Attenuation of very virulent infectious bursal disease virus and comparison of full sequences of virulent and attenuated strains.
ABSTRACT A very virulent strain of infectious bursal disease virus (IBDVks) was isolated from the bursae of Fabricius of IBDV-affected broiler chickens. Following 43 serial passages in specific pathogen-free embryonated eggs, an attenuated strain was established (IBDVmb). Dosages of IBDVmb in the range 10(2) to 10(4) embryo infective dose of 50% were found to be safe and protective for commercial chicks. Chickens vaccinated with live vaccine containing IBDVmb responded with precipitating and type-specific neutralizing antibodies, and were immune to subsequent challenge with a very virulent IBDV. IBDVmb has been used as an attenuated vaccine throughout the world since 1993. A comparison of the full sequences of the virulent and attenuated strains (IBDVks and IBDVmb, respectively) revealed seven nucleotides that were different, four of them leading to changes in the amino-acid sequence. Comparison of the protein sequence of these strains and published sequences of very virulent and attenuated phenotypes lead us to suggest that the novel difference responsible for virulence of the Israeli strains are: residue 272 (VP2, very conserved site) and residue 527 (VP4), both in segment A, and in segment B (VP1) residues 96 and 161 (both conserved). Our study strengthens the possibility that more than one protein is involved in IBDV attenuation. In all reports, including ours, virulence was reduced without affecting antigenicity of the neutralizing epitopes in VP2. This could have practical implications for attenuated-vaccine development.
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ABSTRACT: Rapid and reliable detection and classification of infectious bursal disease viruses (IBDVs) is of crucial importance for disease surveillance and control. This study presents the development and validation of a real-time RT-PCR assay to detect and discriminate very virulent (vv) from non-vv (classic and variant) IBDV strains. The assay uses two fluorogenic, minor groove-binding (MGB) TaqMan probes targeted to a single nucleotide polymorphism (SNP) embedded in a highly conserved genomic region. The analytical sensitivity of the assay was determined using serial dilutions of in vitro-transcribed RNA. The assay demonstrated a wide dynamic range between 10(2) and 10(8) standard RNA copies per reaction. Good reproducibility was also detected, with intra- and inter-assay coefficients of variation ranging from 0.13% to 2.23% and 0.26% to 1.92%, respectively. The assay detected successfully all the assessed vv, classical, and variant field and vaccine strains and correctly discriminated all vvIBDV strains from non-vvIBDV strains. Other common avian RNA viruses tested negative, indicating high specificity of the assay. The high sensitivity, rapidity, reproducibility, and specificity of the real-time RT-PCR assay make this method suitable for general and genotype-specific detection and quantitation.Journal of virological methods 06/2012; 185(1):101-7. · 2.13 Impact Factor
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ABSTRACT: Infectious bursal disease virus (IBDV) causes an economically significant disease of chickens worldwide. Very virulent (vv) IBDV strains have emerged and induce up to 60% mortality. The molecular basis for vvIBDV pathogenicity is not understood and the relative contribution of the two genome segments A and B in this phenomenon is not known. Isolate 94432 was previously shown as being genetically related to vvIBDVs but exhibits an atypical antigenicity and does not cause mortality. Here, the full-length genome of 94432 was determined and a reverse genetics system was established. The molecular clone was rescued and exhibited the same antigenicity and reduced pathogenicity as 94432 isolate. Genetically modified viruses derived from 94432, whose vvIBDV consensus nucleotide sequence was restored in segment A and/or B, were produced and their pathogenicity assessed in specific pathogen free chickens. We found that a valine (position 321) that modifies the most exposed part of the capsid protein VP2 critically modified the antigenicity and partially reduced the pathogenicity of 94432. However, a threonine (position 276) located in the finger domain of the virus polymerase (VP1) contributed even more significantly to attenuation. This threonine is partially exposed in an hydrophobic groove on VP1 surface, which suggests possible interactions between VP1 and another, as yet unidentified, molecule at that this amino acid position. The restored vvIBDV-like pathogenicity was associated with increased replication and lesions in the thymus and spleen. These results demonstrate that both genome segments influence vvIBDV pathogenicity and may provide new targets for the attenuation of vvIBDVs.Journal of Virology 12/2012; · 4.65 Impact Factor
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ABSTRACT: Outbreaks of infectious bursal disease in vaccinated chicken flocks are frequent in Nigeria. For the control of infectious bursal disease, live vaccines based on foreign infectious bursal disease virus (IBDV) strains are used. The present study investigated the phylogenetic relationship between field and vaccine IBDV strains from northwestern Nigeria. Thirty field IBDV strains and three commercial vaccines strains were characterized through sequencing the VP2 hypervariable region. In addition, the complete genome segment A coding region for two vaccines and two field strains was sequenced. The deduced amino acid sequences (position 212 to 331) of IBDV strains from Nigeria and other regions of the world were aligned and possible regional and virulence markers were identified associated with VP2 minor hydrophilic peaks. Reversion to virulence of a vaccine strain with a Q to L mutation at position 253 was observed. Phylogenetic analyses revealed a unique cluster of northwest Nigerian field IBDV strains alone or related to imported characterized classical and very virulent IBDV vaccines. The results suggest that when IBDV strains spread from their region of origin to a different region they mutate alongside indigenous field strains but may retain their identity on the VP2 region.Avian Pathology 07/2013; · 2.04 Impact Factor