<p>Groups of 10 BALB/c mice were vaccinated with 10<sup>6.5</sup> TCID<sub&g... more <p>Groups of 10 BALB/c mice were vaccinated with 10<sup>6.5</sup> TCID<sub>50</sub> of NDV-H5 or NDV-sH5<sup>3</sup>, either via the ON or IM route. One group of mice was mock-vaccinated (PBS) as challenge control. Three weeks after the vaccination, mice were infected with ā¼10 LD<sub>50</sub> of HPAIV H5N1 and weighed daily and observed for clinical signs during 14 days. Graphed for each group are A and B) Kaplan-Meier survival curves indicating percentage of survival p.c., C and D) mean percentage of body weight changes relative to starting weights measured on the day of challenge (day 0) with error bars representing the standard deviation, and E and F) median clinical scores observed after challenge. For reference, each panel includes the same PBS group.</p
<p>Groups of 10 SPF chickens were immunized with 10<sup>7</sup> TCID<sub>... more <p>Groups of 10 SPF chickens were immunized with 10<sup>7</sup> TCID<sub>50</sub> of NDV-H5 or NDV-sH5<sup>3</sup>, either via the ON/IT or IM route. Another group received an IM immunization with sH5<sup>3</sup> adjuvanted in Stimune. As a challenge control, one group of chickens was mock-vaccinated with PBS. Three weeks after the vaccination, birds were challenged with ā¼10<sup>5</sup> TCID<sub>50</sub> of HPAIV H5N1. A and B) Kaplan-Meier survival curves indicating percentage of survival p.c. on each day for each group that was (mock-)vaccinated via the ON/IT (A) or IM (B) route. C and D) Clinical index calculated on basis of clinical signs (as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0044447#s2" target="_blank">materials and methods</a>) observed after challenge for each group that was (mock-)vaccinated ON/IT (C) or IM (D). An index of 3.0 means that all birds died within 24 hours. For reference, each panel includes the same PBS group.</p
a<p>Day (D) post infection on which tracheal and cloacal swabs were collected are indicated... more a<p>Day (D) post infection on which tracheal and cloacal swabs were collected are indicated. Amount of RNA detected in swabs is expressed as log10 TCID50 ml-1 equivalents.āā=ā negative (C<sub>T</sub> >39.0); x ā=ā not tested because chicken did not survive after challenge with HPAIV H5N1.</p
In April 2020, respiratory disease and increased mortality were observed in farmed mink on two fa... more In April 2020, respiratory disease and increased mortality were observed in farmed mink on two farms in the Netherlands. In both farms, at least one worker had been found positive for SARS-CoV-2. Necropsies of the mink revealed interstitial pneumonia, and organ and swab samples tested positive for SARS-CoV-2 RNA by qPCR. Variations in viral genomes point at between-mink transmission on the farms and lack of infection link between the farms. Inhalable dust in the mink houses contained viral RNA, indicating possible exposure of workers.
Respiratory disease and increased mortality occurred in minks on two farms in the Netherlands, wi... more Respiratory disease and increased mortality occurred in minks on two farms in the Netherlands, with interstitial pneumonia and SARS-CoV-2 RNA in organ and swab samples. On both farms, at least one worker had coronavirus disease-associated symptoms before the outbreak. Variations in mink-derived viral genomes showed between-mink transmission and no infection link between the farms. Inhalable dust contained viral RNA, indicating possible exposure of workers. One worker is assumed to have attracted the virus from mink.
Little is known about the intermolecular interactions between the viral proteins of infectious bu... more Little is known about the intermolecular interactions between the viral proteins of infectious bursal disease virus (IBDV). By using the yeast two-hybrid system, which allows the detection of proteināprotein interactions in vivo, all possible interactions were tested by fusing the viral proteins to the LexA DNA-binding domain and the B42 transactivation domain. A heterologous interaction between VP1 and VP3, and homologous interactions of pVP2, VP3, VP5 and possibly VP1, were found by co-expression of the fusion proteins in Saccharomyces cerevisiae. The presence of the VP1āVP3 complex in IBDV-infected cells was confirmed by co-immunoprecipitation studies. Kinetic analyses showed that the complex of VP1 and VP3 is formed in the cytoplasm and eventually is released into the cell-culture medium, indicating that VP1āVP3 complexes are present in mature virions. In IBDV-infected cells, VP1 was present in two forms of 90 and 95 kDa. Whereas VP3 initially interacted with both the 90 and 95 ...
Background: Highly pathogenic avian influenza virus (HPAIV) causes a highly contagious often fata... more Background: Highly pathogenic avian influenza virus (HPAIV) causes a highly contagious often fatal disease in poultry, resulting in significant economic losses in the poultry industry. HPAIV H5N1 also poses a major public health threat as it can be transmitted directly from infected poultry to humans. One effective way to combat avian influenza with pandemic potential is through the vaccination of poultry. Several live vaccines based on attenuated Newcastle disease virus (NDV) that express influenza hemagglutinin (HA) have been developed to protect chickens or mammalian species against HPAIV. However, the zoonotic potential of NDV raises safety concerns regarding the use of live NDV recombinants, as the incorporation of a heterologous attachment protein may result in the generation of NDV with altered tropism and/or pathogenicity. Methodology/Principal Findings: In the present study we generated recombinant NDVs expressing either full length, membrane-anchored HA of the H5 subtype (...
Little is known about the intermolecular interactions between the viral proteins of infectious bu... more Little is known about the intermolecular interactions between the viral proteins of infectious bursal disease virus (IBDV). By using the yeast two-hybrid system, which allows the detection of proteināprotein interactions in vivo, all possible interactions were tested by fusing the viral proteins to the LexA DNA-binding domain and the B42 transactivation domain. A heterologous interaction between VP1 and VP3, and homologous interactions of pVP2, VP3, VP5 and possibly VP1, were found by co-expression of the fusion proteins in Saccharomyces cerevisiae. The presence of the VP1āVP3 complex in IBDV-infected cells was confirmed by co-immunoprecipitation studies. Kinetic analyses showed that the complex of VP1 and VP3 is formed in the cytoplasm and eventually is released into the cell-culture medium, indicating that VP1āVP3 complexes are present in mature virions. In IBDV-infected cells, VP1 was present in two forms of 90 and 95 kDa. Whereas VP3 initially interacted with both the 90 and 95 ...
Inoculation of infl uenza (H5N1) into beagles resulted in virus excretion and rapid seroconversio... more Inoculation of infl uenza (H5N1) into beagles resulted in virus excretion and rapid seroconversion with no disease. Binding studies that used labeled infl uenza (H5N1) showed virus attachment to higher and lower respiratory tract tissues. Thus, dogs that are subclinically infected with infl u-enza (H5N1) may contribute to virus spread. Avian influenza (H5N1) virus has been shown to be infectious not only for birds but also for humans and mammals such as mice, ferrets, and cats. Carnivorous mammals that are susceptible to subtype H5N1 may contribute to spread of the virus; shedding of influenza (H5N1) by pet carnivores may pose a risk to humans. Cats experimentally inoculated with influenza (H5N1) have been shown to be susceptible to infection and to shed virus (1). However, dogs ā susceptibility to this virus is unknown. Unpublished studies indicate that a substantial number of dogs tested in Thailand were positive for antibodies against H5N1 subtype (2). Recently, isolation of infl...
<p>Groups of 10 BALB/c mice were vaccinated with 10<sup>6.5</sup> TCID<sub&g... more <p>Groups of 10 BALB/c mice were vaccinated with 10<sup>6.5</sup> TCID<sub>50</sub> of NDV-H5 or NDV-sH5<sup>3</sup>, either via the ON or IM route. One group of mice was mock-vaccinated (PBS) as challenge control. Three weeks after the vaccination, mice were infected with ā¼10 LD<sub>50</sub> of HPAIV H5N1 and weighed daily and observed for clinical signs during 14 days. Graphed for each group are A and B) Kaplan-Meier survival curves indicating percentage of survival p.c., C and D) mean percentage of body weight changes relative to starting weights measured on the day of challenge (day 0) with error bars representing the standard deviation, and E and F) median clinical scores observed after challenge. For reference, each panel includes the same PBS group.</p
<p>Groups of 10 SPF chickens were immunized with 10<sup>7</sup> TCID<sub>... more <p>Groups of 10 SPF chickens were immunized with 10<sup>7</sup> TCID<sub>50</sub> of NDV-H5 or NDV-sH5<sup>3</sup>, either via the ON/IT or IM route. Another group received an IM immunization with sH5<sup>3</sup> adjuvanted in Stimune. As a challenge control, one group of chickens was mock-vaccinated with PBS. Three weeks after the vaccination, birds were challenged with ā¼10<sup>5</sup> TCID<sub>50</sub> of HPAIV H5N1. A and B) Kaplan-Meier survival curves indicating percentage of survival p.c. on each day for each group that was (mock-)vaccinated via the ON/IT (A) or IM (B) route. C and D) Clinical index calculated on basis of clinical signs (as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0044447#s2" target="_blank">materials and methods</a>) observed after challenge for each group that was (mock-)vaccinated ON/IT (C) or IM (D). An index of 3.0 means that all birds died within 24 hours. For reference, each panel includes the same PBS group.</p
a<p>Day (D) post infection on which tracheal and cloacal swabs were collected are indicated... more a<p>Day (D) post infection on which tracheal and cloacal swabs were collected are indicated. Amount of RNA detected in swabs is expressed as log10 TCID50 ml-1 equivalents.āā=ā negative (C<sub>T</sub> >39.0); x ā=ā not tested because chicken did not survive after challenge with HPAIV H5N1.</p
In April 2020, respiratory disease and increased mortality were observed in farmed mink on two fa... more In April 2020, respiratory disease and increased mortality were observed in farmed mink on two farms in the Netherlands. In both farms, at least one worker had been found positive for SARS-CoV-2. Necropsies of the mink revealed interstitial pneumonia, and organ and swab samples tested positive for SARS-CoV-2 RNA by qPCR. Variations in viral genomes point at between-mink transmission on the farms and lack of infection link between the farms. Inhalable dust in the mink houses contained viral RNA, indicating possible exposure of workers.
Respiratory disease and increased mortality occurred in minks on two farms in the Netherlands, wi... more Respiratory disease and increased mortality occurred in minks on two farms in the Netherlands, with interstitial pneumonia and SARS-CoV-2 RNA in organ and swab samples. On both farms, at least one worker had coronavirus disease-associated symptoms before the outbreak. Variations in mink-derived viral genomes showed between-mink transmission and no infection link between the farms. Inhalable dust contained viral RNA, indicating possible exposure of workers. One worker is assumed to have attracted the virus from mink.
Little is known about the intermolecular interactions between the viral proteins of infectious bu... more Little is known about the intermolecular interactions between the viral proteins of infectious bursal disease virus (IBDV). By using the yeast two-hybrid system, which allows the detection of proteināprotein interactions in vivo, all possible interactions were tested by fusing the viral proteins to the LexA DNA-binding domain and the B42 transactivation domain. A heterologous interaction between VP1 and VP3, and homologous interactions of pVP2, VP3, VP5 and possibly VP1, were found by co-expression of the fusion proteins in Saccharomyces cerevisiae. The presence of the VP1āVP3 complex in IBDV-infected cells was confirmed by co-immunoprecipitation studies. Kinetic analyses showed that the complex of VP1 and VP3 is formed in the cytoplasm and eventually is released into the cell-culture medium, indicating that VP1āVP3 complexes are present in mature virions. In IBDV-infected cells, VP1 was present in two forms of 90 and 95 kDa. Whereas VP3 initially interacted with both the 90 and 95 ...
Background: Highly pathogenic avian influenza virus (HPAIV) causes a highly contagious often fata... more Background: Highly pathogenic avian influenza virus (HPAIV) causes a highly contagious often fatal disease in poultry, resulting in significant economic losses in the poultry industry. HPAIV H5N1 also poses a major public health threat as it can be transmitted directly from infected poultry to humans. One effective way to combat avian influenza with pandemic potential is through the vaccination of poultry. Several live vaccines based on attenuated Newcastle disease virus (NDV) that express influenza hemagglutinin (HA) have been developed to protect chickens or mammalian species against HPAIV. However, the zoonotic potential of NDV raises safety concerns regarding the use of live NDV recombinants, as the incorporation of a heterologous attachment protein may result in the generation of NDV with altered tropism and/or pathogenicity. Methodology/Principal Findings: In the present study we generated recombinant NDVs expressing either full length, membrane-anchored HA of the H5 subtype (...
Little is known about the intermolecular interactions between the viral proteins of infectious bu... more Little is known about the intermolecular interactions between the viral proteins of infectious bursal disease virus (IBDV). By using the yeast two-hybrid system, which allows the detection of proteināprotein interactions in vivo, all possible interactions were tested by fusing the viral proteins to the LexA DNA-binding domain and the B42 transactivation domain. A heterologous interaction between VP1 and VP3, and homologous interactions of pVP2, VP3, VP5 and possibly VP1, were found by co-expression of the fusion proteins in Saccharomyces cerevisiae. The presence of the VP1āVP3 complex in IBDV-infected cells was confirmed by co-immunoprecipitation studies. Kinetic analyses showed that the complex of VP1 and VP3 is formed in the cytoplasm and eventually is released into the cell-culture medium, indicating that VP1āVP3 complexes are present in mature virions. In IBDV-infected cells, VP1 was present in two forms of 90 and 95 kDa. Whereas VP3 initially interacted with both the 90 and 95 ...
Inoculation of infl uenza (H5N1) into beagles resulted in virus excretion and rapid seroconversio... more Inoculation of infl uenza (H5N1) into beagles resulted in virus excretion and rapid seroconversion with no disease. Binding studies that used labeled infl uenza (H5N1) showed virus attachment to higher and lower respiratory tract tissues. Thus, dogs that are subclinically infected with infl u-enza (H5N1) may contribute to virus spread. Avian influenza (H5N1) virus has been shown to be infectious not only for birds but also for humans and mammals such as mice, ferrets, and cats. Carnivorous mammals that are susceptible to subtype H5N1 may contribute to spread of the virus; shedding of influenza (H5N1) by pet carnivores may pose a risk to humans. Cats experimentally inoculated with influenza (H5N1) have been shown to be susceptible to infection and to shed virus (1). However, dogs ā susceptibility to this virus is unknown. Unpublished studies indicate that a substantial number of dogs tested in Thailand were positive for antibodies against H5N1 subtype (2). Recently, isolation of infl...
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