RESUMO
The emergence of West Nile virus (WNV) and Usutu virus (USUV) in Europe resulted in significant outbreaks leading to avifauna mortality and human infections. Both viruses have overlapping geographical, host and vector ranges, and are often co-circulating in Europe. In Germany, a nationwide bird surveillance network was established to monitor these zoonotic arthropod-borne viruses in migratory and resident birds. In this framework, co-infections with WNV and USUV were detected in six dead birds collected in 2018 and 2019. Genomic sequencing and phylogenetic analyses classified the detected WNV strains as lineage 2 and the USUV strains as lineages Africa 2 (n = 2), Africa 3 (n = 3) and Europe 2 (n = 1). Preliminary attempts to co-propagate both viruses in vitro failed. However, we successfully cultivated WNV from two animals. Further evidence for WNV-USUV co-infection was obtained by sampling live birds in four zoological gardens with confirmed WNV cases. Three snowy owls had high neutralizing antibody titres against both WNV and USUV, of which two were also positive for USUV-RNA. In conclusion, further reports of co-infections in animals as well as in humans are expected in the future, particularly in areas where both viruses are present in the vector population.
Assuntos
Doenças das Aves , Coinfecção , Infecções por Flavivirus , Flavivirus , Febre do Nilo Ocidental , Vírus do Nilo Ocidental , Animais , Doenças das Aves/epidemiologia , Aves , Coinfecção/epidemiologia , Coinfecção/veterinária , Flavivirus/genética , Infecções por Flavivirus/veterinária , Alemanha/epidemiologia , Filogenia , Febre do Nilo Ocidental/epidemiologia , Febre do Nilo Ocidental/veterinária , Vírus do Nilo Ocidental/genéticaRESUMO
Infectious pancreatic necrosis virus (IPNV) causes great losses in fish hatcheries world-wide. The detection of IPNV can be challenging in certain circumstances, particularly due to low viral load and the genetic variability of this RNA virus. For the first time, this project created a quantitative triplex real-time reverse transcription PCR (RT-qPCR), including an endogenous control system, for specific, sensitive and rapid detection of IPNV in routine diagnostics. Multiple sequence alignment of 46 nucleotide sequences of the segment A genome obtained from the NCBI database allowed the design of two RT-qPCR systems covering the IPNV genogroup 1 and genogroups 2-5, respectively. The completed triplex RT-qPCR including a salmonid-specific endogenous control showed high specificity and an analytical sensitivity of 20-40 oligonucleotide copies. Testing of dilution series of virus-loaded cell culture suspensions proved equality of the triplex RT-qPCR with virus detection in cell culture and a higher sensitivity than conventional RT-PCR in field samples. In comparative studies of a total of 77 field samples tested, 51 showed identical positive and 19 identical negative results in cell culture and the triplex RT-qPCR. However, seven other samples yielded positive results in the triplex RT-qPCR, but negative results in cell culture.
Assuntos
Genótipo , Vírus da Necrose Pancreática Infecciosa/isolamento & purificação , Reação em Cadeia da Polimerase Multiplex/métodos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Vírus da Necrose Pancreática Infecciosa/classificação , Vírus da Necrose Pancreática Infecciosa/genética , Reação em Cadeia da Polimerase Multiplex/normas , Reação em Cadeia da Polimerase em Tempo Real/normas , Padrões de Referência , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normas , Sensibilidade e EspecificidadeRESUMO
West Nile virus (WNV) is a mosquito-borne viral pathogen of global importance and is considered to be the most widespread flavivirus in the World. Horses, as dead-end hosts, can be infected by bridge mosquito vectors and undergo either subclinical infections or develop severe neurological diseases. The aim of this study was to detect WNV specific antibodies in horses in Germany as an indicator for an endemic circulation of WNV. Sera from more than 5,000 horses (primarily fallen stock animals) were collected in eight different federal states of Germany from 2010 to 2012. Sera were screened by a competitive ELISA and positive reactions were verified by an indirect IgM ELISA and/or by virus neutralization tests (VNT) for WNV and Tick-borne encephalitis virus (TBEV) in order to exclude cross-reacting antibody reactions. In essence WNV specific antibodies could not be detected in any of the horse sera. Not surprisingly, a small number of sera contained antibodies against TBEV. It is noteworthy that equine sera were often collected from horse carcasses and therefore were of poor quality. Nonetheless, these sera were still suitable for WNV ELISA testing, i.e., they did not produce a high background reaction which is a frequently observed phenomenon. According to these data there is no evidence for indigenous WNV infections in horses in Germany at present.
Assuntos
Anticorpos Antivirais/sangue , Ensaio de Imunoadsorção Enzimática/métodos , Doenças dos Cavalos/epidemiologia , Febre do Nilo Ocidental/veterinária , Vírus do Nilo Ocidental/imunologia , Animais , Especificidade de Anticorpos , Reações Cruzadas , Vírus da Encefalite Transmitidos por Carrapatos/imunologia , Flavivirus , Alemanha/epidemiologia , Cavalos , Estudos Soroepidemiológicos , Febre do Nilo Ocidental/epidemiologiaRESUMO
Reemerging bluetongue virus serotype 8 (BTV-8) in Germany was detected first in May 2007 in a sentinel cow and in February 2008 in an export heifer. Reemergence was confirmed by retesting the samples, experimental inoculation, fingerprinting analysis, and virus isolation. Overwintering of BTV-8 and continuous low-level infections are assumed.
