Experimental infection of South American camelids with bluetongue virus serotype 8.

Bluetongue (BT) is an infectious, non-contagious disease of wild and domestic ruminants. It is caused by bluetongue virus (BTV) and transmitted by Culicoides biting midges. Since 1998, BT has been emerging throughout Europe, threatening not only the naïve ruminant population. Historically, South American camelids (SAC) were considered to be resistant to BT disease. However, recent fatalities related to BTV in captive SAC have raised questions about their role in BTV epidemiology. Data on the susceptibility of SAC to experimental infection with BTV serotype 8 (BTV-8) were collected in an animal experiment. Three alpacas (Vicugna pacos) and three llamas (Lama glama) were experimentally infected with BTV-8. They displayed very mild clinical signs. Seroconversion was first measured 6-8 days after infection (dpi) by ELISA, and neutralising antibodies appeared 10-13 dpi. BTV-8 RNA levels in blood were very low, and quickly cleared after seroconversion. However, spleens collected post-mortem were still positive for BTV RNA, over 71 days after the last detection in blood samples. Virus isolation was only possible from blood samples of two alpacas by inoculation of highly sensitive interferon alpha/beta receptor-deficient (IFNAR(-/-)) mice. An in vitro experiment demonstrated that significantly lower amounts of BTV-8 adsorb to SAC blood cells than to bovine blood cells. Although this experiment showed that SAC are generally susceptible to a BTV-8 infection, it indicates that these species play a negligible role in BTV epidemiology.

Coexpression of avian influenza virus H5 and N1 by recombinant Newcastle disease virus and the impact on immune response in chickens.

To analyze the contribution of neuraminidase (NA) toward protection against avian influenza virus (AIV) infection, three different recombinant Newcastle disease viruses (NDVs) expressing hemagglutinin (HA) or NA, or both, of highly pathogenic avian influenza virus (HPAIV) were generated. The lentogenic NDV Clone 30 was used as backbone for the insertion of HA of HPAIV strain A/chicken/Vietnam/P41/05 (H5N1) and NA of HPAIV strain A/duck/Vietnam/TG24-01/05 (H5N1). The HA was inserted between the genes encoding NDV phosphoprotein (P) and matrixprotein (M), and the NA was inserted between the fusion (F) and hemagglutinin-neuraminidase protein (HN) genes, resulting in NDVH5VmPMN1FHN. Two additional recombinants were constructed carrying the HA gene between the NDV P and M genes (NDVH5VmPM) or the NA between F and HN (NDVN1FHN). All recombinants replicated well and stably expressed the HA gene, the NA gene, or both. Chickens immunized with NDVH5VmPMN1FHN or NDVH5VmPM were protected against two different HPAIV H5N1 and also against HPAIV H5N2. In contrast, immunization of chickens with NDVN1FHN induced NDV- and AIV N1-specific antibodies but did not protect the animals against a lethal dose of HPAIV H5N1. Furthermore, expression of AIV N1, in addition to AIV H5 by NDV, did not increase protection against HPAIV H5N1.

Longitudinal 2 years field study of conventional vaccination against highly pathogenic avian influenza H5N1 in layer hens.

A licensed, inactivated vaccine based on a low pathogenic avian influenza virus strain (H5N2) was evaluated in layer hens kept under field conditions during a 2-year period. Vaccine efficacy was investigated by specific antibodies and by challenge-contact experiments using highly pathogenic avian influenza viruses (HPAIV) H5N1. Basic immunization with two applications induced clinical protection. Virus excretion by vaccinated hens was significantly reduced compared to non-vaccinated controls; transmission to non-vaccinated and vaccinated contact birds was not fully interrupted. Vaccination efficacy is influenced by several factors including antigenic relatedness between vaccine and field strains, but also by species, age and type of commercial uses of the host. Limitations and risks of HPAIV vaccination as silent spread of HPAIV and emergence of escape mutants must be considered a priori and appropriate corrective measures have to be installed.

Infectious blood or culture-grown virus: a comparison of bluetongue virus challenge models.

The World Organisation for Animal Health (OIE) currently recommends using infectious ruminant blood as challenge inoculum in bluetongue virus (BTV) vaccination and challenge experiments. The use of virus grown in cultured cells is discouraged because culture passages can lead to changes in virus phenotype, including reduced replication efficiency and virulence in the host, while the OIE considers clinical disease in control animals indispensable evidence of successful infection. In the present study, two groups of five sheep were inoculated with either infectious calf blood lysate or culture-grown bluetongue virus of serotype 8 (BTV-8) (2 × 10(4)TCID(50) and 5 × 10(5)TCID(50), respectively). No pronounced difference in the induction and progression of viraemia as determined by real-time RT-PCR, which is the most objective parameter in the evaluation of vaccine efficacy, was observed. In a second experiment, the virulence of both inocula was confirmed by fatal infection of interferon receptor-deficient mice. The recent availability of highly sensitive molecular methods for the detection of BTV can finally shift the focus away from clinical disease. For the sake of objective and repeatable BTV challenge experiments, the OIE should reconsider its policy on culture-grown virus.

Protection and virus shedding of falcons vaccinated against highly pathogenic avian influenza A virus (H5N1).

Because fatal infections with highly pathogenic avian influenza A (HPAI) virus subtype H5N1 have been reported in birds of prey, we sought to determine detailed information about the birds' susceptibility and protection after vaccination. Ten falcons vaccinated with an inactivated influenza virus (H5N2) vaccine seroconverted. We then challenged 5 vaccinated and 5 nonvaccinated falcons with HPAI (H5N1). All vaccinated birds survived; all unvaccinated birds died within 5 days. For the nonvaccinated birds, histopathologic examination showed tissue degeneration and necrosis, immunohistochemical techniques showed influenza virus antigen in affected tissues, and these birds shed high levels of infectious virus from the oropharynx and cloaca. Vaccinated birds showed no influenza virus antigen in tissues and shed virus at lower titers from the oropharynx only. Vaccination could protect these valuable birds and, through reduced virus shedding, reduce risk for transmission to other avian species and humans.