Epizootic Emergence of Usutu Virus in Wild and Captive Birds in Germany

Norbert Becker,Ute Ziegler,Hanna Jöst,Stefan Bosch,Bernd Hoffmann,Martin Beer,Christina Czajka,Petra Emmerich,Martin Eiden,Stephan Günther,Martin Pfeffer,Michael Wink,Martin H Groschup,Armin Konrad,Dirk W Höper ,Paul Rácz ,Deborah Ehichioya ,Marcin Gabriel ,Klara Tenner-Rácz ,Elisabeth Fichet‐Calvet ,Jonas Schmidt‐Chanasit

doi:10.1371/journal.pone.0032604

Abstract

This study aimed to identify the causative agent of mass mortality in wild and captive birds in southwest Germany and to gather insights into the phylogenetic relationship and spatial distribution of the pathogen. Since June 2011, 223 dead birds were collected and tested for the presence of viral pathogens. Usutu virus (USUV) RNA was detected by real-time RT-PCR in 86 birds representing 6 species. The virus was isolated in cell culture from the heart of 18 Blackbirds (Turdus merula). USUV-specific antigen was demonstrated by immunohistochemistry in brain, heart, liver, and lung of infected Blackbirds. The complete polyprotein coding sequence was obtained by deep sequencing of liver and spleen samples of a dead Blackbird from Mannheim (BH65/11-02-03). Phylogenetic analysis of the German USUV strain BH65/11-02-03 revealed a close relationship with strain Vienna that caused mass mortality among birds in Austria in 2001. Wild birds from lowland river valleys in southwest Germany were mainly affected by USUV, but also birds kept in aviaries. Our data suggest that after the initial detection of USUV in German mosquitoes in 2010, the virus spread in 2011 and caused epizootics among wild and captive birds in southwest Germany. The data also indicate an increased risk of USUV infections in humans in Germany.

Highlights

Usutu virus (USUV) is an arthropod-borne, single-stranded RNA virus and belongs to the Japanese encephalitis virus group within the family Flaviviridae
USUVspecific RNA was detected in the organs of 86 animals from 6 species using USUV-specific real-time reverse transcription–polymerase chain reaction (RT-PCR) (Table 1)
The genetic distance calculated with the complete nucleotide sequence of polyprotein between strain BH65/11-02-03 and the other European USUV strains Vienna 2001, Meise H from 2002, and Budapest from 2005 is $99%

Summary

Introduction

Usutu virus (USUV) is an arthropod-borne, single-stranded RNA virus and belongs to the Japanese encephalitis virus group within the family Flaviviridae. USUV is closely related to West Nile virus (WNV), which killed thousands of birds in North America following its emergence in 1999. USUV was demonstrated to circulate in several other European countries including Hungary, Switzerland, Spain and Italy [3,4]. No serological evidence for USUV circulation was found in birds from Germany between 2007 and 2011 [6,7]. In August 2010, USUV strain 1477 was isolated from a pool of Culex pipiens pipiens mosquitoes that were trapped in the city of Weinheim, southwest Germany [10]. All mosquitoes trapped the year before (during 2009) in southwest Germany tested negative for USUV [10]. No obvious increase in the mortality of wild and captive birds was observed in southwest Germany in 2009 and 2010 [10]

Objectives

Methods

Results

Conclusion