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Serological and molecular investigation into the role of wild birds in the epidemiology of West Nile virus in Greece

George Valiakos12, Antonia Touloudi12, Labrini V Athanasiou23, Alexios Giannakopoulos1, Christos Iacovakis12, Periklis Birtsas45, Vassiliki Spyrou6, Zisis Dalabiras2, Liljana Petrovska7 and Charalambos Billinis12*

Author Affiliations

1 Laboratory of Microbiology and Parasitology, Faculty of Veterinary Medicine, University of Thessaly, 224 str. Trikalon, Karditsa, 43100, Greece

2 Laboratory of Zoonoses Research, Institute of Biomedical Research and Technology (BIOMED/CERETETH), Larissa, Greece

3 Department of Medicine, Faculty of Veterinary Medicine, University of Thessaly, Karditsa, Greece

4 Department of Forestry and Natural Environment Administration, Technological Education Institute of Larissa, Karditsa, Greece

5 Hunting Federation of Macedonia and Thrace, Thessaloniki, Greece

6 Department of Animal Production, Technological Education Institute of Larissa, Larissa, Greece

7 Department of Bacteriology, Animal Health and Veterinary Laboratories Agency, Weybridge, UK

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Virology Journal 2012, 9:266  doi:10.1186/1743-422X-9-266

Published: 12 November 2012



A West Nile virus (WNV) disease outbreak occurred in 2010 in northern Greece with a total of 262 laboratory-confirmed human cases and 35 deaths. A serological and molecular surveillance was conducted on samples of hunter-harvested wild birds prior to and during the outbreak.


Serum and tissue samples from 295 resident and migratory wild birds, hunter-harvested during the 2009–2010 and 2010–2011 hunting seasons at the epicenter of the outbreak in northern Greece, were tested for the presence of WNV-specific antibodies by immunofluorescence assay and virus neutralization test. WNV neutralizing antibodies were detected in 53 avian samples. Fourteen positive sera were obtained from birds hunter-harvested up to 8 months prior to the human outbreak. Specific genetic determinants of virulence (His249Pro NS3 mutation, E-glycosylation motif) were recognized in a WNV lineage 2 strain isolated from a hunter-harvested Eurasian magpie and a nucleotide mismatch was revealed between this strain and a mosquito WNV strain isolated one month earlier in the same area.


This is the first report regarding exposure of wild birds to WNV prior to the 2010 outbreak, in Greece. Results provide evidence of the implication of wild birds in a local enzootic cycle that could allow maintenance and amplification of the virus before and during the outbreak. Findings of past exposure of migratory birds to WNV upon their arrival in Greece during autumn migration, suggest avian species with similar migration traits as candidates for the introduction of WNV into Greece. The possibility that an endemic circulation of WNV could have caused the outbreak, after an amplification cycle due to favorable conditions cannot be excluded.

Greece; Molecular epidemiology; Outbreak; Serological surveillance; West nile virus; Wild birds