A COMBINATION VACCINE AGAINST PCV2 AND PRRS VIRUS INFECTION COMPRISING ALBUMIN

Abstract

The present invention pertains to a vaccine for use in prophylactically treating an animal against an infection with porcine circovirus type 2 (PCV2) and an infection with PRRS virus, the vaccine comprising in combination non-replicating immunogen of porcine circovirus type 2 and live attenuated PRRS virus, wherein the vaccine additionally comprises albumin.

Claims

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16. A vaccine comprising in combination a non-replicating immunogen of porcine circovirus type 2 (PCV2), a live attenuated porcine reproductive and respiratory syndrome (PRRS) virus, and albumin.

17. A vaccine of claim 16, wherein the vaccine comprises ovalbumin.

18. A vaccine of claim 16, wherein the albumin concentration is between 0.1 and 10% (w/w).

19. A vaccine of claim 16, wherein the non-replicating immunogen of PCV2 is recombinantly expressed ORF2 protein of PCV2.

20. A vaccine of claim 16, wherein the non-replicating immunogen of PCV2 is baculovirus expressed ORF2 protein of PCV2.

21. A vaccine of claim 16, wherein the vaccine comprises in addition non-replicating immunogen of Mycoplasma hyopneumoniae.

22. A vaccine for use in prophylactically treating an animal against an infection with porcine circovirus type 2 (PCV2), an infection with porcine reproductive and respiratory syndrome (PRRS) virus, or an infection of both PCV2 and PRRS virus, wherein the vaccine comprises in combination non-replicating immunogen of PCV2, live attenuated PRRS virus, and albumin.

23. A vaccine of claim 22, wherein said vaccine is administered into the dermis of the animal.

24. A vaccine of claim 22, wherein said vaccine is administered by a single dose.

25. A vaccine of claim 22, wherein said vaccine is administered with a needle-less vaccination device.

26. A vaccine of claim 22, wherein immunogen of PCV2 and the live attenuated PRRS virus are combined in the vaccine within 24 hours before administration.

27. A vaccine of claim 22, wherein immunogen of PCV2 and the live attenuated PRRS virus are combined in the vaccine within 6 hours before administration.

28. A vaccine of claim 26, wherein prior to combination of the immunogens, the albumin is present in combination with the immunogen of PRRS virus.

29. A method for prophylactically treating an animal against an infection with porcine circovirus type 2 (PCV2), an infection with porcine reproductive and respiratory syndrome (PRRS) virus, or an infection of both PCV2 and PRRS virus, by administrating to the animal a vaccine comprising in combination non-replicating immunogen of PCV2, live attenuated PRRS virus, and albumin.

30. A method of manufacturing a vaccine comprising the non-replicating immunogen of porcine circovirus type 2 (PCV2), the live attenuated porcine reproductive and respiratory syndrome (PRRS) virus, and albumin, for administration to an animal to prophylactically treat the animal against an infection with PCV2, an infection with PRRS virus, or an infection of both PCV2 and PRRS virus.

Description

EXAMPLES

[0034] Experiment 1

[0035] In a first experiment the effect of the addition of a PCV2 ORF2 subunit vaccine on the viability of a live attenuated PRRS virus vaccine was established, with or without the presence of bovine serum albumine in the final vaccine. For this the PRRS virus titer was measured (log 10 TCID50/ml) in a dilution on MA-104 cells (African green monkey kidney cells) one hour after combining the vaccines. As a control, the viability of the same PRRS virus vaccine was measured without adding the PCV2 vaccine. The results are indicated below in Table 1 for vaccines wherein the (aimed at) start titer of the PRRS virus was 4 (log 10). In the combination vaccine 0.3% (3 grams per litre vaccine) serum albumin was added. Table 2 gives the same results for vaccines wherein the (aimed at) start titer of the PRRS virus was 5 (log 10). In this latter combination vaccine also 0.3% (3 grams per litre vaccine) serum albumin was added. The results indicate that albumin has a significant effect on PRRS virus viability in the combination vaccine.

TABLE-US-00001 TABLE 1 Effect of PCV2 vaccine on PRRS virus viability Sample (PRRS start titer 4 log10) PRRS virus viability Control (PRRS virus vaccine) 3.3 PCV/PRRS, no albumin 0 PCV/PRRS, 0.3% albumin 3.0

TABLE-US-00002 TABLE 2 Effect of PCV2 vaccine on PRRS virus viability Sample (PRRS start titer 5 log10) PRRS virus viability Control (PRRS virus vaccine) 4.5 PCV/PRRS, no albumin 2.2 PCV/PRRS, 0.3% albumin 4.8

[0036] Experiment 2

[0037] In the second experiment different proteins were tested for their effect on the viability of PRRS virus in a PCV2/PRRS virus combination vaccine. In this experiment the combined PCV2/PRRS virus samples with a final concentration of (alleged) PRRS virus stabiliser of 1% (w/w), were tested for PRRS viability (starting titre of 5 log 10) as described under Experiment 1. For this, the stabilisers were mixed with the PRRS virus vaccine and thereafter the PCV2 vaccine was added. The following proteinaceous virus stabilisers (next to bovine serum albumin) were tested: [0038] Vegetable peptone (Sigma Aldrich 18332-500G-F) [0039] Vegetable peptone No 1 (Sigma Aldrich 61854-500G-F) [0040] Vegetable peptone No 2 (Sigma Aldrich 19942-500G-F) [0041] Soybean peptone (Sigma Aldrich 70178-100G) [0042] Skimmed milk (Campina, The Netherlands) [0043] Ovalbumin (Sigma Aldrich, A5253-250G) [0044] NZ-amine (casein hydrolysate; lab product)

[0045] The results are indicated here below in Table 3.

TABLE-US-00003 TABLE 3 Effect of PCV2 vaccine on PRRS virus viability Sample PRRS virus titer Control (PRRS virus vaccine) 6.4 Bovine serum albumin 6.4 Vegetable peptone 4.2 Vegetable peptone No 1 4.3 Vegetable peptone No 2 4.0 Soybean peptone 4.0 Skimmed milk 4.8 Ovalbumin 6.1 NZ-amine 4.3

[0046] It appeared that only with albumin (either of bovine or chicken egg source), the PRRS virus titer decrease could be (almost completely) prevented.

[0047] Experiment 3

[0048] Objective

[0049] The objective of this study was to evaluate efficacy and safety of PCV2/Mhyo/PRRS combination vaccines and in particular to show the effect of the addition of albumin on the PRRS efficacy. The efficacy towards protection against infection with PCV2 was evaluated by assessing anti-ORF2 serology. The efficacy against infection with Mycoplasma hyopneumoniae was evaluated by comparing the serological response with that of the commercially available Mhyo vaccine Porcilis Mhyo (MSD Animal Health, Boxmeer, The Netherlands). The efficacy against an infection with PRRS virus was evaluated by assessing serology and the PRRs viraemia upon challenge with a pathogenic PRRS strain, 4 weeks post vaccination.

[0050] Experimental Design

[0051] The progeny of 10 sows was available for this study. A total of 40 animals were allotted to 4 groups of 10 piglets each. All animals were transferred to an animal facility when they were approximately 4 weeks old. Groups 1 to 4 were intradermally vaccinated using the IDAL vaccinator into the right side of the neck. Groups 1 and 2 each received an ORF2 protein based PCV2 vaccine comprising in addition Mhyo bacterin (the same antigen as in the commercially available product Porcilis M Hyo), and 3% ovalbumin (group 1) or no albumin (group 2). In these combination vaccines a live PRRS virus vaccine (Porcilis PRRS) was reconstituted. The vaccines used Montanide IMS 251, available from SEPPIC, France as adjuvant. Each vaccine contained 9 g/dose of the ORF2 protein, and Mhyo antigen at 1-2 times the concentration of the M Hyo antigen in the commercially available vaccine Porcilis M Hyo ID ONCE. The PRRS vaccine was a freeze-dried vaccine and was reconstituted immediately before administration to contain 10.sup.4.5 TCID.sub.50 of virus per dose of 200 l using the appropriate PCV2 vaccine or a diluent. Group 3 only received the PRRS vaccine and group 4 remained unvaccinated and served as (challenge) control. All piglets were observed daily for clinical signs. The animals were challenge-infected with pathogenic PRRS virus (type I) when they were approximately 8 weeks old (day 28). The challenge material contained (a calculated dose of) 5.3 log 10 TCID50 of the virus in 2 ml. The material was intra-nasally administered, 1 ml per nostril. At the end of the observation period (49 days after vaccination corresponding to 21 days post challenge) all pigs were sacrificed. Blood samples (via v. jugularis) were taken from all animals individually on day 0, 14, 28 (right before challenge), 31, 35, 38, 42 and 49 and tested for the presence of PRRS virus, for antibodies against PRRSV, PCV2 and Mhyo.

[0052] Results

[0053] No animals showed any clinical signs due to vaccination and rectal temperatures remained within 1.5 C. from controls. The vaccines are thus regarded safe.

[0054] Regarding Mhyo, the serological response of the combination vaccine appears to be comparable to that as obtainable with the commercially available vaccine Porcilis M Hyo (no numerical results depicted in a figure). It may thus be assumed that the vaccine protects against infection with Mhyo.

[0055] The results of the PCV2 serological response are given in FIG. 1. It appears that the two combination vaccines induce a positive anti-ORF2 antibody response which means that the vaccines induce protection against infection with wild-type PCV2.

[0056] The results of the PRRS serological response are given in FIG. 2. It appears that the two combination vaccines, like the commercially available PRRS vaccine, induce a positive anti-PRRS antibody response before challenge. This is an indication that the vaccines provide protection against PRRS virus infection. It can also be seen that the serology was improved in the vaccine comprising the albumin. In FIG. 3 the viraemia data are given. These results are in line with the results of FIG. 2. It appears that all three vaccines provide protection against PRRS virus infection since viraemia levels are lower than the level in the control animals (group 4) at each point in time. Still, the combination vaccine comprising albumin provides better protection against challenge than the combination vaccine without the albumin.