvant.

The EHV-1 Ts mutants of the present invention are preferably administered to ordinary seronegative animals from a few days to several years of age, including mares. Said vaccine can be administered to animals by non-parenteral routes of administration, including, but not limited to, oral administration, spray administration, aerosol administration, intraocular and intranasal administration. Alternatively, said vaccine may be administered by parenteral routes of administration. Said vaccine is preferably administered subcutaneously or intranasally.

Basically, the Ts-mutant EHV-1 virus is administered in an amount effective to induce protection against EHV-1 infection. The dose usually depends on the route of administration, the time of administration, and also on the age, state of health and diet of the vaccinated animal. The specified virus can be introduced in an amount of from 10 ² to 10 ⁹ , preferably from 10 ³ to 10 ⁵ bfu / dose, and more preferably 10 ² bfu / dose per animal.

The vaccines of the present invention can also be administered simultaneously or alternately with other live or inactivated vaccines. These additional vaccines may be administered non-parenterally or parenterally. These additional vaccines are preferably recommended for parenteral administration.

Examples

1. Isolation and evaluation of the temperature-sensitive mutant strain TS C147 EHV-1

Almost confluent one-day 75 cm ² monolayers of horse skin cells were infected with a multiplicity of infection (MH) of 0.001 EHV-1. The inoculum (2.0 ml) was adsorbed (1 hour, 37 ° C), removed and the monolayers washed with PBS, and then re-fed with tissue culture medium (25 ml) containing 40 μg / ml 5-bromo-2-deoxyuridine, and incubated at 34 ° C. At maximum CPD (7 days after inoculation), the culture was harvested (frozen at -40 ° C and then thawed at 37 ° C), dialyzed overnight against PBS at 4 ° C, titrated to EHV-1 infection in ED cells at 37 ° C, and then cloned at 34 ° C in ED cells cultured in 96-well microtiter plates. Wells with one EHV-1 lesion were identified, left for cultivation to maximum CPD, and then a small (20 μl out of the total 200 μl) sample was used for phenotypic typing at permissive or limiting temperatures using ED cells. Then the temperature-sensitive clones were passaged on JCK line (Jay's Calf Kidney-Intervet) on the cells of the calf kidneys to obtain the initial and working seed.

2. Temperature-sensitive phenotype of strain TS C147 EHV-1

Strain TS C147 EHV-1 at the level of the initial vaccine virus (MSV) + 1 ° was titrated in parallel in the cells of a bovine lung embryo (BEL26-Intervet line), in calf kidney cells, JCK line (Jay's Calf Kidney-Intervet), in horse skin cells (ED), and in cells of the clone W7 C5 horse skin (line ED-W7 C5-Intervet) at 37 ° C and 38.5 ° C. The virus at the level of (MSV) + 1 ° passage did not show growth at 38.5 ° C. The results are presented in table 1.

3. The TS C147 EHV-1 strain has EHV-4-like characteristics

The criterion for differentiation between EHV-1 and EHV-4 is their ability to replicate in rabbit kidney cells (RK13). EHV-1 strains replicate well in RK13 cells, however, these cells are immune to EHV-4 strains. Strain TS C147 EHV-1 at the level of (MSV) + 1 °, its parent strain of wild-type EHV1, strain EHV-1 deficient in the early gene (IE EHV-1), pathogenic strain CHLi EHV-1 and field isolate EHV-4 titrated in parallel at 37 ° C in RK13 cells and in horse skin cells (ED). The results presented in table 2 showed that 4 strains of EHV-1, including strain TS C147 EHV-1 and strain EHV-4 are replicated in ED cells, however, strain TS C147 EHV-1 and strain EHV-4 do not show growth in RK13 cells.

Table 1. Relative titers of strain TS C147 EHV-1 at 37 ° C and 38.5 ° C in different cell lines of cows and horses

Titers ^a) (log ₁₀ TCID ₅₀ / ml) at 37 ° C and 38.5 ° C in various types of cow and horse cells The passage level of the virus TSC 147 (MSV) + 1 ° BEL: 37 ° C
38.5 ° C 5.2
<1.1 JCK: 37 ° C
38.5 ° C 5,4
<1.1 ED: 37 ° C
38.5 ° C 5,4
<1.1 ED W48 C10: 37 ° C
38.5 ° C 5.7
<1.1 ED W7 C5: 37 ° C
38.5 ° C 5.7
<1.1

a) Titers after 5-day incubation; the titers, represented as <1.1 log ₁₀ TCID ₅₀ / ml, did not produce foci of EHV-1 detected at the lowest dilution of 4 × 200 μl (10 ^-1 ) of the virus tested by titration.

Table 2. The ability of rabbit kidney cells to support the replication of strain TS C147 EHV-1

Virus Relative titer (log ₁₀ TCID ₁₀ / ml) at 37 ° C in RK13 and ED cells RK13 cells ED cells EHV-1TS C147 MSV + 1 ° <1.1 ^a 5.7 EHV-1,040 5.7 5.7 EHV-1 IE ^- 6.0 6.2 EHV-1 CHLi 5.7 6.0 Ehv-4 <1.1 3,7

a) Titers represented as <1.1 log ₁₀ TCID ₅₀ / ml did not give the EHV-1 CPD detected at the lowest dilution of 4 × 200 μl (10 ^-1 ) upon titration.

4. Clinical and virological protection of common ponies against EHV-1 and EHV-4 infections

Of 29 ordinary ponies with weakly EHV-1-neutralizing or non-neutralizing EHV-1 antibody (BH), 15 animals were vaccinated intranasally (in) with a dose of 5.3 log ₁₀ TCID _{50 of} TS C147 EHV-1 strain, and 14 ponies were left unvaccinated as an unvaccinated control. 1 month after one vaccination (in), 8 vaccinated and 8 unvaccinated (control) ponies were infected (in) with the field strain EHV-1, and a group of 7 EHV-1 vaccinated and 6 control animals were infected (in) with fresh field isolate EHV-4. After vaccination and infection, the animals were continuously monitored for clinical reactions, virus excretion into nasal mucus, infected white blood cells (viremia) and EHV-1 neutralizing antibodies.

In 11/15 ponies, the vaccine virus propagated to low titers (maximum titers in nasal mucus were 1.5-3.0 log ₁₀ TCID ₅₀ / ml) for 1-8 days, and in 7 of 15 animals for 1-4 days, he also led to viremia with a low white blood cell count (only detectable). However, all 15 ponies were seroconverted. In contrast, EHV-1 was not isolated from nasal mucus or blood from 14 control ponies observed daily for 10 or 14 days, respectively, and these animals remained seronegative for EHV-1 after infection. A similar level of hyperthermia was observed in 10 animals in each of the two (vaccinated and control) groups. These results are systematized in table 3.

After intranasal EHV-1 infection, a significant decrease in the release of the virus into the nasal mucus was observed in vaccinated ponies compared with the release of the virus in control animals. Similarly, a single vaccination prevented leukocyte viremia in 7 of 8 ponies, while one pony was almost virus-positive for 1 day. In contrast, almost all 8 unvaccinated ponies experienced viremia, 7 ponies for 3-4 days, and 1 pony 1 day. All 8 control ponies had a moderate to high fever for 1-6 days, and all 8 vaccinated animals remained healthy. None of the 8 vaccinated animals showed an anamnestic response to infection, while all 8 control animals showed a response with the formation of a significant amount of EHV-1 neutralizing antibody. These results are systematized in table 4.

After intranasal infection with EHV-4, the virus was isolated from nasal mucus in one of 7 vaccinated ponies in one case, while in all 6 control ponies, the virus was isolated at a fairly high titer on day 2-3, with the exception of only one pony. None of the 7 EHV-1 vaccinated ponies experienced viremia, in contrast to 3 of the 6 control ponies who experienced viremia on days 1–3. EHV-4 infection led to hyperthermia in 3 of 6 control animals on day 2-3, but none of the 7 vaccinated ponies had this hyperthermia. 4 out of 7 vaccinated animals and 5 out of 6 control animals showed a slight increase in respiratory rate (15-20 exhalations / minutes) for 1-3 and 2-6 days, respectively. These results are systematized in table 5.

Table 3:
Post vaccination results
Result - Amount + ve / total amount (range of maximum activity and duration) Parameter Vaccinated Control The exit of the virus into the mucus 11/15 (1.5-3.0 log ₁₀ TCID ₅₀ / ml, 1-8 days) 0/14 Leukocyte viremia 7/5 (low 1-4 days) 0/14 Seroconversion 15/15 0/14 Hyperthermia (= 38.5 ° C) (between 1 and 10 days) 10/15 (38.5-39.3, 1-3 days) 10/14 (38.5-38.8, 1-3 days)

Table 4:
After infection with EHV-1
Result - Amount + ve / total amount (range of maximum activity and duration) Parameter Vaccinated Control The exit of the virus into the mucus 5/8 (1.5 log ₁₀ / ml, 1-2 days) 8/8 (2.2-3.4 log ₁₀ TCID ₅₀ / ml, 4-6 days) Leukocyte viremia 1/8 (low, 1 day) 8/8 (3-4 days; 1 day for one animal The production of antibodies against EHV-1 (VN) 0/8 (= 4x magnification) 8/8 Hyperthermia 0/8 8/8 (38.9-41.0,
1-6 days) Breath 3/8 (15-20 exhalations / min, 1 day) 2/8 (15-20 exhalations / min, 1 day)

Table 5:
Results after EHV-1 infection
Result - Amount + ve / total amount (range of maximum activity and duration) Parameter Vaccinated Control The exit of the virus into the mucus 1/7 (1.5 log ₁₀ / ml, 1 day) 6/6 (1.5-3.7 log ₁₀ TSID ₅₀ / ml, 1-3 days) Leukocyte viremia 0/7 3/6 (1-3 days) The production of antibodies against EHV-1 (VN) 1/7 (= 4x magnification) 5/6 Hyperthermia 0/7 3/6 (38.6-38.8, 2-3 days) Breath 4/7 (15-20 exhalations / min, 1-3 days) 5/6 (15-20 exhalations / min, 2-6 days)

Protecting horses from paresis and abortion caused by EHV-1 infection

Of the 12 mares with weakly EHV-1 neutralizing or generally non-neutralizing EHV-1 antibody (VN), 6 animals were vaccinated intranasally (in) for approximately 6 months of pregnancy, and then all 12 mares were infected (in) with a pathogenic strain of EHV -1 at the stage of pregnancy, which is critical for EHV-1-induced abortion, namely around the 9th month of pregnancy. After vaccination, the animals were continuously monitored for clinical reactions, virus excretion into nasal mucus, infected white blood cells (viremia) and an EHV-1 neutralizing antibody.

Although the vaccine virus was not isolated from nasal mucus in any of the 6 vaccinated mares, however, mild (temporary) viremia (1-3 days) was detected in 5-6 mares, and seroconversion was observed in all 6 animals with the formation of a significant level of VN antibody against EHV-1. None of the 6 control mares that were observed in parallel with vaccinated animals for 10-14 days showed no release of EHV-1 from nasal mucus or leukocytes, however, in one of 6 animals, after about 2.5 months, was observed seroconversion. These results are systematized in table 6.

After infection, a significant (2 out of 6 compared to 5 out of 6 and 1.5-1.7 (1.5-3.0 log ₁₀ TCID ₅₀ / ml, within 1-2 days compared to 245-3, 7 log ₁₀ TCID ₅₀ / ml, within 1-6 days) decreased virus output into the nasal mucus in vaccinated mares. Similarly, none of the 6 vaccinated mares showed viremia, while 5 of the 6 unvaccinated control mares had viremia. In the control group, 5 out of 6 mares showed high fever for 1-5 days, 3 mares also developed paresis, accompanied by severe jaundice, and 2 mares had there was disintegration of the mucous plug in the cervix with signs of miscarriage. One of the two animals died and the second animal was urgently euthanized. Both animals had dead foals. Three mares had abortions. Fetal tissue from all 5 embryos was positive on EHV -1 In contrast, all 6 vaccinated mares had healthy foals.The clinical response observed only in vaccinated mares was short-term (1 day) hyperthermia in one of the 6 mares. The control mare, which had a normal foal, actually detected seroconversion immediately before infection. These results are systematized in table 7.

Table 6:
Post vaccination results
Result - Amount + ve / total amount (range of maximum activity and duration) Parameter Vaccinated (group 1) Control (group 2) ^a The exit of the virus into the mucus 0/6 Not observed Leukocyte viremia 5/6 (low,
1-3 days) 0/6 Hyperthermia 1/6 (1 day) Not watched ^a Gray conversion 6/6 (month after vaccination) 1 * / 6 (approximately 3 months after vaccination)

^a = not observed, as the animals were kept isolated from the vaccinated group.

Table 7: Results after infection

Result - Amount + ve / total amount (range of maximum activity and duration)

Parameter Vaccinated (group 1) Control (group 2) The exit of the virus into the mucus 2/6 (1.5-1.7 log ₁₀ TCID ₅₀ / ml, 1-2 days) 5/6 (2.4-3.7 log ₁₀ TCID ₅₀ / ml, 1-6 days) Leukocyte viremia 0/6 5/6 The production of antibodies against EHV-1 (VN) 0/6 5/6 + Hyperthermia 1/6 (1 day) 5/6 (1-5 days) Paresis 0/6 3/6 (fatal in two mares) Jaundice 0/6 2/6 Death 0/6 2/6 (1 mare died, 1 mare with severe paralysis, jaundice and a rapid decrease in body temperature was subjected to urgent euthanasia, Abortion 0/6 5/6

+ The control mare showed a seronegative result when taking blood samples 3 months after vaccination of group 1 mares, but showed seroconversion before infection.

6. Safety TS C147 EHV-1 for mares

Four mares at approximately 9 months of gestation (critical stage for EHV-1 abortion) were naturally inoculated with a 10-fold protective dose and observed for abortion. The results presented in table 8 showed that all 4 mares showed EHV-1 seroconversion, and one of the 4 mares showed short-term viremia, however, the birth went well. Three of the 4 foals showed negative results for virus-neutralizing (BH) antibodies against EHV-1 in blood samples taken before feeding by their mother, while one foal gave a positive result for the BH antibody, which was due to colostrum (since he was born in the interval between observations in the early hours). These results are systematized in table 8.

Table 8:
Overdose safety for pregnant mares during pregnancy, which is critical for abortions caused by EHV-1 Mare No. Exit to nasal mucus Viremia BH antibody against EHV-1 at dose and after 3 weeks Childbirth and antibody ^a thirteen c -ve <2.0 3,5 Normal <2.0 14A from + ve (3 days) <2.0 6.0 Normal 4.0 ^b fifteen from -ve <2.0 5,0 Normal <2.0 fifteen from -ve <2.0 6.0 Normal <2.0

a = EHV-1 neutralizing antibody at birth;

b = born between monitoring intervals in the early hours and blood samples were taken from this foal at least 3 hours after birth;

c = studies are incomplete, that is, ongoing.

7. Lack of transmission of TS C147 EHV-1 between target species

Studies on the transmission of the virus from one animal to another have been carried out in EHV-naive (all types) weaning foals (foals that lack a specific pathogen, SPF foals).

Two SPF foals were inoculated intranasally (in) with a 10-fold protective dose of TS C147 EHV-1 strain at the passage level of the original vaccine virus + 1 °, and virus-positive nasal mucus collected over several days was used to similarly infect another pair of SPF foals . After inoculation (i.n.), the foals were observed for (i) virus isolation in the nasal mucus, (ii) clinical reactions and (iii) EHV-1 seroconversion.

In foals inoculated with TS C147 EHV-1 at (MSV) + 1 °, virus was secreted into nasal mucus, and they were all seroconverted. However, a pool of virus-positive nasal mucus samples did not infect another pair of EHV-naive foals, as indicated by the absence of EHV-1 isolation from their nasal mucus and the absence of EHV-1 seroconversion. These results were confirmed by a study using another 4 SPF foals, 2 of which were inoculated (MSV) + 1 °, and 2 more foals were injected with virus-positive nasal mucus taken from the first two foals. The results are systematized in tables 9 and 10.

Table 9:
Transmission of TS C 147 EHV-1 strain to “EHV naive foals”
Passage one: Foals 1 and 2 were intranasally inoculated with TS C147 EHV-1 strain (10-fold protective dose) at (MSV) + 1 ° Parameter Result (+/-, interval and duration) Isolation of the virus into nasal mucus 2/2 (1.7-5.0 log ₁₀ TCID ₅₀ / ml, 5-6 days) EHV-1 seroconversion 2/2 (took blood 2 weeks after inoculation and CF-test) Passage two: foals 5 and 6 were intranasally inoculated with virus-positive nasal mucus taken from foals 1 and 2 Parameter Result (+/-, interval and duration) Isolation of the virus into nasal mucus 0/2 Seroconversion 0/2

Table 10:
Transmission of TS C147 EHV-1 strain to EHV-naive foals
Passage one: Foals 7 and 8 were intranasally inoculated with TS C147 EHV-1 strain (10-fold protective dose) at (MSV) + 1 ° Parameter Result (+/-, interval and duration) Isolation of the virus into nasal mucus 2/2 (1.5-3 / 7 log ₁₀ TSID ₅₀ / ml, 4-8 days) EHV-1 seroconversion 2/2 (took blood 2 weeks after inoculation and CF-test) Passage two: foals 9 and 10 were intranasally inoculated with virus-positive nasal mucus taken from foals 7 and 8 Parameter Result (+/-, interval and duration) Isolation of the virus into nasal mucus 0/2 EHV-1 seroconversion 0/2

Claims (4)

1. Temperature-sensitive (Ts) strain of horse abortion virus (EHV-1), deposited in ECACC under accession number V99061001, designed to prevent and / or treat EHV-1 infections in equines. 2. A pharmaceutical composition comprising a temperature-sensitive strain according to claim 1 and a pharmaceutically acceptable excipient or carrier. 3. A vaccine for the prevention and / or treatment of equine EHV-1 infections, comprising the temperature-sensitive strain of claim 1 and a pharmaceutically acceptable carrier or diluent. 4. A method of immunizing an animal to protect against EHV-1 infections, comprising administering to said animal a vaccine according to p.