Comparative efficacy of two next-generation Rift Valley fever vaccines.

Rift Valley fever virus (RVFV) is a re-emerging zoonotic bunyavirus of the genus Phlebovirus. A natural isolate containing a large attenuating deletion in the small (S) genome segment previously yielded a highly effective vaccine virus, named Clone 13. The deletion in the S segment abrogates expression of the NSs protein, which is the major virulence factor of the virus. To develop a vaccine of even higher safety, a virus named R566 was created by natural laboratory reassortment. The R566 virus combines the S segment of the Clone 13 virus with additional attenuating mutations on the other two genome segments M and L, derived from the previously created MP-12 vaccine virus. To achieve the same objective, a nonspreading RVFV (NSR-Gn) was created by reverse-genetics, which not only lacks the NSs gene but also the complete M genome segment. We have now compared the vaccine efficacies of these two next-generation vaccines and included the Clone 13 vaccine as a control for optimal efficacy. Groups of eight lambs were vaccinated once and challenged three weeks later. All mock-vaccinated lambs developed high fever and viremia and three lambs did not survive the infection. As expected, lambs vaccinated with Clone 13 were protected from viremia and clinical signs. Two lambs vaccinated with R566 developed mild fever after challenge infection, which was associated with low levels of viral RNA in the blood, whereas vaccination with the NSR-Gn vaccine completely prevented viremia and clinical signs.

Efficacy of a pandemic (H1N1) 2009 virus vaccine in pigs against the pandemic influenza virus is superior to commercially available swine influenza vaccines.

In April 2009 a new influenza A/H1N1 strain, currently named "pandemic (H1N1) influenza 2009" (H1N1v), started the first official pandemic in humans since 1968. Several incursions of this virus in pig herds have also been reported from all over the world. Vaccination of pigs may be an option to reduce exposure of human contacts with infected pigs, thereby preventing cross-species transfer, but also to protect pigs themselves, should this virus cause damage in the pig population. Three swine influenza vaccines, two of them commercially available and one experimental, were therefore tested and compared for their efficacy against an H1N1v challenge. One of the commercial vaccines is based on an American classical H1N1 influenza strain, the other is based on a European avian H1N1 influenza strain. The experimental vaccine is based on reassortant virus NYMC X179A (containing the hemagglutinin (HA) and neuraminidase (NA) genes of A/California/7/2009 (H1N1v) and the internal genes of A/Puerto Rico/8/34 (H1N1)). Excretion of infectious virus was reduced by 0.5-3 log(10) by the commercial vaccines, depending on vaccine and sample type. Both vaccines were able to reduce virus replication especially in the lower respiratory tract, with less pathological lesions in vaccinated and subsequently challenged pigs than in unvaccinated controls. In pigs vaccinated with the experimental vaccine, excretion levels of infectious virus in nasal and oropharyngeal swabs, were at or below 1 log(10)TCID(50) per swab and lasted for only 1 or 2 days. An inactivated vaccine containing the HA and NA of an H1N1v is able to protect pigs from an infection with H1N1v, whereas swine influenza vaccines that are currently available are of limited efficaciousness. Whether vaccination of pigs against H1N1v will become opportune remains to be seen and will depend on future evolution of this strain in the pig population. Close monitoring of the pig population, focussing on presence and evolution of influenza strains on a cross-border level would therefore be advisable.

Bovine viral diarrhoea virus antigen in foetal calf serum batches and consequences of such contamination for vaccine production.

A protocol to test foetal calf serum (FCS) for contamination with bovine viral diarrhoea virus (BVDV) is described. Following this protocol, which combines cell culture methods and detection of pestivirus RNA, seven batches of FCS were tested. Infectious BVDV was detected in four of those batches. One of the remaining batches contained a relatively high number of non-infectious BVDV particles. A sample of this batch was formulated with aluminium hydroxide and aluminium phosphate as adjuvant into an experimental vaccine preparation. This product was injected twice into BVDV seronegative cattle with a 4 week interval. Blood samples taken 4 weeks after the second application were negative for BVDV specific antibodies. Our data stress that detection of BVDV RNA is not sufficient for a complete risk assessment on FCS. Discrimination between infectious and non-infectious BVDV is essential. This can only be achieved by cell culture methods.

Leukopenia and thrombocytopenia in pigs after infection with bovine viral diarrhoea virus-2 (BVDV-2).

The aim of this study was to investigate whether bovine viral diarrhoea virus-2 (BVDV-2) is pathogenic for pigs, which organs become infected and whether or to which extent the virus is excreted into the environment. Ten pigs were observed for clinical reactions after infection with a BVDV-2 strain, that has been shown to be pathogenic in calves under experimental conditions. Samples were taken to monitor thrombocyte and leukocyte counts as well as antibody development. Post mortem examinations were performed at 7, 11 and 27 days after infection. Tissue samples were collected for virus isolation, histological and immunohistological examination. All ten pigs became infected and BVDV could be re-isolated from the lymphocytes, the plasma and different lymphatic organs. The infection passed clinically inapparent, apart from a slight increase in body temperature in some animals. Some animals developed a slight leukopenia and/or thrombocytopenia. There were no macroscopic or histological lesions observed that could specifically be related to the inoculation of BVDV-2. With respect to all parameters studied, the infection and the consequences thereof were clearly less pronounced in pigs as compared to cattle, the natural host. Our results indicate, that pigs infected with BVDV-2 might develop antibodies that cross-react in tests for antibodies against classical swine fever virus.

Investigations towards an efficacious and safe strangles vaccine: submucosal vaccination with a live attenuated Streptococcus equi.

As part of a search for a safe and efficacious strangles vaccine, several different vaccines and different vaccination routes were tested in foals. The degree of protection was evaluated after an intranasal challenge with virulent Streptococcus equi by clinical, postmortem and bacteriological examinations. Inactivated vaccines containing either native purified M-protein (500 microg per dose) or whole S equi cells (10(10) cells per dose) administered at least twice intramuscularly at intervals of four weeks, did not protect against challenge. Different live attenuated S equi mutants administered at least twice at intervals of four weeks by the intranasal route were either safe but not protective or caused strangles. In contrast, a live attenuated deletion mutant administered intramuscularly, induced complete protection but also induced unacceptable local reactions at the site of vaccination. Submucosal vaccination in the inner side of the upper lip with the live attenuated mutant at > or =10(8) colony-forming units per dose, appeared to be safe and efficacious in foals as young as four months of age. The submucosal vaccinations caused small transient swellings that resolved completely within two weeks, and postmortem no vaccine remnants or other abnormalities were found at the site of vaccination.

Chimeras in noncoding regions between serotypes I and II of segment A of infectious bursal disease virus are viable and show pathogenic phenotype in chickens.

Two serotypes, I and II, have been identified for infectious bursal disease virus (IBDV), a member of the family BIRNAVIRIDAE: Here, the generation by reverse genetics of IBDV chimeras in segment A of the bisegmented genome is reported. The 5- and 3'-noncoding regions (NCRs) of a serotype II strain were exchanged with the NCRs of a full-length cDNA clone of segment A of a serotype I strain. Isolated chimeric viruses were characterized in cell culture and susceptible chickens. The results show that IBDV chimeras in segment A were able to replicate in cell culture and that VP1 encoded by a serotype I segment B is functionally active with serotype I NCRs as well as with serotype II NCRs. Chimeric viruses infected susceptible chickens and caused mild depletion of bursal cells. Thus, the noncoding regions of segment A are not responsible for the different pathotypes of IBDV serotypes I and II.

Immunohistochemical and serological investigation of experimental Ornithobacterium rhinotracheale infection in chickens.

Immunohistochemical techniques were used to prove that Ornithobacterium rhinotracheale was the causative agent of lesions in the air sacs and lungs in chickens, but only after infection with Newcastle Disease virus (NDV). At first, the bacteria attached to the epithelium of the air sacs. Subsequently, they infiltrated the air sacs, and caused thickening of the air sacs, the formation of oedematous and granulomatous tissue, and accumulation of macrophages. The infection peaked at 5 to 9 days, after which recovery was seen. In the lungs, some areas with bronchially-associated lymphoid tissue were affected. The other organs investigated were shown not to be affected. In the absence of NDV infection, aerosol exposure of chickens to O. rhinotracheale only resulted in minimal and temporary microscopic air sac lesions. No O. rhinotracheale cells or fragments could be detected at any time point later than 2 days post-exposure. In spite of the absence of visible lesions, chickens exposed to O. rhinotracheale without prior NDV infection reacted serologically. The duration and the titre of this immune response was indistinguishable from that obtained in chickens exposed after NDV infection. Thus, infection with O. rhinotracheale appears to be restricted to the respiratory tract, with lesions only evident in birds previously infected with NDV, even though a strong serological response can be established in the absence of prior viral infection.

Vaccination of dogs against Babesia canis infection.

This paper describes the clinico-pathological parameters measured in dogs that were vaccinated against Babesia canis using soluble parasite antigens (SPA) and then challenged. The packed cell volume (PCV) and the plasma creatinine value decreased immediately after challenge. Actual PCV values could be predicted in the first 5-6 days of the infection, assuming that creatinine values were modulated by increase of plasma volume. This association no longer existed after that time, and observations indicated splenic involvement in reduction of numbers of circulating erythrocytes. The anaemia due to B. canis infection appears to be the result of a multifactorial process including plasma volume increase, erythrocyte retention in the spleen and erythrocyte destruction, partly due to parasite proliferation. Vaccination limited the reduction of PCV values, and the development of splenomegaly. Differences in protection between vaccinated and control animals became apparent 6 days after infection, when a memory immune response becomes operative, and the onset of recovery of vaccinated animals correlated with the onset of antibody production against SPA.

Experimental infection in turkeys and chickens with Ornithobacterium rhinotracheale.

Ornithobacterium rhinotracheale was found to cause growth retardation in both turkeys and chickens after experimental intra-air sac administration and to cause growth retardation together with airsacculitis and pneumonia after aerosol administration. Both turkey and chicken isolates of O. rhinotracheale were able to induce the same kind of respiratory inflammations and weight-gain losses in chickens as well as turkeys. Turkey rhinotracheitis virus was found to have a triggering effect on the O. rhinotracheale infection in turkeys, and Newcastle disease virus and to a lesser extent infectious bronchitis virus showed triggering effects on the O. rhinotracheale infection in chickens. Ornithobacterium rhinotracheale could be reisolated from affected organs of experimentally infected birds.

A mouse model for testing the pathogenicity of equine herpes virus-1 strains.

A mouse model was developed for testing the pathogenicity of equine herpes virus-1 (EHV-1) strains. The model was validated with EHV-1 strains that are known to be of a low or high pathogenicity in horses. From all parameters tested, the safety index, which was calculated from the body weights of the mice after infection, proved to be the best predictive parameter. When this parameter was used, good and reliable correlations were found with the pathogenicity of the EHV-1 strains in horses. This method enabled the differentiation between the two experimental EHV-1 strains whose genetic backgrounds were supposedly equal.

Field experiments to evaluate the feasibility of eradication of Aujeszky's disease virus by different vaccination schedules.

In the present report, the extent of the reduction in Aujeszky's disease virus (ADV) dissemination achieved when pigs were intensively vaccinated with gI-deleted vaccines under field circumstances, was examined. On widely dispersed breeding-fattening farms, a gI-negative status was most rapidly obtained and the rate of new waves of infections was lowest when the attenuated Bartha strain was administered to both the sows and the fatteners. It was more difficult not only to reach but also to keep a gI-negative status on farms on which the sows were vaccinated with an inactivated vaccine and the fatteners with the attenuated Bartha strain or when the fattening pigs were not vaccinated at all. In a densely populated area, 9 of the 17 farms had gI-positive fatteners at the start of the intensive vaccination programme in which the attenuated Bartha strain was given to both the sows and the fatteners. Antibodies were not detected in the sera of the fatteners of each farm at some time during the experiments, but the fatteners on 7 of the 18 farms still showed antibodies against gI after 20 months of vaccination. At the end of the experiment, the percentage of fatteners with antibodies on these farms was markedly reduced compared with the percentage at the start of the experiment. Therefore, elimination of field virus may be feasible if intensive vaccination is carried out over a sufficiently long period of time. However, the high rate of reinfections experienced either due to reintroduction of the virus or to recrudescence should be a warning against too much optimism, particularly in regions with a dense swine population.