Immunization with a DNA vaccine candidate in chronic hepatitis C patients is safe, well tolerated and does not impair immune response induction after anti-hepatitis B vaccination.

BACKGROUND: In the present study, we evaluated the safety of CIGB-230, a novel vaccine candidate based on the mixture of a plasmid for DNA immunization, expressing hepatitis C virus (HCV) structural antigens, with a recombinant HCV Core protein. METHODS: Fifteen HCV chronically-infected volunteers with detectable levels of HCV RNA genotype 1b, who were nonresponders to previous treatment with interferon plus ribavirin, were intramuscularly injected with CIGB-230 on weeks 0, 4, 8, 12, 16 and 20. Individuals were also immunized at weeks 28, 32 and 36 with a recombinant vaccine against hepatitis B. Adverse events were recorded and analyzed. Blood samples were taken every 4 weeks up to month 12 for hematological, biochemical, virological and immunological analysis. RESULTS: All patients completed the treatment with CIGB-230. Adverse events were only slight (83.6%) or moderate (16.4%). No significant differences in hematological and biochemical parameters, including serum aminotransferases, were detected between the baseline and post-treatment state. Induction of a CD4+ T lymphocyte response against a particular region in HCV E1, spanning amino acids 230-312 in HCV polyprotein, was detected in 42.8% of patients during treatment with CIGB-230. The ability of T cells to proliferate in response to mitogenic stimulation was not weakened. Most individuals (78.6%) were seroprotected after anti-hepatitis B vaccination and 42.8% were hyper-responders (antibody titers > 100 UI/ml). No anti-mitochondrial, anti-nuclear and anti-extractable nuclear antigen antibodies were generated during immunization with CIGB-230. CONCLUSIONS: Vaccination with CIGB-230 in HCV chronically-infected individuals was safe, well tolerated and did not impair the ability to respond to non-HCV antigens.

Recombinant in vitro assembled hepatitis C virus core particles induce strong specific immunity enhanced by formulation with an oil-based adjuvant.

In the present work, immunogenicity of recombinant in vitro assembled hepatitis C virus core particles, HCcAg.120-VLPs, either alone or in combination with different adjuvants was evaluated in BALB/c mice. HCcAg.120-VLPs induced high titers of anti-HCcAg.120 antibodies and virus-specific cellular immune responses. Particularly, HCcAg.120-VLPs induced specific delayed type hypersensitivity, and generated a predominant T helper 1 cytokine pro file in immunized mice. In addition, HCcAg.120-VLPs prime splenocytes proliferate in vitro against different HCcAg.120-specific peptides, depending on either the immunization route or the adjuvant used. Remarkably, immunization with HCcAg.120-VLPs/Montanide ISA888 formulation resulted in a significant control of vaccinia virus titer in mice after challenge with a recombinant vaccinia virus expressing HCV core protein, vvCore. Animals immunized with this formulation had a marked increase in the number of IFN-gamma producing spleen cells, after stimulation with P815 cells infected with vvCore. These results suggest the use of recombinant HCV core particles as components of therapeutic or preventive vaccine candidates against HCV.

Giardia duodenalis: effects of an ozonized sunflower oil product (Oleozon) on in vitro trophozoites.

The ozonized sunflower oil product (Oleozon) was investigated to explore its cytotoxic activity on Giardia duodenalis in vitro cultivated trophozites. Oleozon produced inactivation of Giardia trophozoites in a dose- and cell density-dependent manner. Thirty microliter of Oleozon with peroxide index value of 500 equivalent-mmol of activated oxygen per kilogram were used to achieve a 100% inhibition (<-4.00 log unit) of trophozoites from an initial inoculum of 15x10(4) cells. This potent effect was confirmed by transmission electron microscopy where morphological deterioration of superficial structures mainly in the ventral disc, and formation of a great number of micro vesicles in the cytoplasm were found. We concluded that a direct chemical-oxidation attack by the active substances from Oleozon is one of the causes of the parasitocidal effect of this product. We suggest that the dose and cell density-dependent effect must be taken into account when prescription of this product for giardiasis treatment in humans.

A C-terminal truncated hepatitis C virus core protein variant assembles in vitro into virus-like particles in the absence of structured nucleic acids.

Little is known about the assembly pathway or structure of the hepatitis C virus (HCV). In this work a truncated HCcAg variant covering the first 120 aa (HCcAg.120) with a 32 aa N-terminal fusion peptide (6x Histag-Xpress epitope) was purified as a monomer under strong denaturing conditions. In addition, minor HCcAg.120 peaks exhibiting little different molecular mass by SDS-PAGE which possibly represents alternative forms harboring the N-termini of HCcAg.120 were detected. Analysis using gel filtration chromatography showed that HCcAg.120 assembled into high molecular weight structures in vitro in the absence of structured nucleic acids. The negative-stain electron microscopy analysis revealed that these structures correspond with spherical VLPs of uniform morphology and size distribution. The diameters of these particles ranged from 20 to 43nm with an average diameter of approximately 30 nm and were specifically immunolabelled with a mouse monoclonal antibody against the residues 5-35 of HCcAg. Results presented in this work showed that HCcAg.120 assembled in vitro into VLPs in the absence of structured nucleic acids with similar morphology and size distribution to those found in sera and hepatocytes from HCV-infected patients. Therefore, these VLPs would be important to elucidate the mechanisms behind the ability of HCcAg to assemble into a nucleocapsid structure.

[Usefulness of 2 coproparasitological methods and their utilization in an antigiardiasis therapeutic trial]. / Utilidad de 2 métodos coproparasitológicos y su empleo en un ensayo terapéutico antigiardiásico.

A study of 456 children aged 1-5 from 4 day care centers of San Miguel del Padron municipality was conducted in 1998, to evaluate the diagnosis of Giardia lamblia and other intestinal protozoa by using comparatively the coproparasitological diagnostic methods of direct examination and Ritchie's concentration technique or formol-ether. Besides, a therapeutical trial was developed with tinidazole and albendazole for treating the infection caused by G lamblia. Ritchie's concentration technique was more effective than the microscopic direct examination for diagnosing Giardia lamblia, Entamoeba histolytical/Entamoeba dispar and Cyclospora cayetanensis. It was demonstratyed that the serial examination was more sensitive than the analysis of just one sample (p< 0.01). On the other hand, tinidazole proved to be more efficient than albendazole to treat the infection produced by G. lamblia, with a greater cure percentage (72 % vs. 334.6%), (p < 0.01).

Recombinant in vitro assembled hepatitis C virus core particles induce strong specific immunity enhanced by formulation with an oil-based adjuvant

In the present work, immunogenicity of recombinant in vitro assembled hepatitis C virus core particles, HCcAg.120-VLPs, either alone or in combination with different adjuvants was evaluated in BALB/c mice. HCcAg.120-VLPs induced high titers of anti-HCcAg.120 antibodies and virus-specific cellular immune responses. Particularly, HCcAg.120-VLPs induced specific delayed type hypersensitivity, and generated a predominant T helper 1 cytokine pro file in immunized mice. In addition, HCcAg.120-VLPs prime splenocytes proliferate in vitro against different HCcAg.120-specific peptides, depending on either the immunization route or the adjuvant used. Remarkably, immunization with HCcAg.120-VLPs/Montanide ISA888 formulation resulted in a significant control of vaccinia virus titer in mice after challenge with a recombinant vaccinia virus expressing HCV core protein, vvCore. Animals immunized with this formulation had a marked increase in the number of IFN-γ producing spleen cells, after stimulation with P815 cells infected with vvCore. These results suggest the use of recombinant HCV core particles as components of therapeutic or preventive vaccine candidates against HCV.

Aislamiento y Axenizacion de Giardia Lamblia en ninos procedentes de Circulos Infantiles de Ciudad de La Habana / Isolation and purification of Giardia lamblia in children from day care centers in Havana City

Se reporta el aislamiento de Giardia lamblia procedentes de ninos de 3 circulos infantiles de Ciudad de La Habana. El 2,8 por ciento de los aislamientos fue logrado utilizando la desenquistacion in vitro, mientras que utilizando al gerbil como modelo animal para la desenquistacion del parasito se logro el 97,2 por ciento, lo que demostro la superioridad de este ultimo metodo

Ensayo inmunoenzimático en fase sólida para la detección de antígenos de Giardia lamblia / Immunoenzymatic assay in solid phase for the detection of Giardia lamblia antigens

Se normalizó un sandwich ELISA para la detección de antígenos de Giardia lamblia en heces humanas. Se estudiaron 175 muestras: 77 positivas, 61 negativas a quistes y/o trofozoítos por el examen directo de las heces y 19 muestras positivas a otros parásitos diferentes de G. lamblia. La sensibilidad de la técnica fue de 94,8 por ciento y la especificidad de 98,3 por ciento, el método detecta una concentración de antígenos de 31 ng. El procedimiento es simple, sensible y específico, por lo que pudiera ser útil para el diagnóstico y en estudios epidemiológicos

Parasitosis intestinales en 4 círculos infantiles de San Miguel del Padrón, Ciudad de La Habana, 1998 / Intestinal parasitic infections in child day-care centers located in San Miguel del Padrón municipality, Havana City, 1998

Se realizó un estudio coproparasitológico a 456 niños, de 1 a 5 años de edad pertenecientes a 4 círculos infantiles del municipio San Miguel del Padrón, durante el mes de noviembre de 1998, con la finalidad de conocer el comportamiento de las principales especies parásitas intestinales y en especial cómo se encontraba Giardia lamblia afectando a esta población infantil. Fueron recolectadas por cada niño, 3 muestras fecales frescas en días alternos, las que se procesaron con los métodos coproparasitológicos de examen directo y técnica de concentración de Ritchie. El parásito identificado con mayor frecuencia en este estudio fue G. lamblia, se encontraron 249 casos positivos, para una prevalencia de 54,6 porciento. El segundo lugar lo ocupó Blastocystis hominis (29,6 porciento), seguido por Endolimax nana (23,9 porciento). Las coccidias, Cryptosporidium parvum y Cyclospora cayetanensis, fueron detectadas con una baja frecuencia, 0,6 y 1,5 porciento respectivamente; la mayoría de los casos en un solo círculo. Estos resultados confirman que G. lamblia es el parásito más prevalente en los círculos infantiles, con un pico entre las edades de 2 a 4 años y sin diferencia entre los dos sexos

Utilidad de 2 métodos coproparasitológicos y su empleo en un ensayo terapéutico antigiardiásico

Se realizó un estudio a 456 niños con edades de 1 a 5 años, pertenecientes a 4 guarderías infantiles del municipio San Miguel del Padrón, en el mes de noviembre de 1998; para evaluar el diagnóstico de Giardia lamblia y otros protozoos intestinales, utilizando comparativamente los métodos de diagnóstico coproparasitológicos de examen directo y la técnica de concentración de Ritchie o formol-éter. Además, se desarrolló un ensayo terapéutico, utilizando tinidazol y albendazol, para el tratamiento de la infección por G. lamblia. La técnica de concentración de Ritchie fue más efectiva que el examen microscópico directo para el diagnóstico de Giardia lamblia, Entamoeba histolytica/ Entamoeba dispar y Cyclospora cayetanensis y se demostró la mayor sensibilidad del examen seriado sobre el análisis de una sola muestra (p< 0,01). Por otra parte, el tinidazol demostró mayor eficacia que el albendazol para el tratamiento de la infección por G. lamblia, con un mayor porcentaje de curación (72 % vs. 34,6 %) (p< 0,01).

A study of 456 children aged 1-5 from 4 day care centers of San Miguel del Padrón municipality was conducted in November, 1998, to evaluate the diagnosis of Giardia lamblia and other intestinal protozoa by using comparatively the coproparasitological diagnostic methods of direct examination and Ritchie's concentration technique or formol-ether. Besides, a therapeutical trial was developed with tinidazole and albendazole for treating the infection caused by G. lamblia. Ritchie's concentration technique was more effective than the microscopic direct examination for diagnosing Giardia lamblia, Entamoeba histolytical/Entamoeba dispar and Cyclospora cayetanensis. It was demonstratyed that the serial examination was more sensitive than the analysis of just one sample (p< 0.01). On the other hand, tinidazole proved to be more efficient than albendazole to treat the infection produced by G. lamblia, with a greater cure percentage (72 % vs. 334.6 %), (p < 0.01).

Sensibilidad de Giardia lamblia a extractos de Artemisia absinthium y Artemisia vulgaris in vitro / Sensitivity of Giardia lamblia to Artemisia absinthium and Artemisia vulgaris extracts in vitro

Se evaluó el efecto de 2 extractos alcohólicos de Artemisia (Artemisia absinthium y Artermisia vulgaris) y una fracción de lactonas sesquiterpénicas de A. absinthium sobre el crecimiento in vitro de Giardia lamblia. Los resultados obtenidos muestran tantos por cientos de inhibición del crecimiento superiores al 70 por ciento con el extracto de A. absinthium, aun con la mayor dilución testada, no así con el extracto de A. vulgaris con el cual sólo con la fracción más concentrada se obtuvo el 30 por ciento de inhibición. La actividad del extracto de lactonas de A. absinthium fue de 86,6 por ciento y 57,7 por ciento de inhibición del crecimiento para las concentraciones de 166 y 83 ug/mL, respectivamente