RESUMEN
OBJECTIVE: To investigate the efficacy of a vaccine formulation of the 19 kDa conserved carboxyl-terminal fragment of Plasmodium yoelii merozoite surface protein-1 (PyMSP1(19)) formulated with CpG ODN 1826 and Montanide ISA51 or ISA720 when used to immunize mice by a single injection. METHODS: Groups of BALB/c mice were immunized parenterally with one, two or four injections with PBS or PyMSP1(19) formulated with CpG ODN in ISA51 or ISA720. Sera were collected weekly and assessed for total IgG and IgG subclass titers. Protection was tested by challenge infection with P. yoelii YM. RESULTS: Interestingly, single injection immunization showed the same kinetics of antibody responses as two- or four-injection immunization. However, the peak antibody response induced by PyMSP1(19) in CpG ODN and ISA51 appeared earlier than that induced by PyMSP1(19) in CpG ODN and ISA720 (28 days vs 41 days). At day 63 after the first injection, the PyMSP1(19)-specific IgG antibody levels by single injection and four-injection immunizations were not different. However, the levels of the IgG2a antibody subclass were significantly lower by single injection immunization with PyMSP1(19) in CpG ODN and ISA720. The antibodies were sustained at high levels for at least 20 weeks. After challenge infection, all mice immunized by a single injection of PyMSP1(19) in CpG ODN and ISA51 survived with low-grade parasitemia, while 50% of mice immunized with PyMSP1(19) in CpG ODN and ISA720 died with high levels of parasitemia. CONCLUSION: These findings suggest that MSP1(19) immunization by a single injection can induce protective immunity, particularly when formulated with an appropriate strong adjuvant.
Asunto(s)
Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Vacunas contra la Malaria/inmunología , Proteína 1 de Superficie de Merozoito/inmunología , Oligodesoxirribonucleótidos/inmunología , Plasmodium yoelii/inmunología , Adyuvantes Inmunológicos/metabolismo , Animales , Femenino , Vacunas contra la Malaria/administración & dosificación , Vacunas contra la Malaria/química , Manitol/administración & dosificación , Manitol/análogos & derivados , Manitol/química , Manitol/inmunología , Proteína 1 de Superficie de Merozoito/administración & dosificación , Proteína 1 de Superficie de Merozoito/química , Ratones , Ratones Endogámicos BALB C , Ácidos Oléicos/administración & dosificación , Ácidos Oléicos/química , Ácidos Oléicos/inmunología , Oligodesoxirribonucleótidos/administración & dosificación , Oligodesoxirribonucleótidos/químicaRESUMEN
Due to the indistinguishable morphology between Entamoeba histolytica (pathogenic) and Entamoeba dispar (non pathogenic), PCR-based assays were conducted. Based on microscopy, suspected Entamoeba cells were detected in 30 out of 455 fecal samples obtained from individuals residing at Thai/Myanmar border region. The target genes for PCR amplification included genes encoding small subunit rRNA (SSU-rRNA), chitinase and serine rich Entamoeba protein. PCR primers derived from SSU-rRNA gene amplified both E. histolytica and E. dispar genes producing an amplicon of 1,080 bp, and detected 3 out of 30 samples. PCR primers derived from chitinase gene of E. histolytica generating amplicons of 500 and 1,260 bp, samples were positive in 12 out of 30 samples. Due the large difference of gene encoding serine rich protein between E. histolytica and E. dispar, two specific sets of primers were designed. SREH-primer set, specific for E. histolytica, generated amplicons of 550 and 700 bp and detected 22 out of 30 samples. SED-primer set, specific to E. dispar, produced an amplicon of 550 bp, and together with a nested primer pair generating an amplicon of 477 bp, detected 16 out of 30 samples. Thus, detection of single and mixed infections of the two Entamoeba species could be effectively achieved directly from DNA extracted from feces without the need to culture the parasites.
Asunto(s)
Disentería Amebiana/diagnóstico , Entamoeba histolytica/aislamiento & purificación , Heces/parasitología , Reacción en Cadena de la Polimerasa/métodos , Animales , Disentería Amebiana/epidemiología , Entamoeba histolytica/genética , Humanos , Mianmar/epidemiología , Prevalencia , ARN Ribosómico/genética , Tailandia/epidemiologíaRESUMEN
Merozoite surface protein 1 (MSP1) is the major protein on the surface of the plasmodial merozoite, and its carboxy terminus, the 19-kDa fragment (MSP1(19)), is highly conserved and effective in induction of a protective immune response against malaria parasite infection in mice and monkeys. However, the duration of the immune response has not been elucidated. As such, we immunized BALB/c mice with a standard four-dose injection of recombinant Plasmodium yoelii MSP1(19) formulated with Montanide ISA51 and CpG oligodeoxynucleotide (ODN) and monitored the MSP1(19)-specific antibody levels for up to 12 months. The antibody titers persisted constantly over the period of time without significant waning, in contrast to the antibody levels induced by immunization with Freund's adjuvant, where the antibody levels gradually declined to significantly lower levels 12 months after immunization. Investigation of immunoglobulin G (IgG) subclass longevity revealed that only the IgG1 antibody level (Th2 type-driven response) decreased significantly by 6 months, while the IgG2a antibody level (Th1 type-driven response) did not change over the 12 months after immunization, but the boosting effect was seen in the IgG1 antibody responses but not in the IgG2a antibody responses. After challenge infection, all immunized mice survived with negligibly patent parasitemia. These findings suggest that protective immune responses to MSP1(19) following immunization using oil-based Montanide ISA51 and CpG ODN as an adjuvant are very long-lasting and encourage clinical trials for malaria vaccine development.