Faeces as a novel material to estimate lyssavirus prevalence in bat populations.

Rabies is caused by infection with a lyssavirus. Bat rabies is of concern for both public health and bat conservation. The current method for lyssavirus prevalence studies in bat populations is by oral swabbing, which is invasive for the bats, dangerous for handlers, time-consuming and expensive.

PCR diagnostics: In silico validation by an automated tool using freely available software programs.

PCR diagnostics are often the first line of laboratory diagnostics and are regularly designed to either differentiate between or detect all pathogen variants of a family, genus or species. The ideal PCR test detects all variants of the target pathogen, including newly discovered and emerging variants, while closely related pathogens and their variants should not be detected. This is challenging as pathogens show a high degree of genetic variation due to genetic drift, adaptation and evolution.

Detection and Characterization of Distinct Alphacoronaviruses in Five Different Bat Species in Denmark.

Bat populations harbour a multitude of viruses; some of these are pathogenic or potentially pathogenic in other animals or humans. Therefore, it is important to monitor the populations and characterize these viruses. In this study, the presence of coronaviruses (CoVs) in different species of Danish bats was investigated using active surveillance at different geographical locations in Denmark.

Molecular Epidemiology and Evolution of European Bat Lyssavirus 2.

Bat rabies cases in Europe are mainly attributed to two lyssaviruses, namely European Bat Lyssavirus 1 (EBLV-1) and European Bat Lyssavirus 2 (EBLV-2). Prior to the death of a bat worker in Finland in 1985, very few bat rabies cases were reported. Enhanced surveillance in the two subsequent years (1986-1987) identified 263 cases (more than a fifth of all reported cases to date).

Evaluation of a multiplex real-time PCR for detection of four bacterial agents commonly associated with bovine respiratory disease in bronchoalveolar lavage fluid.

Background: Pasteurella multocida, Mannheimia haemolytica, Histophilus somni and Trueperella pyogenes are four bacterial agents commonly associated with bovine respiratory disease (BRD). In this study a bacterial multiplex real-time PCR (the RespoCheck PCR) was evaluated for the detection in bronchoalveolar lavage fluid (BALF) of these four bacterial agents.

Results: The analytical sensitivity of the multiplex real-time PCR assay determined on purified DNA and on bacterial cells of the four target pathogens was one to ten fg DNA/assay and 4 × 10 to 2 × 10 CFU/assay.

Mycoplasma detection by triplex real-time PCR in bronchoalveolar lavage fluid from bovine respiratory disease complex cases.

Background: In this study we evaluated the RespoCheck Mycoplasma triplex real-time PCR for the detection in bronchoalveolar lavage fluid (BALF) of Mycoplasma (M.) dispar, M. bovis and M.

Molecular double-check strategy for the identification and characterization of European Lyssaviruses.

The "gold standard" for post-mortem rabies diagnosis is the direct fluorescent antibody test (FAT). However, in the case of ante-mortem non-neural sample material or decomposed tissues, the FAT reaches its limit, and the use of molecular techniques can be advantageous. In this study, we developed and validated a reverse transcription PCR cascade protocol feasible for the classification of samples, even those for which there is no epidemiological background knowledge.

A step forward in molecular diagnostics of lyssaviruses--results of a ring trial among European laboratories.

Rabies is a lethal and notifiable zoonotic disease for which diagnostics have to meet the highest standards. In recent years, an evolution was especially seen in molecular diagnostics with a wide variety of different detection methods published. Therefore, a first international ring trial specifically designed on the use of reverse transcription polymerase chain reaction (RT-PCR) for detection of lyssavirus genomic RNA was organized.

Schmallenberg virus in Culicoides spp. biting midges, the Netherlands, 2011.

To determine which species of Culicoides biting midges carry Schmallenberg virus (SBV), we assayed midges collected in the Netherlands during autumn 2011. SBV RNA was found in C. scoticus, C.

Circulation of group 2 coronaviruses in a bat species common to urban areas in Western Europe.

Fecal samples of 211 bats representing 13 different bat species from 31 locations in the Netherlands were analyzed for the presence of coronaviruses (CoV) using a genus-wide reverse transcription (RT)-polymerase chain reaction. CoVs are known for their high potential for interspecies transmission, including zoonotic transmission with bats as reservoir hosts. For the first time, a group 2 CoV was found in a bat, Pipistrellus pipistrellus, in Europe.

A stable full-length yellow fever virus cDNA clone and the role of conserved RNA elements in flavivirus replication.

Yellow fever virus (YF) is the prototype member of the Flavivirus genus. Here, we report the successful construction of a full-length infectious cDNA clone of the vaccine strain YF-17D. YF cDNA was cloned into a low-copy-number plasmid backbone and stably maintained in several E.

Yellow fever virus replicons as an expression system for hepatitis C virus structural proteins.

Chimeric yellow fever virus (YF) RNAs were constructed in which the YF structural genes were replaced by the hepatitis C virus (HCV) structural genes or fusions between the YF and HCV structural genes. Interestingly, RNA replication required nucleotide complementarity between the 3'-located conserved sequence 1 and an RNA sequence located in the 5' end of the YF capsid sequence. The (chimeric-)HCV structural proteins were efficiently expressed and processed, and the native E1/E2 heterodimer was formed.