Production of a falcon herpesvirus vaccine.

Ten common kestrels (Falco tinnunculus) were used for this falcon herpes vaccine experiment. Four kestrels were subcutaneously given 1 ml of an attenuated falcon herpesvirus that had originally been isolated from the liver of an American prairie falcon (Falco mexicanus). This virus was then passaged 100 times on chicken embryo fibroblast cells (CEF-cells). Another 4 kestrels were given subcutaneously an inactivated falcon herpesvirus vaccine derived from the same American field strain. This vaccine was concentrated, inactivated by heat and betapropiolactone and emulsified in complete Freund's adjuvans. Two further kestrels served as controls and were not vaccinated. Twenty-one days after vaccination, all 10 kestrels were challenged with passage 3 of the American falcon herpesvirus. The 2 control kestrels died 6 days after challenge and 3 of those given the inactivated herpes vaccine died 9 days after challenge, with typical lesions of herpesvirus inclusion body hepatitis. Before the vaccination experiment, all 10 kestrels were free of serum neutralising antibodies to the falcon herpesvirus. Twenty-one days after vaccination, all 4 kestrels vaccinated with the attenuated vaccine, and one vaccinated with the killed vaccine, had seroconverted, having shown no symptoms to the challenge with a low passage virulent American herpesvirus strain. Following the challenge their antibody titres to falcon herpesvirus increased. No herpesvirus was isolated from any of the cloacal swabs taken during this experiment, indicating that there is no danger for any other birds from the attenuated herpesvirus vaccine. This experiment clearly shows that an attenuated falcon herpesvirus vaccine can protect kestrels from fatal inclusion body hepatitis.     

Plasma chemistry reference values in hybrid falcons in relation to their species of origin

Twenty-one blood chemistry parameters were determined in 127 peregrine falcons (Falco peregrinus), 79 gyr-peregrine hybrids and 166 gyr-saker hybrids. These parameters, together with those previously established in 53 gyr falcons (Falco rusticolus) and 234 saker falcons (Falco cherrug), were compared. There were statistically significant differences in 15 of the parameters between the saker and peregrine falcons; the saker and gyr falcons; the saker and gyr-saker hybrids; the peregrine and gyr falcons; and the peregrine and gyr-peregrine hybrids, but not between the gyr falcons and the gyr-saker falcon hybrids.     

The effect of dose, route and virulence of bovine herpesvirus 1 vaccine on experimental respiratory disease in cattle.

Three experiments were conducted with calves in which, following intramuscular or intranasal vaccination with virulent or attenuated bovine herpesvirus 1, calves were protected against bovine herpesvirus 1 -- Pasteurella haemolytica challenge. Calves receiving low doses of vaccine had lower levels of antibody and greater evidence of virus replication upon challenge than those receiving higher doses. In contrast 11/13 unvaccinated controls had fibrino-purulent pneumonia following challenge. The immune response developed later in younger calves and those given low doses of vaccine. Neutralizing antibodies to bovine herpes-virus 1 were not found in nasal secretions, but were present in serum seven days after vaccination. Bovine herpesvirus 1 was isolated before challenge from nasal secretions of calves vaccinated intranasally or intramuscularly with virulent virus but not those vaccinated intramuscularly with vaccine virus. It was concluded that both routes of vaccination with either virulent or attenuated bovine herpesvirus 1 provided protection from challenge with homologous or heterologous bovine herpesvirus 1 and that live vaccines should contain at least 10(3) plaque forming units/dose for effective immunization.     

ISCOM of BHV-1 envelope glycoproteins protected calves against both disease and infection.

A subunit vaccine in the form of immunostimulating complex (iscom) was prepared to contain the envelope glycoproteins of bovine herpesvirus type 1 (BHV-1). This iscom preparation was tested in a vaccination experiment on 4-month-old calves seronegative to BHV-1. In this experiment, four groups with three animals per group were used. Two groups were vaccinated with the iscom preparation twice, four weeks apart, one group with 50 micrograms and the other with 100 micrograms per calf. The third group received a commercial inactivated whole-virus vaccine applying the same vaccination program. The fourth group served as control. Two weeks after the second vaccination, all the animals were challenge-infected intranasally with a virulent BHV-1 strain and four days later with a virulent Pasteurella multocida--this in order to mimic hard field conditions. When exposed to challenge infection, all the animals vaccinated with the iscom were fully protected, i.e., no virus could be recovered from their nasal secretions and no clinical symptoms were recorded. In contrast, the animals vaccinated with the commercial vaccine, responded to challenge with moderate fever and loss of appetite, and virus was isolated from the nasal secretions. The animals in the control group developed severe clinical symptoms. In the sera of iscom-vaccinated animals, the virus neutralization titers reached levels of 1/3500 or higher.     

Enhancement of the immune response and virological protection of calves against bovine herpesvirus type 1 with an inactivated gE-deleted vaccine

Four immunisation protocols based on inactivated and attenuated commercially available marker vaccines for bovine herpesvirus type 1 (BHV-1) were compared. The first group of calves were vaccinated with an attenuated vaccine administered intranasally and an inactivated vaccine injected subcutaneously, four weeks apart; the second group were vaccinated twice with the attenuated vaccine, first intranasally and then intramuscularly; the third group were vaccinated twice subcutaneously with the inactivated vaccine; and the fourth group were vaccinated twice intramuscularly with the attenuated vaccine. A control group of calves were not vaccinated. The cellular and humoral immune responses were highest in the two groups which received at least one injection of the inactivated vaccine. Virological protection was observed in all the vaccinated groups after a challenge infection and reactivation by treatment with dexamethasone, but the calves which received one dose of the inactivated vaccine as a booster or two doses of the inactivated vaccine excreted significantly less of the challenge virus than the calves which were vaccinated only with the attenuated preparation.     

Falcon adenovirus infection in breeding Taita falcons (Falco fasciinucha).

Four female and 3 male Taita falcons (Falco fasciinucha) out of a breeding colony of 14 Taita falcons (7 pairs) died during the breeding season after showing lethargy and anorexia for 1 to 2 days. All animals were submitted for necropsy. Gross lesions in the female falcons were characterized by anemia secondary to marked hemorrhage into the ovary and oviduct, serofibrinous effusion into the cardioabdominal cavity and serosal petechiae. In addition, marked necrotizing splenitis and pulmonary hemorrhage were present. Histologically, the female falcons had mild necrotizing hepatitis with numerous intranuclear inclusion bodies and necrotizing splenitis with rare inclusion bodies. There were no gross lesions in the male falcons, and the histological lesions were characterized by urate deposition and rare intranuclear inclusion bodies in the renal tubular epithelial cells. Adenoviral particles were found by electron microscopy in the cloacal contents of the female Taita falcons but not in the male falcons. DNA in situ hybridization revealed widespread aviadenoviral nucleic acid within the nuclei of hepatocytes, renal tubular epithelial cells, and adrenal cells in the female falcons but no aviadenoviral nucleic acid in 1 male falcon and only a low quantity of adenoviral nucleic acid in the liver and kidney of another male Taita falcon. PCR amplified aviadenoviral DNA in the liver and intestine of all Taita falcons. The amplicons were sequenced, and the virus was identified as falcon adenovirus. The deaths of the female and male birds were attributed to the aviadenovirus infection.     

Plasma concentrations of voriconazole in falcons

Doses of 12.5 mg voriconazole/kg bodyweight administered every 12 hours by crop gavage to six falcons for 14 days provided peak plasma concentrations of more than 1 microg/ml, but the trough concentrations were lower and sometimes undetectable. Administering the same doses incorporated into meat that was fed to one falcon for seven days and to three falcons for up to 91 days provided similar plasma concentrations.     

Induction of immunity against pneumonic pasteurellosis following experimental infection in calves.

Immunity against pneumonic pasteurellosis was studied in calves after recovery from experimental respiratory disease with Pasteurella haemolytica. Nine calves were exposed to aerosols of parainfluenza-3 virus and Pasteurella haemolytica A1 six days apart to produce respiratory disease. After recovery from the disease, these nine principal and four control calves were challenged with aerosols of bovine herpesvirus 1 and P. haemolytica A1 four days apart. With this viral-bacterial challenge, the nine principal animals failed to develop clinical responses to this bacterial challenge and their lungs did not show the growth of P. haemolytica on cultures, whereas two of four control calves had elevated temperatures and developed necropurulent pneumonia with the isolation of P. haemolytica from the lungs. The principal calves had developed high levels of cytotoxin neutralizing antibodies in their sera following parainfluenza-3 virus-P. haemolytica infection. This demonstrated that immunity against pneumonic pasteurellosis can be achieved, with a suggestion that further search for an effective vaccine for P. haemolytica is warranted.     

A live attenuated glycoprotein E negative bovine herpesvirus 1 vaccine induces a partial cross-protection against caprine herpesvirus 1 infection in goats

Taking into account the close antigenic relationship between bovine herpesvirus 1 (BoHV-1) and caprine herpesvirus 1 (CpHV-1), a live attenuated glycoprotein E (gE) negative BoHV-1 vaccine was assessed in goats with the aim to protect against CpHV-1 infection. Vaccine safety was evaluated by intranasal inoculation of two groups of goats with either a gE-negative BoHV-1 vaccine or a virulent BoHV-1. The length of viral excretion and the peak viral titre were reduced with the gE-negative vaccine. To assess the efficacy, two goats were inoculated intranasally twice 2 weeks apart with a gE-negative BoHV-1 vaccine. Four weeks later, immunised and control goats were challenged with CpHV-1. A 2 log(10) reduction in the peak viral titre was observed and the challenge virus excretion lasted 2 days more in immunised than in control goats. These data indicate the safety and the partial efficacy of a live attenuated gE-negative BoHV-1 vaccine intranasally administrated in goats.     

Molecular assays for detection of falcon adenovirus.

Falcon adenovirus is a newly recognized member of the family Aviadenoviridae and includes 2 closely related strains that are pathogenic to several species of falcons. Peregrine falcons appear to be one of the primary reservoirs, but recent outbreaks suggest that other carrier species probably exist. To allow screening of captive birds for virus shedding and investigations of disease outbreaks, conventional and real-time, quantitative polymerase chain reaction (PCR) assays and an in situ hybridization technique were developed. The diagnostic protocols were used on tissue and fecal samples from 7 species or subspecies of falcons infected with adenovirus as well as adenoviruses from other birds and mammals. The assays were specific for falcon adenovirus and detected both strains of virus in fecal samples from living animals or frozen and formalin-fixed, paraffinized tissues. Together with established serologic tests for falcon adenovirus, these molecular assays are valuable tools for management and conservation of falcons in captivity and the wild.     

Experimental Bovine Pneumonic Pasteurellosis II. Genesis and Prevention

Two experiments were conducted. In the first, 16 crossbred Hereford calves were divided into two equal groups. The first group was vaccinated intranasally with a commercial vaccine against bovid herpesvirus 1 and the second group was unvaccinated. The calves were later exposed to an aerosol of bovid herpesvirus 1 (strain 108) for five minutes. Four calves from each group were subjected to transportation and four calves from each group were kept in an environmental chamber for four days. Four days after viral aerosol all calves were exposed to an aerosol of Pasteurella haemolytica and the same subgroups were again transported or held in the chamber for a further four days.

The calves that did not die from pneumonia were necropsied ten days after the final day of transport. Pulmonary lesions were present in both vaccinated and control animals but were less extensive in the vaccinated calves. Six of eight vaccinated but none of the eight control calves survived.

In the second experiment, eight crossbred Hereford calves were divided into two equal groups. One group was vaccinated with bovid herpesvirus 1 (strain 108) and the other acted as controls. Four weeks later all calves were sequentially exposed to aerosols of bovid herpesvirus 1 (strain 108) and P. haemolytica four days apart. Three of the four controls but none of the vaccinates died from pneumonia. Every lobe of the lungs in all the controls was affected by pneumonia while no pulmonary lesions were found in the vaccinated calves. The differences in efficacy of the modes of vaccination and the possible role of transport stress are discussed.

     

Experimental bovine pneumonic pasteurellosis. I. Prevention of the disease.

Fifteen three- to six-month old Hereford-cross calves were divided into three groups. The first group was inoculated with bovid herpersvirus 1 (Strain 108), the second with a commercial intranasal vaccine against bovid herpesvirus 1 and the third group acted as controls. At least three weeks after vaccination, all calves were weaned, placed in an environmental chamber at 25.0 degrees C (days) and -13.3 degrees C (nights) and challenged with an aerosol of bovid herpesvirus 1 followed four days later by an aerosol of Pasteurella haemolytica. All surviving calves were sacrificed four days after the second aerosol. None of the calves inoculated with bovid herpesvirus 1 virus or the commercial vaccine developed a generalized pneumonia, although there were one or two nodules (4--8 mm diameter) in two of the calves given the commercial vaccine. Four of the five control calves had extensive lobar pneumonia at necropsy, two of the five died from the disease. Details of the clinical, pathological, bacteriological, virological and some of the serological findings are reported.     

Outbreak of microsporidiosis caused by Enterocytozoon bieneusi in falcons

Four falcons from a private collection of 137 falcons in Abu Dhabi (UAE) died suddenly in summer 2005. In order to screen for a possible disease among the remaining falcons in the aviary, all other birds were caught, examined and treated if necessary. Most of the falcons suffered from massive lice infestation and 74 falcons additionally from a heavy Caryospora sp. burden. Endoscopy revealed yellowish plaques on intestines, livers or kidneys in 70 birds (51.1% morbidity). Proliferative serositis was seen in 17 out of 24 necropsied birds with plaques on intestines, livers or kidneys, which did not resemble any known disease in falcons. However, apart from 20 falcons, which died within a 6-week period after the initial examinations due to advanced disease stages, all other falcons responded well to the treatment with dimetridazole (Emtryl), indicating protozoal disease. Immunohistochemistry confirmed the presence of microsporidial antigen. The final diagnosis of Enterocytozoon (E.) bieneusi genotype D was confirmed with materials from 6 birds by PCR and sequencing. To our knowledge this is the first report of microsporidiosis caused by E. bieneusi in raptors in general and in falcons in particular. However, it is still unclear for how long E. bieneusi was present in the falcon flock, and which role it played in the development of the disease. Predisposing factors such as high temperature and overcrowding in the aviary induced immune suppression causing massive lice infestation as well as coccidiosis, thus paving the way for invasion with microsporidial spores.     

Comparison of the efficacy of three commercial bacterins in preventing canine leptospirosis

Twenty-four specific pathogen-free beagles were randomly allocated into four groups (three vaccinated groups and one control group) and inoculated at nine and 12 weeks of age with one of three commercial inactivated Leptospira vaccines: A (Vanguard 7; Pfizer Santé Animale), B (Dohyvac 7L; Fort Dodge), and C (Nobivac DHPPi + Lepto; Intervet International); the control group received Nobivac DHPPi (Intervet International). Seven weeks after the second vaccination all the dogs were challenged with Leptospira interrogans serogroup canicola. All the vaccinated dogs developed a mild serological response (microscopic agglutination titres) after the booster vaccination. A significant serological response after the challenge was observed, particularly in the controls. The challenge induced fever and clinical disorders in the control group, whereas in the vaccinated groups the clinical signs were mild. Blood cultures became positive in all control dogs, and in one of six dogs vaccinated with vaccine A and two of four dogs vaccinated with vaccine B; none of the six dogs vaccinated with vaccine C was leptospiraemic at any stage of the experiment. Urine cultures were positive in all the control dogs two weeks after the challenge. One of six dogs vaccinated with vaccine A and two of four dogs vaccinated with vaccine B shed bacteria in their urine after the challenge, but none of the dogs vaccinated with vaccine C shed bacteria in their urine at any time during the experiment.     

Drop of egg production in chickens by experimental infection with an avian metapneumovirus strain PLE8T1 derived from swollen head syndrome and the application to evaluate vaccine

Decreases in egg production and increased incidence of abnormal eggs due to malformation of egg shells were observed in specific pathogen free (SPF) 173-day-old laying hens inoculated intravenously with an avian metapneumovirus (aMPV) strain PLE8T1. This strain was derived from an isolate from broiler birds exhibiting swollen head syndrome (SHS). Some SPF birds inoculated with the virus showed, slight diarrhea without any respiratory symptoms. Thus, the PLE8T1 strain was used as a challenge virus to evaluate efficacy of aMPV vaccines. SPF chickens which received a live attenuated aMPV vaccine (NEMOVAC; Merial) at 7 or 77 days old and an inactivated aMPV vaccine (OVO-4; Merial) at 105 days old were protected against poor egg production caused by the challenge with the PLE8T1 strain. Thus, aMPV, the PLE8T1 strain passaged 22 times after isolation, from birds exhibiting SHS, could induce a drop in egg production in laying hens accompanied by malformation of egg shells. It was suggested that this challenge system could be applied to evaluate the efficacy of aMPV vaccine.     

Update on vaccination of cattle and wildlife populations against tuberculosis

A recombinant bovine herpesvirus 5 lacking thymidine kinase and glycoprotein E genes (BoHV-5gEΔTKΔ) was evaluated as a live experimental vaccine. In a first experiment, ten-months-old calves were vaccinated intramuscularly (n=9) or remained as controls (n=8) and 42 days later were challenged with BoHV-5 or BoHV-1 intranasally. The four control calves challenged with BoHV-5 developed severe depression and neurological signs and were euthanized in extremis at days 13 and 14 pos-infection (pi); the five vaccinated animals challenged with BoHV-5 remained healthy. The titers of virus shedding were reduced (p<0.01) from days 3 to 7 post-infection (pi) in vaccinated animals. Control and vaccinated calves challenged with BoHV-1 presented mild transient respiratory signs; yet the magnitude of virus shedding was reduced (p<0.05) in vaccinated animals (days 5, 9 and 11pi). In a second experiment, young calves (100-120 days-old) were vaccinated (n=15) or kept as controls (n=5) and subsequently challenged with a BoHV-1 isolate. Control calves developed moderate to severe rhinitis and respiratory distress; two were euthanized in extremis at days 5 and 9 pi, respectively. In contrast, vaccinated animals were protected from challenge and only a few developed mild and transient nasal signs. The duration and titers of virus shedding after challenge were reduced (p<0.05) in vaccinated animals comparing to controls. In both experiments, vaccinated animals developed antibodies to gE only after challenge. These results demonstrate homologous and heterologous protection and are promising towards the use of the recombinant BoHV-5gEΔTKΔ in vaccine formulations to control BoHV-5 and BoHV-1 infections.     

Efficacy of an intranasal modified live bovine respiratory syncytial virus and temperature-sensitive parainfluenza type 3 virus vaccine in 3-week-old calves experimentally challenged with PI3V

Two experimental parainfluenza type 3 virus (PI3V) challenge studies were undertaken to evaluate the efficacy of a single intranasal dose of an attenuated live vaccine containing modified live bovine respiratory syncytial virus (BRSV) and temperature-sensitive PI3V in 3-week-old calves. In the first study, vaccine efficacy was evaluated in colostrum deprived calves. Nasal shedding of PI3V was highly significantly reduced in vaccinated calves challenged 10 days or 21 days after vaccination. In the second study, vaccine efficacy was assessed in calves with maternal antibodies against PI3V by challenge 66 days post-vaccination. Vaccination also significantly reduced PI3V excretion after challenge in this study. In both studies, clinical signs after challenge were very mild and were not different between vaccinated and control calves.     

Limited efficacy of West Nile virus vaccines in large falcons (Falco spp.)

West Nile virus (WNV) can lead to fatal diseases in raptor species. Unfortunately, there is no vaccine which has been designed specifically for use in breeding stocks of falcons. Therefore the immunogenicity and protective capacity of two commercially available WNV vaccines, both approved for use in horses, were evaluated in large falcons. One vaccine contained adjuvanted inactivated WNV lineage 1 immunogens, while the second represented a canarypox recombinant live virus vector vaccine. The efficacy of different vaccination regimes for these two vaccines was assessed serologically and by challenging the falcons with a WNV strain of homologous lineage 1. Our studies show that the recombinant vaccine conveys a slightly better protection than the inactivated vaccine, but moderate (recombinant vaccine) or weak (inactivated vaccine) side effects were observed at the injection sites. Using the recommended 2-dose regimen, both vaccines elicited only sub-optimal antibody responses and gave only partial protection following WNV challenge. Better results were obtained for both vaccines after a third dose, i.e. alleviation of clinical signs, absence of fatalities and reduction of virus shedding and viraemia. Therefore the consequences of WNV infections in falcons can be clearly alleviated by vaccination, especially if the amended triple administration scheme is used, although side effects at the vaccination site must be accepted.     

Vaccination of calves with contaminated batches of bovine herpes virus 1 vaccines did not result in infection with bovine virus diarrhea virus

The aim of the experiment was to study whether bovine herpesvirus 1 (BHV1) marker vaccine batches known to be contaminated with bovine virus diarrhoea virus (BVDV) type 1 could cause BVD in cattle. For this purpose, four groups of cattle were used. The first group (n = 4 calves, the positive control group), was vaccinated with vaccine from a batch contaminated with BVDV type 2. The second group (n = 4 calves, the negative control group), was vaccinated with vaccine from a batch that was not contaminated with BVDV. The third group (n = 39 calves), was vaccinated with a vaccine from one of four batches contaminated with BVDV type 1 (seronegative experimental group). The fourth group (n = 6 seropositive heifers), was vaccinated with a vaccine from one of three batches known to be contaminated with BVDV type 1. All cattle were vaccinated with an overdose of the BHV1 marker vaccine. At the start of the experiment, all calves except those from group 4 were seronegative for BVDV and BHV1. The calves from group 4 had antibodies against BVDV, were BVDV-free and seronegative to BHV1. After vaccination, the positive control calves became severely ill, had fever for several days, and BVDV was isolated from nasal swabs and white blood cells. In addition, these calves produced antibodies to BVDV and BHV1. No difference in clinical scores of the other groups was seen, nor were BVDV or BVDV-specific antibody responses detected in these calves; however, they did produce antibodies against BHV1. The remainder of each vaccine vial used was examined for the presence of infectious BVDV in cell culture. From none of the vials was BVDV isolated after three subsequent passages. This indicates that BVDV was either absent from the vials or was present in too low an amount to be isolated. Thus vaccination of calves with vaccines from BHV1 marker vaccine batches contaminated with BVDV type 1 did not result in BVDV infections.     

Evaluation of the dosage of ivermectin in falcons

Twelve groups of falcons, each containing three female gyrfalcon-peregrine falcon hybrids (Falco rusticolus x Falco peregrinus) were injected intramuscularly with a single dose of ivermectin ranging from 0.2 mg/kg to 11 mg/kg bodyweight, and a control group was injected with water. Doses of ivermectin between 0.2 and 5 mg/kg failed to produce clinical signs of illness in the birds. Four birds which received either 6, 7 or 8 mg/kg showed slight clinical signs, and all the birds receiving 9 to 11 mg/kg showed more or less severe clinical signs of anorexia, apathy and sedation. Slight changes in the mean plasma activities of aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase (AP) were detected in the group dosed with 5 mg/kg, and higher dosages caused marked changes in these enzymes as well as in the mean plasma activity of lactate dehydrogenase. The mean activity of AP decreased, and the activities of the other enzymes increased. A dosage of 2 to 3 mg/kg ivermectin is recommended as a safe and effective antiparasitic drug for falcons and it has been used successfully to treat infestations of Serratospiculum species.