No abstract

This study explored a novel system combining plant-based production and the elastin-like peptide (ELP) fusion strategy to produce vaccinal antigens against tuberculosis. Transgenic tobacco plants expressing the mycobacterial antigens Ag85B and ESAT-6 fused to ELP (TBAg-ELP) were generated. Purified TBAg-ELP was obtained by the highly efficient, cost-effective, inverse transition cycling (ICT) method and tested in mice. Furthermore, safety and immunogenicity of the crude tobacco leaf extracts were assessed in piglets. Antibodies recognizing mycobacterial antigens were produced in mice and piglets. A T-cell immune response able to recognize the native mycobacterial antigens was detected in mice. These findings showed that the native Ag85B and ESAT-6 mycobacterial B- and T-cell epitopes were conserved in the plant-expressed TBAg-ELP. This study presents the first results of an efficient plant-expression system, relying on the elastin-like peptide fusion strategy, to produce a safe and immunogenic mycobacterial Ag85B-ESAT-6 fusion protein as a potential vaccine candidate against tuberculosis.

Microsporidia are fungal-like unicellular eukaryotes which develop as obligate intracellular parasites. They differentiate into resistant spores that are protected by a thick cell wall composed of glycoproteins and chitin. Despite an extensive description of the fibrillar structure of this wall, very little is known about its protein components and deposit mechanisms. In this study on the human pathogen Encephalitozoon cuniculi, we identify by mass spectrometry the target of polyclonal antibodies previously raised against a 33-kDa protein located at the outer face of the parasite plasma membrane. This 254-amino acid protein is encoded by the ECU11_0510 open reading frame and presents two isoforms of 33 and 55 kDa. Sequence analysis supports an assignment to the polysaccharide deacetylase family with a suspected chitin deacetylase activity (EcCDA). As demonstrated by TEM studies, EcCDA is present at the plasma membrane of the early stages of E. cuniculi life-cycle. At the sporoblast stage, the enzyme accumulates especially in paramural bodies which are convolutions of the plasma membrane opened to the wall. The identification of an EcCDA homologue in the insect parasite Antonospora locustae (ex Nosema locustae) suggests a widespread distribution of this enzyme among Microsporidia. This characterization of a new microsporidian surface protein creates new perspectives to understand spore wall formation and spore resistance.

Echovirus 30 is one of the enterovirus serotypes isolated most frequently in meningitis cases. The genetic diversity of echovirus 30 was investigated in patients hospitalised during an outbreak in 2000 in Clermont-Ferrand, France. A nested reverse transcription-PCR (RT-PCR) assay was developed for qualitative analysis of the echovirus 30 VP1 encoding sequence directly from cerebrospinal fluid. The viral sequences obtained for 22 patients were compared with those of virus isolates obtained from nine patients with echovirus 30 meningitis admitted to hospital in 1996-1997 and with echovirus 30 sequences from international databases. In 2000, meningitis cases were caused by two virus variants (C3 and C4) distinct genetically from the other two variants (C1 and C2) identified during the period 1996-1997. A detailed phylogenetic analysis established that the C1, C2, and C3 variants had close relatives among viruses previously identified in other geographical areas. The C4 variant had not been described earlier. The genomic differences observed between the four echovirus 30 variants arose at synonymous sites indicating that the viruses shared similar antigenic sites in the VP1 encoding sequence. Overall, these observations suggest wide circulation of different echovirus 30 variants and periodic importation of new viruses. The apparent displacement observed between virus variants did not result from a selective advantage caused by antigenic variation.