Use of organic solvent to prevent protein adsorption in CE-MS experiments.

Protein adsorption onto capillary wall often hampers CE separations, particularly in the CZE mode. Electrostatic interactions are not the only factors affecting adsorption, as hydrophobic interactions and/or protein conformational changes are also involved in the adsorption phenomenon. Numerous methods can be used to reduce or avoid adsorption, such as (i) addition of low molecular weight molecules in the BGE, (ii) use of surfactants, or (iii) capillary coatings. However, most of these methods are not MS-compatible. In this study, we evaluated the addition of organic solvent as an alternative MS-compatible method to decrease protein adsorption. The effect of the solvent addition was emphasized using classical methods for estimating reversible and irreversible adsorption. In many cases, organic solvents were effective at decreasing adsorption. However, the influence of the organic solvent on protein adsorption should be evaluated case-by-case in CE method development.

Very virulent infectious bursal disease virus: reduced pathogenicity in a rare natural segment-B-reassorted isolate.

The purpose of this study was to compare the molecular epidemiology of infectious bursal disease virus (IBDV) segments A and B of 50 natural or vaccine IBDV strains that were isolated or produced between 1972 and 2002 in 17 countries from four continents, with phenotypes ranging from attenuated to very virulent (vv). These strains were subjected to sequence and phylogenetic analysis based on partial sequences of genome segments A and B. Although there is co-evolution of the two genome segments (70 % of strains kept the same genetic relatives in the segment A- and B-defined consensus trees), several strains (26 %) were identified with the incongruence length difference test as exhibiting a significantly different phylogenetic relationship depending on which segment was analysed. This suggested that natural reassortment could have occurred. One of the possible naturally occurring reassortant strains, which exhibited a segment A related to the vvIBDV cluster whereas its segment B was not, was thoroughly sequenced (coding sequence of both segments) and submitted to a standardized experimental characterization of its acute pathogenicity. This strain induced significantly less mortality than typical vvIBDVs; however, the mechanisms for this reduced pathogenicity remain unknown, as no significant difference in the bursal lesions, post-infectious antibody response or virus production in the bursa was observed in challenged chickens.

Extensive antigenic changes in an atypical isolate of very virulent infectious bursal disease virus and experimental clinical control of this virus with an antigenically classical live vaccine.

The 99323 Egyptian isolate of infectious bursal disease (IBD) virus (IBDV) was identified during an international survey of acute IBD cases. Its unique antigenicity was characterized by a markedly reduced binding of neutralizing monoclonal antibodies 3, 4, 5, 6, 8 and 9 in an antigen-capture enzyme-linked immunosorbent assay. Nucleotide sequencing of the genome region encoding the VP2 major immunogenic domain in 99323 revealed amino acid changes occurring at positions critical for antigenicity, but phylogenetic analysis demonstrated that 99323 was related to typical, very virulent IBDV (e.g. isolate 89163). Protection experimentally afforded by an antigenically classical live IBD vaccine was investigated in specific pathogen free chickens challenged with 99323 or 89163. Both viruses were similarly controlled, as evaluated by clinical signs, growth retardation, bursa-to-body weight ratios and histological lesions of the bursa after challenge. These results document that an active antibody response to a classical live antigen may clinically control infection by an antigenically atypical very virulent IBDV.