ABSTRACT
AIMS: Currently, immunoinformatic approaches have shown promise in rapidly and cost-effectively identifying new antigens from the Leptospira proteome. Chimeric multiepitope proteins offer a strategy with significant potential for implementation in serology and the design of effective vaccines. METHODS AND RESULTS: In this study, we detail the immunoinformatic analyses and design of a new recombinant chimeric protein constructed with epitopes identified from the sequences of ErpY-like and LemA proteins, previously identified as potential antigens for controlling leptospirosis. We expressed the chimeric protein using Escherichia coli heterologous systems, evaluated its antigenicity using serum from naturally infected patients, and its immunogenicity in mice as an animal model, with Freund as an adjuvant. The resulting recombinant chimeric protein, named rErpY-LemA, was successfully expressed and purified using a prokaryotic system, with an expected mass of 35 kDa. Serologic assays using serum samples from naturally infected patients demonstrated recognition of the chimera protein by antibodies present in sera. Animals immunized with the chimera exhibited a significant IgG antibody response from the 7th day (P<0.001), persisting until day 49 of experimentation, with a titer of 1:12,800 (P<0.05). Notably, significant production of IgA, IgM, and IgG subclasses was observed in animals immunized with the chimera. CONCLUSIONS: These results highlight the promising role of immunoinformatics in rapidly identifying antigens and the potential of chimeric multiepitope proteins in developing effective strategies for leptospirosis control.
ABSTRACT
El objetivo de esta investigación fue aislar y caracterizar bacterias solubilizadoras de fosfatos (BSF) asociadas a la rizosfera de Baccharis macrantha y Viburnum triphyllum, y evaluar su capacidad para solubilizar fosfatos en condiciones in vitro. Además se determinó el efecto de la inoculación de las cepas de BSF más eficientes sobre el crecimiento de B. macrantha. Las muestras de suelo rizosférico de B. macrantha y V. triphyllum fueron colectadas en los meses de mayo-período de lluvia y septiembre-período seco del 2012. Para la cuantificación de bacterias heterótrofas cultivables y BSF se empleó el método de recuento en placa en los medios Agar Tripticasa de Soya y Pikovskaya (PVK) respectivamente. La capacidad de solubilización de fosfatos de las cepas aisladas se estimó a partir del diámetro de los halos formados alrededor de las colonias en el medio de cultivo PVK después de 7 días de incubación a 28 °C. Los ensayos de inoculación en B. macrantha se realizaron con las BSF más eficientes. La inoculación de las BSF B. firmusy P. fluorescens de forma individual y como inoculante combinado mostro un efecto benéfico, incrementando significativamente el porcentaje de germinación de semillas, la altura de la plántula, la longitud de la raíz y el peso seco de B. macrantha. La inoculación de BSF podría ser considerada una estrategia para mejorar el crecimiento y establecimiento de B. macrantha en pastizales abandonados.
The objectives of this research was to isolate and characterize phosphate solubilizing bacteria (BSF) associated to the rhizosphere of Baccharis macrantha and Viburnum triphyllum, and to assess their ability to solubilize phosphate under conditions in vitro. Furthermore to determine the effect of inoculation of the strains BSF more efficient on the growth of B. macrantha. Rhizosphere soil samples of B. macrantha and V. triphyllum were collected in the months of May-rainy season and September-period dry the 2012. Trypticase Soya Agar and Pikovskaya (PVK) were used for quantification of culturable heterotrophic bacteria and BSF, respectively. The phosphate solubilizing capacity of the isolated strains was estimated from the diameter of the halo around the colonies formed in the culture medium PVK after 7 days incubation at 28 °C. Inoculation assays were performed with more efficient BSF in B. macrantha. Inoculation of BSF Bacillus firmus and Pseudomona fluorescens individually and as inoculant combined showed a beneficial effect, significantly increasing the percentage of seed germination, seedling height, root length and dry weight of B . macrantha. Inoculation the BSF could be considered a strategy to improve the growth and development of B. macrantha in abandoned pastures.
