BackgroundLeishmaniasis is caused by intracellular Leishmania parasites that induce a T-cell mediated response associated with recognition of CD4+ and CD8+ T cell Line 1Lineepitopes. Identification of CD8+ antigenic determinants is crucial for vaccine and therapy development. Herein, we developed an open-source software dedicated to search and compile data obtained from currently available on line prediction algorithms.Methodology/Principal FindingsWe developed a two-phase algorithm and implemented in an open source software called EPIBOT, that consolidates the results obtained with single prediction algorithms, generating a final output in which epitopes are ranked. EPIBOT was initially trained using a set of 831 known epitopes from 397 proteins from IEDB. We then screened 63 Leishmania braziliensis vaccine candidates with the EPIBOT trained tool to search for CD8+ T cell epitopes. A proof-of-concept experiment was conducted with the top eight CD8+ epitopes, elected by EPIBOT. To do this, the elected peptides were synthesized and validated for their in vivo cytotoxicity. Among the tested epitopes, three were able to induce lysis of pulsed-target cells.ConclusionOur results show that EPIBOT can successfully search across existing prediction tools, generating a compiled list of candidate CD8+ epitopes. This software is fast and a simple search engine that can be customized to search over different MHC alleles or HLA haplotypes.

B1 cells, a subset of B lymphocytes whose developmental origin, phenotype, and function differ from that of conventional B2 cells, are the main source of “natural” IgM but can also respond to infection by rapidly producing pathogen-specific IgM directed against T-independent antigens. Francisella tularensis (Ft) is a Gram-negative bacterium that causes tularemia. Infection with Ft Live Vaccine Strain activates B1 cells for production of IgM directed against the bacterial LPS in a process incompletely understood. Here we show that immunization with purified Ft LPS elicits production of LPS-specific IgM and IgG3 by B1 cells independently of TLR2 or MyD88. Immunization, but not infection, generated peritoneum-resident memory B1 cells that differentiated into LPS-specific antibody secreting cells (ASC) upon secondary challenge. IL-5 was rapidly induced by immunization with Ft LPS and was required for production of LPS-specific IgM. Antibody-mediated depletion of ILC2 indicated that these cells were the source of IL-5 and were required for IgM production. IL-25, an alarmin that strongly activates ILC2, was rapidly secreted in response to immunization or infection and its administration to mice significantly increased IgM production and B1 cell differentiation to ASC. Conversely, mice lacking IL-17RB, the IL-25 receptor, showed impaired IL-5 induction, IgM production, and B1 ASC differentiation in response to immunization. Administration of IL-5 to Il17rb-/- mice rescued these B1 cells-mediated responses. Il17rb-/- mice were more susceptible to infection with Ft LVS and failed to develop immunity upon secondary challenge suggesting that LPS-specific IgM is one of the protective adaptive immune mechanisms against tularemia. Our results indicated that immunization with Ft LPS triggers production of IL-25 that, through stimulation of IL-5 release by ILC2, promotes B1 cells activation and differentiation into IgM secreting cells. By revealing the existence of an IL-25-ILC2-IL-5 axis our results suggest novel strategies to improve vaccination against T-independent bacterial antigens.

This study aimed to verify the potential of three Aspergillus and Bacillus species as growth promoters in cotton plants under greenhouse conditions. The experiment was conducted with a completely randomized design with seven treatments (six microorganisms plus one control) and five replicates until the flowering stage at 70 days after emergence. The inoculation of cotton plants with Bacillus velezensis (Bv188) and Bacillus subtilis (Bs248 and Bs290) had a positive effect on total nitrogen extraction (899.31, 962.18, and 755.41 mg N/kg dry matter, respectively) compared to the control (459.31 mg N/kg dry weight), total phosphorus extraction (121.94, 124.31, and 99.27 mg P/kg dry matter, respectively) compared to the control (65.10 mg P/kg dry matter), and total dry matter (41.08, 43.59, and 49.86 g/plant, respectively) compared to the control (26.70 g/plant), as well as biomass carbon (72.26, 35.18, and 14.7 mg/kg soil, respectively). Cotton plants inoculated with Aspergillus brasiliensis (F111), Aspergillus sydowii (F112), and Aspergillus sp. (versicolor section) (F113) had higher total nitrogen extraction (953.33, 812.59, and 891.62 mg N/kg dry matter, respectively) compared to the control (459.31 mg N/kg dry matter), a higher total phosphorus (122.30, 104.86, and 118.45 mg P/kg dry matter, respectively) compared to the control (65.10 mg P/kg dry matter), a higher total dry matter (37.52, 37.41, and 53.02 g/plant) compared to the control (26.70 g/plant), and greater respiratory activity (14.98, 10.43, and 7.11 mg CO2/100 g soil, respectively) compared to the control (3.5 mg CO2/100 g soil). The fungi A. brasiliensis (F111) and A. sydowii (F112) promoted higher phosphorus absorption by cotton plants, which was reflected by the lower amount of nutrients in the soil (7.10 and 16.96 g P/dm3 soil) than in the control (26.91 g P/dm3 soil). The results suggest that B. subtilis 248 promoted an increase in phosphorus extracted from the roots and total and phosphorous compounds from the root dry matter and increased the value of soil respiratory activity, and this bacterium could be used as an inoculant in cotton crops.

Objective. To develop, validate, and patent a Restraining Device for Small Animal Imaging Exams (RDSAIE) that allows exams to be comfortably conducted without risks to animals and professionals. Methods. A RDSAIE with a mobile cover and shelf was built with transparent acrylic material. A total of six anesthetized rabbits were used to perform the following imaging exams of the skull: Cone Beam Computed Tomography, Magnetic Resonance Imaging, and Scintigraphy. Results. The device showed great functionality and full visibility of the animal behavior, which remained fully stabilized and immobilized in either the horizontal or vertical position without the need for a person to remain in the test room to assist them. The procedures were performed without difficulty, and images of good resolution and without artifacts were obtained. Conclusion. The RDSAIE is comfortable, safe, efficient, and ergonomic. It allows the easy placement of animals in different body positions, including the vertical, the maintenance of postural stability, and full visibility. It may be constructed for animals heavier than 4 kg and it is adaptable for translational studies in anima nobile.

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