Institute Virologia

Germany

Institute Virologia

Germany

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Pauly D.,Robert Koch Institute | Chacana P.A.,Institute Virologia | Brembs B.,Free University of Berlin | Schade R.,Charité - Medical University of Berlin
Journal of Visualized Experiments | Year: 2011

Hens can be immunized by means of i.m. vaccination (Musculus pectoralis, left and right, injection volume 0.5-1.0 ml) or by means of Gene-Gun plasmid-immunization. Dependent on the immunogenicity of the antigen, high antibody-titres (up to 1:100,000 - 1:1,000,000) can be achieved after only one or 3 - 4 boost immunizations. Normally, a hen lays eggs continuously for about 72 weeks, thereafter the laying capacity decreases. This protocol describes the extraction of total IgY from egg yolk by means of a precipitation procedure (PEG. Polson et al. 1980). The method involves two important steps. The first one is the removal of lipids and the second is the precipitation of total IgY from the supernatant of step one. After dialysis against a buffer (normally PBS) the IgY-extract can be stored at -20°C for more than a year. The purity of the extract is around 80%, the total IgY per egg varies from 40-80 mg, dependent on the age of the laying hen. The total IgY content increases with the age of the hen from around 40 mg/egg up to 80 mg/egg (concerning PEG precipitation). The laying capacity of a hen per year is around 325 eggs. That means a total potential harvest of 20 g total IgY/year based on a mean IgY content of 60 mg total IgY/egg (see Table 1). © 2011 Journal of Visualized Experiments.


Legisa D.M.,Institute Virologia | Perez Aguirreburualde M.S.,Institute Virologia | Gonzalez F.N.,Institute Virologia | Marin-Lopez A.,Research Center en Sanidad Animal | And 4 more authors.
Vaccine | Year: 2015

Bluetongue virus (BTV), the causative agent of bluetongue disease (BT) in domestic and wild ruminants, is worldwide distributed. A total of 27 serotypes have been described so far, and several outbreaks have been reported. Vaccination is critical for controlling the spread of BTV. In the last years, subunit vaccines, viral vector vaccines and reverse genetic-based vaccines have emerged as new alternatives to conventional ones. In this study, we developed an experimental subunit vaccine against BTV4, with the benefit of targeting the recombinant protein to antigen-presenting cells. The VP2 protein from an Argentine BTV4 isolate was expressed alone or fused to the antigen presenting cell homing (APCH) molecule, in the baculovirus insect cell expression system. The immunogenicity of both proteins was evaluated in guinea pigs and cattle. Titers of specific neutralizing antibodies in guinea pigs and cattle immunized with VP2 or APCH-VP2 were high and similar to those induced by a conventional inactivated vaccine. The immunogenicity of recombinant proteins was further studied in the IFNAR(-/-) mouse model where the fusion of VP2 to APCH enhanced the cellular immune response and the neutralizing activity induced by VP2. © 2015 Elsevier Ltd.


Bluetongue virus (BTV), the causative agent of bluetongue disease (BT) in domestic and wild ruminants, is worldwide distributed. A total of 27 serotypes have been described so far, and several outbreaks have been reported. Vaccination is critical for controlling the spread of BTV. In the last years, subunit vaccines, viral vector vaccines and reverse genetic-based vaccines have emerged as new alternatives to conventional ones. In this study, we developed an experimental subunit vaccine against BTV4, with the benefit of targeting the recombinant protein to antigen-presenting cells. The VP2 protein from an Argentine BTV4 isolate was expressed alone or fused to the antigen presenting cell homing (APCH) molecule, in the baculovirus insect cell expression system. The immunogenicity of both proteins was evaluated in guinea pigs and cattle. Titers of specific neutralizing antibodies in guinea pigs and cattle immunized with VP2 or APCH-VP2 were high and similar to those induced by a conventional inactivated vaccine. The immunogenicity of recombinant proteins was further studied in the IFNAR(-/-) mouse model where the fusion of VP2 to APCH enhanced the cellular immune response and the neutralizing activity induced by VP2.


Gillet N.A.,University of Liège | Hamaidia M.,University of Liège | de Brogniez A.,University of Liège | Gutierrez G.,Institute Virologia | And 4 more authors.
PLoS Pathogens | Year: 2016

Retroviruses are not expected to encode miRNAs because of the potential problem of self-cleavage of their genomic RNAs. This assumption has recently been challenged by experiments showing that bovine leukemia virus (BLV) encodes miRNAs from intragenomic Pol III promoters. The BLV miRNAs are abundantly expressed in B-cell tumors in the absence of significant levels of genomic and subgenomic viral RNAs. Using deep RNA sequencing and functional reporter assays, we show that miRNAs mediate the expression of genes involved in cell signaling, cancer and immunity. We further demonstrate that BLV miRNAs are essential to induce B-cell tumors in an experimental model and to promote efficient viral replication in the natural host. © 2016 Gillet et al.


PubMed | Institute Virologia, National Veterinary Research Institute and University of Liège
Type: Journal Article | Journal: PLoS pathogens | Year: 2016

Retroviruses are not expected to encode miRNAs because of the potential problem of self-cleavage of their genomic RNAs. This assumption has recently been challenged by experiments showing that bovine leukemia virus (BLV) encodes miRNAs from intragenomic Pol III promoters. The BLV miRNAs are abundantly expressed in B-cell tumors in the absence of significant levels of genomic and subgenomic viral RNAs. Using deep RNA sequencing and functional reporter assays, we show that miRNAs mediate the expression of genes involved in cell signaling, cancer and immunity. We further demonstrate that BLV miRNAs are essential to induce B-cell tumors in an experimental model and to promote efficient viral replication in the natural host.


Soria L.A.,University of Buenos Aires | Corva P.M.,University of the Sea | Huguet M.J.,University of Buenos Aires | Mino S.,Institute Virologia | Miquel M.C.,University of Buenos Aires
BAG - Journal of Basic and Applied Genetics | Year: 2010

The bovine CAPN1 gene encodes the large subunit of μ-calpain, which is thought to be one of the most important enzymes involved in postmortem beef tenderization. Three SNPs in CAPN1 (SNP 316, SNP 530 and SNP 4751) have been associated with beef tenderness in different beef cattle breeds. The objective of this work was to implement genotyping strategies for CAPN1 markers as part of a project pursuing the identifi cation and validation of molecular markers associated with bovine meat quality and composition. Three PCR-RFLP methods were designed to determine genotypes of 64 bulls (11 Angus, 43 Brangus and 10 Brahman). Unexpected patterns resulting from the PCR-RFLP analysis at SNP 316 and SNP 530 were resolved by cloning and sequencing and lead to the discovery of three substitutions not previously described. These mutations could be useful in population studies, such as the determination of the relative contribution of the Angus and Brahman breeds to the Brangus.


PubMed | Institute Virologia
Type: | Journal: BMC veterinary research | Year: 2012

Parainfluenza virus type 3 (PIV3) was isolated from dairy buffaloes (Bubalus bubalis) naturally affected with respiratory and reproductive clinical conditions.Examination of nasal and vaginal swabs collected from 12 diseased buffaloes led to the isolation of three paramyxovirus isolates from two animals. Antigenic, morphological and biological characteristics of these three isolates were essentially similar to those of members of the Paramyxoviridae family. Antigenic analysis by direct immunofluorescence and cross neutralization test placed these isolates together with bovine parainfluenza virus type 3 (BPIV3). Nucleotide and amino acid phylogenetic analysis of partial matrix gene sequences of the buffalo isolates and six field BPIV3 isolates from bovines in Argentina were studied. Buffalo isolates were similar to genotype B (BPIV3b) while the six BPIV3 isolates were similar to genotypes A (BPIV3a) and C (BPIV3c).This is the first characterization of BPIV3 in water buffalo.According to the samples analyzed, in Argentina, the genotype B was found in buffalo and the genotypes A and C were found in cattle.


PubMed | Institute Virologia
Type: Journal Article | Journal: Veterinary microbiology | Year: 2012

Bovine Group A Rotavirus (RVA) is one of the main causes of neonatal calf diarrhea worldwide. The present study reports the genotyping of bovine RVA strains circulating in Argentinean cattle from 2004 to 2010. Additionally, a new set of typing primers was designed and tested to differentiate between G8 and G6 (lineage III and IV) RVA strains. Bovine RVA was detected in 30% (435/1462) of the tested samples, corresponding to 49% (207/423) of the studied outbreaks with a similar detection rates in beef (53%; 67/127) and dairy herds (52%; 65/126). The RVA strains circulating in Argentinean cattle belonged to the common bovine genotypes G6 (lineages III and IV), G8, G10, P[5] and P[11]. A different RVA G/P-genotype distribution was found between the exploitation types, with the combination G6(IV)P[5] being by fare the most prevalent RVA strain in beef herds (58%), whereas a more even distribution of G6(III)P[11] (15%), G10P[11] (17%), G6(IV)P[5] (14%), and G6(IV)P[11] (6%) RVA strains was detected in dairy herds. G8 RVA strains were found in two dairy farms in calves co-infected with G8+G6(III)P[11]. A high percentage of co-infections and co-circulation of RVA strains with different genotypes during the same outbreak were registered in both exploitation types (20% of the outbreaks from beef herds and 23% from dairy herds), indicating a potential environment for reassortment. This finding is significant because G10P[11] and G6(III)P[11] strains may possess zoonotic potential. Continuous surveillance of the RVA strains circulating in livestock provides valuable information for a better understanding of rotavirus ecology and epidemiology.

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