RESUMO
OBJECTIVE: To determine the candidate genes for engineering vaccine of Ascaris lumbricoides. METHODS: pMD18-T-ALAg and plasmid expression vector pET-28a(+) were digested with BamH I and EcoR I and linked to each other. The resultant plasmid pET-28a(+)-ALAg was transferred into E. coli BL21 (DE3) and its expression was induced with IPTG, and the recombinant ALAg(rALAg) was purified. A total of 30 mice were equally divided into 3 groups, the mice in each group were injected with rALAg-FCA, FCA and PBS respectively, then they were attacked by infectious eggs of Ascaris (3 600 per mouse). The IgG levels in sera of mice in each group were detected by indirect ELASA. RESULTS: rALAg was recognized by the sera from repeatedly Ascaris lumbricoides inoculated rabbits. The numbers of larvae of Ascaris lumbricoides from liver and lung of mice were 25.30 +/- 4.55 in the rALAg-FCA group and 57.60 +/- 5.76 in the PBS group, respectively, the former being the reducing rate of 69.26%, and the difference among the 3 groups showed statistical significances (P < 0.01). The IgG levels (A450 value) of the rALAg-FCA, FCA and PBS groups were 0.858 +/- 0.003, 0.149 +/- 0.004 and 0.134 +/- 0.004, respectively, there were statistical differences among them (P < 0.01). CONCLUSION: ALAg can be used as a candidate gene of genetic engineering vaccine of Ascaris lumbricoides.
Assuntos
Vetores Genéticos/genética , Vacinas Sintéticas/genética , Animais , Ascaríase/imunologia , Ascaris lumbricoides/isolamento & purificação , Eletroforese em Gel de Poliacrilamida , Engenharia Genética , Vetores Genéticos/imunologia , Camundongos , Coelhos , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Vacinas Sintéticas/imunologiaRESUMO
Cysteine proteinases 112 (EhCP112) of Entamoeba histolytica are considered important for ameba pathogenicity. The recombinant gene was obtained by cloning and expression of the EhCP112 gene in heterologous host Escherichia coli BL-21 (DE3), were used to evaluate their ability to induce immune protective responses in minipig against challenge infection in a minipig-E. histolytica model. There was a 46.29% reduction (P<0.001) in the group of recovery of challenged E. histolytica compared with that in the control group. Specific anti-EhCP112 antibodies from immune protected minipig had significantly higher levels of immunoglobulin G (IgG) (P<0.001). This is a first report demonstrating that a recombinant form of EhCP112 generated in E. coli, to immunize a minipig model of E. histolytica, and there is significant protection. This study may help to understand the EhCP112 for human in the future.
Assuntos
Cisteína Proteases/imunologia , Entamoeba histolytica/enzimologia , Entamebíase/prevenção & controle , Vacinas Protozoárias/normas , Animais , Anticorpos Antiprotozoários/sangue , Antígenos de Protozoários/imunologia , Western Blotting , Cisteína Proteases/genética , Modelos Animais de Doenças , Entamoeba histolytica/genética , Entamoeba histolytica/imunologia , Feminino , Soros Imunes/imunologia , Imunoglobulina G/sangue , Coelhos , Suínos , Porco Miniatura , Vacinas Sintéticas/normasRESUMO
Cysteine proteinases 4 (EhCP4) of Entamoeba histolytica are considered important for ameba pathogenicity. The recombinant gene was obtained by cloning and expression of the EhCP4 gene in heterologous host Escherichia coli BL-21 (DE3), were used to evaluate their ability to induce immune protective responses in minipig against challenge infection in a minipig-E. histolytica model. There was a 53.16% reduction (P<0.001) in the group of recovery of challenged E. histolytica compared with that in the control group. Specific anti-EhCP4 antibodies from immune protected minipig had significantly higher levels of immunoglobulin G (IgG) (P<0.001). This is a first report demonstrating that a recombinant form of EhCP4 generated in E. coli, to immunize a minipig model of E. histolytica, and there is significant protection. This study may help to understand the EhCP4 for human in the future.
Assuntos
Antígenos de Protozoários/metabolismo , Cisteína Proteases/metabolismo , Entamoeba histolytica/enzimologia , Entamoeba histolytica/imunologia , Entamebíase/prevenção & controle , Vacinas Protozoárias , Animais , Antígenos de Protozoários/genética , Antígenos de Protozoários/imunologia , Clonagem Molecular , Cisteína Proteases/genética , Cisteína Proteases/imunologia , Entamoeba histolytica/patogenicidade , Feminino , Regulação Enzimológica da Expressão Gênica , Modelos Animais , Coelhos , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , Suínos , Porco MiniaturaRESUMO
We used a real-time PCR assay and indirect fluorescent antibody (IFA) assay to detect genomic DNA of Salmonella Enteritidis in the internal organs of quails after an oral challenge. The results showed that S. Enteritidis was detected in all the samples at different time points. This study will assist a future understanding of the pathogenesis of S. Enteritidis.
Assuntos
Doenças das Aves/microbiologia , Codorniz/microbiologia , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Salmonelose Animal/microbiologia , Salmonella enteritidis/patogenicidade , Animais , Doenças das Aves/genética , Técnica Indireta de Fluorescência para Anticorpo/veterinária , Salmonelose Animal/genética , Salmonelose Animal/patologia , Salmonella enteritidis/genéticaRESUMO
This research was undertaken to determine the population of a high-virulence strain of Salmonella enterica serovar Enteritidis in partridge by a fluorescent quencher PCR assay and to correlate these findings with the results obtained from the immunohistochemical localization and histopathological examinations of selected Salmonella enterica serovar Enteritidis-infected tissues. To make the results meaningful, a side-by-side bacteriology method (indirect immuno-fluorescent antibody staining) was performed too. The results of indirect immuno-fluorescent antibody staining and immunohistochemical localization were similar to the fluorescent quencher PCR assay. The time course of the appearance of bacterial antigens and tissue lesions in various tissues was coincident with the levels of the bacterial DNA loads at the infection sites. This suggests that Salmonella enterica serovar Enteritidis loads in internal organs are closely correlated with the progression of the infection.
