RESUMO
Se desarrolló una metodología para la selección de cepas de Vibrio cholerae O1 y O139 modificadas genéticamente, con el objetivo de obtener candidatos vacunales atenuados orales contra el cólera. A las cepas modificadas se les realizó caracterización microbiológica, susceptibilidad bacteriana y diferentes pruebas biológicas (dosis letal media, capacidad colonizadora, adherencia en ratones, intestino ligado e inoculación intraduodenal en conejos como pruebas de virulencia y potencia). Las cepas 81, 638, 638T y 1333 fueron evaluadas en ensayos clínicos para determinar su reactogenicidad e inmunogenicidad. Todas las cepas fueron sensibles a la tetraciclina y doxiclicina y mostraron su atenuación e inmunogenicidad en modelos animales, resultando las cepas 638 y 1333 inmunogénicas y no reactogénicas en voluntarios(AU)
Assuntos
Vibrio cholerae/imunologia , Vacinas , Modelos Animais , Ensaios Clínicos como AssuntoRESUMO
Se desarrolló una metodología para la selección de cepas de Vibrio cholerae O1 y O139 modificadas genéticamente, con el objetivo de obtener candidatos vacunales atenuados orales contra el cólera. A las cepas modificadas se les realizó caracterización microbiológica, susceptibilidad bacteriana y diferentes pruebas biológicas (dosis letal media, capacidad colonizadora, adherencia en ratones, intestino ligado e inoculación intraduodenal en conejos como pruebas de virulencia y potencia). Las cepas 81, 638, 638T y 1333 fueron evaluadas en ensayos clínicos para determinar su reactogenicidad e inmunogenicidad. Todas las cepas fueron sensibles a la tetraciclina y doxiclicina y mostraron su atenuación e inmunogenicidad en modelos animales, resultando las cepas 638 y 1333 inmunogénicas y no reactogénicas en voluntarios
Assuntos
Ensaios Clínicos como Assunto , Modelos Animais , Vacinas , Vibrio choleraeRESUMO
Vibrio cholerae 638 is a living candidate cholera vaccine strain attenuated by deletion of the CTXPhi prophage from C7258 (O1, El Tor Ogawa) and by insertion of the Clostridium thermocellum endoglucanase A gene into the hemagglutinin/protease coding sequence. This vaccine candidate was previously found to be well tolerated and immunogenic in volunteers. This article reports a randomized, double-blind, placebo-controlled trial conducted to test short-term protection conferred by 638 against subsequent V. cholerae infection and disease in volunteers in Cuba. A total of 45 subjects were enrolled and assigned to receive vaccine or placebo. The vaccine contained 10(9) CFU of freshly harvested 638 buffered with 1.3% NaHCO(3), while the placebo was buffer alone. After vaccine but not after placebo intake, 96% of volunteers had at least a fourfold increase in vibriocidal antibody titers, and 50% showed a doubling of at least the lipopolysaccharide-specific immunoglobulin A titers in serum. At 1 month after vaccination, five volunteers from the vaccine group and five from the placebo group underwent an exploratory challenge study with 10(9) CFU of DeltaCTXPhi attenuated mutant strain V. cholerae 81. Only two volunteers from the vaccine group shed strain 81 in their feces, but none of them experienced diarrhea; in the placebo group, all volunteers excreted the challenge strain, and three had reactogenic diarrhea. An additional 12 vaccinees and 9 placebo recipients underwent challenge with 7 x 10(5) CFU of virulent strain V. cholerae 3008 freshly harvested from a brain heart infusion agar plate and buffered with 1.3% NaHCO(3). Three volunteers (25%) from the vaccine group and all from the placebo group shed the challenge agent in their feces. None of the 12 vaccinees but 7 volunteers from the placebo group had diarrhea, and 2 of the latter exhibited severe cholera (>5,000 g of diarrheal stool). These results indicate that at 1 month after ingestion of a single oral dose (10(9) CFU) of strain 638, volunteers remained protected against cholera infection and disease provoked by the wild-type challenge agent V. cholerae 3008. We recommend that additional vaccine lots of 638 be prepared under good manufacturing practices for further evaluation.
Assuntos
Vacinas contra Cólera/administração & dosagem , Vacinas contra Cólera/imunologia , Cólera/prevenção & controle , Vibrio cholerae/imunologia , Administração Oral , Adolescente , Adulto , Anticorpos Antibacterianos/sangue , Bacteriófagos/genética , Celulase/genética , Vacinas contra Cólera/genética , Clostridium thermocellum , Método Duplo-Cego , Fezes/microbiologia , Deleção de Genes , Hemaglutininas/genética , Humanos , Masculino , Peptídeo Hidrolases/genética , Vacinação , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Vibrio cholerae/genética , Vibrio cholerae/patogenicidade , Vibrio cholerae/virologiaRESUMO
A methodology was developed for the selection of genetically modified strains of Vibrio cholerae 01 and 0139 aimed at obtaining oral attenuated candidate vaccines against cholera. The modified strains underwent microbiological characterization, bacterial susceptibility and different biological tests (mean lethal dose, colonizing capacity, adherence in mice, ligated intestine and intraduodenal inoculation in rabbits as virulence and potency tests. The strains 81, 638, 638T and 1333 were evaluated in clinical trials to determine their reactogenicity and immunogenicity. All the strains were sensitive to tetracycline and doxoclycine. They showed their attenuation and immunogenicity in animal models. The strains 638 and 1333 proved to be immunogenic and non reactogenic in volunteers.
Assuntos
Vacinas contra Cólera , Cólera/prevenção & controle , Vibrio cholerae/imunologia , Administração Oral , Análise de Variância , Animais , Vacinas contra Cólera/administração & dosagem , Vacinas contra Cólera/efeitos adversos , Vacinas contra Cólera/genética , Vacinas contra Cólera/imunologia , Vacinas contra Cólera/isolamento & purificação , Interpretação Estatística de Dados , Doxiciclina/farmacologia , Humanos , Camundongos , Microscopia Eletrônica de Transmissão , Modelos Animais , Coelhos , Tetraciclina/farmacologia , Vacinas Atenuadas , Vibrio cholerae/efeitos dos fármacos , Vibrio cholerae/genética , Vibrio cholerae/ultraestruturaRESUMO
The antimicrobial susceptibility and the presence of a heat-stable toxin were researched into 100 non-01 Vibrio cholerae strains sent by 7 different health centers to the National Reference Laboratory of Acute Diarrheal Diseases in "Pedro KourÝ" Tropical Medicine Institute. The presence of 20 toxigenic non-01 Vibrio cholerae was detected, a figure substantially higher than that reported in other geographic areas, except for endemic areas. This result will make it possible to set epidemiological alert in Cuba because these strains can be infected by CTX phages (element transporting genes that encode for choleric toxin) which will give such strains an epidemic potential similar to that of the etiologic agent of cholera. The identified strains could be studied as possible cholera vaccine candidates.
Assuntos
Humanos , Animais , Lactente , Pré-Escolar , Camundongos , Toxina da Cólera/isolamento & purificação , Vibrio cholerae , Técnicas de Tipagem Bacteriana , Cuba , Fezes , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase , Sorotipagem , Toxina da Cólera/análise , Vibrio choleraeRESUMO
Se investigó la susceptibilidad antimicrobiana y la presencia de toxina termoestable en 100 cepas de Vibrio cholerae No-01 remitidas de 7 diferentes centros de salud del país al Laboratorio Nacional de Referencia de Enfermedades Diarreicas Agudas del Instituto de Medicina Tropical Pedro Kourí. Se demostró la presencia de 20 por ciento Vibrio cholerae No-01 toxigénico, esta cifra resultó considerablemente más alta que la reportada en otras áreas geográficas, excepto en zonas epidémicas. Este resultado permitirá establecer una alerta epidemiológica en Cuba, pues estas cepas pueden ser infectadas por el fago CTX (elemento que transporta los genes que codifican para la toxina colérica); lo que les conferiría un potencial epidémico similar al del agente etiológico del cólera. Las cepas identificadas podrían estudiarse como posibles candidatas a la vacuna del cólera(AU)
