RESUMO
FMDV O1 subtype undergoes antigenic variation under diverse growth conditions. Of particular interest is the amino acid variation observed at position 56 within the structural protein VP3. Selective pressures influence whether histidine (H) or arginine (R) is present at this position, ultimately influencing in vitro plaque morphology and in vivo pathogenesis in cattle. Using reverse genetics techniques, we have constructed FMDV type O1 Campos variants differing only at VP3 position 56, possessing either an H or R (O1Ca-VP3-56H and O1Ca-VP3-56R, respectively), and characterized their in vitro phenotype and virulence in the natural host. Both viruses showed similar growth kinetics in vitro. Conversely, they had distinct temperature-sensitivity (ts) and displayed significantly different pathogenic profiles in cattle and swine. O1Ca-VP3-56H was thermo stable and induced typical clinical signs of FMD, whereas O1Ca-VP3-56R presented a ts phenotype and was nonpathogenic unless VP3 position 56 reverted in vivo to either H or cysteine (C).
Assuntos
Proteínas do Capsídeo/química , Vírus da Febre Aftosa/patogenicidade , Febre Aftosa/virologia , Sequência de Aminoácidos , Animais , Variação Antigênica , Sequência de Bases , Proteínas do Capsídeo/biossíntese , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Bovinos , Linhagem Celular , Cricetinae , Vírus da Febre Aftosa/química , Vírus da Febre Aftosa/genética , Vírus da Febre Aftosa/imunologia , Variação Genética , Testes de Neutralização , Fenótipo , RNA Viral/genética , Análise de Sequência de RNA , Ovinos , Suínos , TemperaturaRESUMO
To explore the role in viral pathogenesis of the region located between the two functional AUG (inter-AUG) in foot-and-mouth disease virus (FMDV), we derived viruses containing transposon (tn) inserts from a mutagenized cDNA infectious clone of FMDV (pA24-WT). Mutant viruses containing an in-frame 57-nt transposon insertion grew at a slower rate and had a smaller plaque size phenotype than the parental virus (A24-WT). A mutant virus containing a 51-nt deletion in inter-AUG had a similar phenotype in cell culture to that of A24-WT. When tested by aerosol inoculation in cattle (3 animals per virus), the deletion mutant was fully virulent as was A24-WT. Mutant viruses containing insertions in inter-AUG did not cause clinical disease or viremia. However, viruses that partially or totally removed the tn insertion during animal infection reverted to virulence in 2 inoculated steers. Therefore, this study identified inter-AUG as an FMDV viral virulence determinant in cattle infected by aerosol route.
Assuntos
Códon , Vírus da Febre Aftosa/genética , Vírus da Febre Aftosa/patogenicidade , Poliproteínas/genética , Proteínas Virais/genética , Sequência de Aminoácidos , Animais , Bovinos , Dados de Sequência Molecular , Mutação , Poliproteínas/biossíntese , Biossíntese de Proteínas , Virulência , Replicação ViralRESUMO
The use of transgenic plants as vectors for the expression of viral and bacterial antigens has been increasingly tested as an alternative methodology for the production of experimental vaccines. Here, we report the production of transgenic alfalfa plants containing the genes encoding the polyprotein P1 and the protease 3C of foot and mouth disease virus (FMDV). The immunogenicity of the expressed products was tested using a mouse experimental model. Parenterally immunized mice developed a strong antibody response and were completely protected when challenged with the virulent virus. This report demonstrates the possibility of using transgenic plants to express polyprotein P1 and the protease 3C of FMDV and their utilization as effective experimental immunogens.
Assuntos
Vírus da Febre Aftosa/imunologia , Febre Aftosa/imunologia , Medicago sativa/genética , Plantas Geneticamente Modificadas/genética , Vacinas Virais/uso terapêutico , Agrobacterium tumefaciens/genética , Animais , Capsídeo/imunologia , DNA Viral/genética , Febre Aftosa/prevenção & controle , Injeções Intraperitoneais , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transformação Genética , Vacinas Sintéticas/uso terapêutico , Vacinas Virais/biossínteseRESUMO
A procedure for the simultaneous determination of lead and cadmium in TiO(2) by differential pulse anodic stripping voltammetry (ASV) has been developed. The key feature of the method is the use of triethanolamine (TEA) to remove titanium interference: TiO(2) undergoes acidic digestion with HF/H(2)SO(4) at atmospheric pressure, TEA is added to the HCl solution of the residue and the solution is analysed using a standard ASV instrumentation, equipped with a hanging mercury drop electrode. The calibration curves for both lead and cadmium are linear up to 50 mugl(-1) of solution, and the detection limits are 1 mugl(-1), corresponding to 1 mugg(-1) of TiO(2). Method reliability was tested by comparing the results with those given by electrothermal atomic absorption spectroscopy. The method has been successfully applied for determination of both contaminants in powdered titanium dioxide (raw materials) and in titanium dioxide-containing cosmetics (sunscreen products).
RESUMO
Ratones inmunizados con antígeno 140 S de Aftovirus tipo C3 Arg84 fueron inoculados con Sarcoma 180 TG. Los líquidos ascíticos obtenidos por punción ventral contenían niveles de anticuerpos semejantes a los alcanzados por los sueros, según fueron determinados por las técnicas de seronertralización y ELISA; siendo sus volúmenes de 10 a 20 veces mayores que los correspondientes sueros sanguíneos