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Pathogenicity of two recent Western Mediterranean West Nile virus isolates in a wild bird species indigenous to Southern Europe: the red-legged partridge.

Sotelo E, Gutierrez-Guzmán AV, del Amo J, Llorente F, El-Harrak M, Pérez-Ramírez E, Blanco JM, Höfle U, Jiménez-Clavero MA - Vet. Res. (2011)

Bottom Line: West Nile virus (WNV) is an emerging zoonotic pathogen whose geographic spread and incidence in humans, horses and birds has increased significantly in recent years.All inoculated birds became viremic and showed clinical disease, with mortality rates of 70% and 30%, respectively.These results show that Western Mediterranean WNV variants can be pathogenic for some European bird species, such as the red-legged partridge.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centro de Investigación en Sanidad Animal del Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (CISA-INIA), Ctra Algete-El Casar, s/n, 28130 Valdeolmos (Madrid), Spain. majimenez@inia.es.

ABSTRACT
West Nile virus (WNV) is an emerging zoonotic pathogen whose geographic spread and incidence in humans, horses and birds has increased significantly in recent years. WNV has long been considered a mild pathogen causing self-limiting outbreaks. This notion has changed as WNV is causing large epidemics with a high impact on human and animal health. This has been particularly noteworthy since its introduction into North America in 1999. There, native bird species have been shown to be highly susceptible to WNV infection and disease with high mortalities. For this reason, the effect of WNV infection in North American bird species has been thoroughly studied by means of experimental inoculations in controlled trials. To a lesser extent, European wild birds have been shown to be affected clinically by WNV infection. Yet experimental studies on European wild bird species are lacking. The red-legged partridge (Alectoris rufa) is a gallinaceous bird indigenous to the Iberian Peninsula, widely distributed in South Western Europe. It plays a key role in the Mediterranean ecosystem and constitutes an economically important game species. As such it is raised intensively in outdoor facilities. In this work, red-legged partridges were experimentally infected with two recent WNV isolates from the Western Mediterranean area: Morocco/2003 and Spain/2007. All inoculated birds became viremic and showed clinical disease, with mortality rates of 70% and 30%, respectively. These results show that Western Mediterranean WNV variants can be pathogenic for some European bird species, such as the red-legged partridge.

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Mean daily blood viral genome load (a) and viremia titers (b), plotted for two groups of 10 red-legged partridges: one inoculated with WNV Morocco/2003 (open squares) and another inoculated with WNV Spain/2007 (closed circles). Each group was alternatively sampled every other day as described in the text. Each point represents the mean of five individuals, or of the surviving individuals of each group. Error bars represent the standard error of the mean. The efficiency of the semi-quantitative real-time RT-PCR assay for viral genome load determinations was E = 0.908.
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Figure 2: Mean daily blood viral genome load (a) and viremia titers (b), plotted for two groups of 10 red-legged partridges: one inoculated with WNV Morocco/2003 (open squares) and another inoculated with WNV Spain/2007 (closed circles). Each group was alternatively sampled every other day as described in the text. Each point represents the mean of five individuals, or of the surviving individuals of each group. Error bars represent the standard error of the mean. The efficiency of the semi-quantitative real-time RT-PCR assay for viral genome load determinations was E = 0.908.

Mentions: Both WNV strains assayed were pathogenic for red-legged partridges. All inoculated partridges showed clinical signs (e.g. loss of appetite, ruffled feathers, paralysis, and unresponsiveness), yet those infected with Morocco/2003 were more severely affected than those infected with Spain/2007. Firstly, the onset of symptoms began one day earlier (4 vs. 5 dpi) in partridges inoculated with Morocco/2003 than in those inoculated with Spain/2007. Secondly, mortality was higher (70% vs. 30%) and started earlier (5 vs. 6 dpi) with Morocco/2003 than with Spain/2007 (Figure 1). The differences observed in survival rates over time between both groups of inoculated partridges were statistically significant (p < 0.05). Clinical signs and mortality ceased by 9 dpi in both inoculated groups. Viral genome was detectable from 1 to 7 dpi (Figure 2a) in both inoculations. Viremia was detectable between 1 and 6 dpi for Spain/2007 and slightly longer (up to 7 dpi) for Morocco/2003. Peak viremia titers, and peak viral genome loads were found at 3 dpi for both isolates. Mean peak viremia was higher after inoculation with Morocco/2003 (107.2 pfu/mL) than with Spain/2007 (106.9 pfu/mL) (Figure 2b), similarly to what occurred with viral genome loads (Figure 2a) although in both cases the differences between both groups were not statistically significant (p > 0.05). All surviving partridges developed WNV-neutralizing antibodies (NtAb) (Figure 3), which were observed at 10 dpi or later. In inoculated partridges, specific antibodies to WNV as revealed by competitive ELISA were observed slightly earlier (6 dpi) than NtAb (Figure 3). Oral and cloacal virus shedding was observed between 3 and 7 dpi for both viruses. Virus shedding was more consistently found in oropharyngeal than in cloacal swabs for both strains, particularly in early stages (3 dpi, Figure 4). Post-mortem examination of lethally infected partridges revealed systemic infection, as the virus was detected in the brain, heart, spleen and liver by means of real time RT-PCR (Table 1). Upon necropsy, infected partridges were in poor body condition. Gross lesions were similar in both WNV-inoculated groups, and were characterized mainly by ascites, diffuse pallor of the myocardium, enlarged liver and petechia on the renal surface, injected blood vessels in the intestinal tract, and, in three birds, congestion of superficial cerebral vessels.


Pathogenicity of two recent Western Mediterranean West Nile virus isolates in a wild bird species indigenous to Southern Europe: the red-legged partridge.

Sotelo E, Gutierrez-Guzmán AV, del Amo J, Llorente F, El-Harrak M, Pérez-Ramírez E, Blanco JM, Höfle U, Jiménez-Clavero MA - Vet. Res. (2011)

Mean daily blood viral genome load (a) and viremia titers (b), plotted for two groups of 10 red-legged partridges: one inoculated with WNV Morocco/2003 (open squares) and another inoculated with WNV Spain/2007 (closed circles). Each group was alternatively sampled every other day as described in the text. Each point represents the mean of five individuals, or of the surviving individuals of each group. Error bars represent the standard error of the mean. The efficiency of the semi-quantitative real-time RT-PCR assay for viral genome load determinations was E = 0.908.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3037891&req=5

Figure 2: Mean daily blood viral genome load (a) and viremia titers (b), plotted for two groups of 10 red-legged partridges: one inoculated with WNV Morocco/2003 (open squares) and another inoculated with WNV Spain/2007 (closed circles). Each group was alternatively sampled every other day as described in the text. Each point represents the mean of five individuals, or of the surviving individuals of each group. Error bars represent the standard error of the mean. The efficiency of the semi-quantitative real-time RT-PCR assay for viral genome load determinations was E = 0.908.
Mentions: Both WNV strains assayed were pathogenic for red-legged partridges. All inoculated partridges showed clinical signs (e.g. loss of appetite, ruffled feathers, paralysis, and unresponsiveness), yet those infected with Morocco/2003 were more severely affected than those infected with Spain/2007. Firstly, the onset of symptoms began one day earlier (4 vs. 5 dpi) in partridges inoculated with Morocco/2003 than in those inoculated with Spain/2007. Secondly, mortality was higher (70% vs. 30%) and started earlier (5 vs. 6 dpi) with Morocco/2003 than with Spain/2007 (Figure 1). The differences observed in survival rates over time between both groups of inoculated partridges were statistically significant (p < 0.05). Clinical signs and mortality ceased by 9 dpi in both inoculated groups. Viral genome was detectable from 1 to 7 dpi (Figure 2a) in both inoculations. Viremia was detectable between 1 and 6 dpi for Spain/2007 and slightly longer (up to 7 dpi) for Morocco/2003. Peak viremia titers, and peak viral genome loads were found at 3 dpi for both isolates. Mean peak viremia was higher after inoculation with Morocco/2003 (107.2 pfu/mL) than with Spain/2007 (106.9 pfu/mL) (Figure 2b), similarly to what occurred with viral genome loads (Figure 2a) although in both cases the differences between both groups were not statistically significant (p > 0.05). All surviving partridges developed WNV-neutralizing antibodies (NtAb) (Figure 3), which were observed at 10 dpi or later. In inoculated partridges, specific antibodies to WNV as revealed by competitive ELISA were observed slightly earlier (6 dpi) than NtAb (Figure 3). Oral and cloacal virus shedding was observed between 3 and 7 dpi for both viruses. Virus shedding was more consistently found in oropharyngeal than in cloacal swabs for both strains, particularly in early stages (3 dpi, Figure 4). Post-mortem examination of lethally infected partridges revealed systemic infection, as the virus was detected in the brain, heart, spleen and liver by means of real time RT-PCR (Table 1). Upon necropsy, infected partridges were in poor body condition. Gross lesions were similar in both WNV-inoculated groups, and were characterized mainly by ascites, diffuse pallor of the myocardium, enlarged liver and petechia on the renal surface, injected blood vessels in the intestinal tract, and, in three birds, congestion of superficial cerebral vessels.

Bottom Line: West Nile virus (WNV) is an emerging zoonotic pathogen whose geographic spread and incidence in humans, horses and birds has increased significantly in recent years.All inoculated birds became viremic and showed clinical disease, with mortality rates of 70% and 30%, respectively.These results show that Western Mediterranean WNV variants can be pathogenic for some European bird species, such as the red-legged partridge.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centro de Investigación en Sanidad Animal del Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (CISA-INIA), Ctra Algete-El Casar, s/n, 28130 Valdeolmos (Madrid), Spain. majimenez@inia.es.

ABSTRACT
West Nile virus (WNV) is an emerging zoonotic pathogen whose geographic spread and incidence in humans, horses and birds has increased significantly in recent years. WNV has long been considered a mild pathogen causing self-limiting outbreaks. This notion has changed as WNV is causing large epidemics with a high impact on human and animal health. This has been particularly noteworthy since its introduction into North America in 1999. There, native bird species have been shown to be highly susceptible to WNV infection and disease with high mortalities. For this reason, the effect of WNV infection in North American bird species has been thoroughly studied by means of experimental inoculations in controlled trials. To a lesser extent, European wild birds have been shown to be affected clinically by WNV infection. Yet experimental studies on European wild bird species are lacking. The red-legged partridge (Alectoris rufa) is a gallinaceous bird indigenous to the Iberian Peninsula, widely distributed in South Western Europe. It plays a key role in the Mediterranean ecosystem and constitutes an economically important game species. As such it is raised intensively in outdoor facilities. In this work, red-legged partridges were experimentally infected with two recent WNV isolates from the Western Mediterranean area: Morocco/2003 and Spain/2007. All inoculated birds became viremic and showed clinical disease, with mortality rates of 70% and 30%, respectively. These results show that Western Mediterranean WNV variants can be pathogenic for some European bird species, such as the red-legged partridge.

Show MeSH
Related in: MedlinePlus