Aplicação da alta pressão na conservação de bombons

O presente trabalho teve por objectivo a avaliação do tempo de vida útil de bombons submetidos ao tratamento de alta pressão, utilizando como termo de comparação um ensaio controlo, onde os bombons foram apenas mantidos a 20°C e um ensaio em que as amostras foram conservadas a 4°C. Para tal foi necessário a monitorização das propriedades físico-químicas, microbiológicas e estruturais dos bombons e a optimização das variáveis envolvidas no processo de produção e conservação: i) seleção do tipo de tratamento (alta pressão, refrigeração ou testemunho); ii) seleção dos ciclos de tratamento (400MPa/2,5minutos ou 500MPa/1minuto); iii) temperatura de conservação (20 °C, ou 4 °C); iv) tempo de conservação (dos 0 aos 180dias). Relativamente à revisão bibliográfica, foi feita pesquisa acerca dos desenvolvimentos históricos da tecnologia de alta pressão, princípios gerais e descrição do processo, efeito do tratamento, assim como de factores que o possam influenciar. As metodologias seguidas incluíram a produção de bombons, com recheio constituído por natas e chocolate branco e cobertura de chocolate negro, seguida da avaliação físico-química (humidade, aw, pH, cor), microbiológica (mesófilos aeróbios totais, bolores e leveduras) e de reologia. Os melhores resultados observara.se no ensaio mantido a 4°C. Comparativamente, no tratamento de alta pressão, o ensaio a 400MPa foi o mais negativo relativamente à humidade e o ensaio a 500MPa o menos eficaz ao nível do aw, pH, cor e modulo de armazenamento (G’). Contudo, nas análises microbiológicas, este ultimo foi mais eficiente que o anterior (400MPa) e que o testemunho (0,1MPa/20°C)

A Genome-Wide Phenotypic Analysis of <i>Saccharomyces cerevisiae</i>’s Adaptive Response and Tolerance to Chitosan in Conditions Relevant for Winemaking

In the wine industry, the use of chitosan, a non-toxic biodegradable polysaccharide with antimicrobial properties, has been gaining interest with respect to envisaging the reduction in the use of sulfur dioxide (SO2). Although the mechanisms of toxicity of chitosan against fungal cells have been addressed before, most of the studies undertaken used other sources of chitosan and/or used conditions to solubilize the polymer that were not compatible with winemaking. Herein, the effect of a commercial formulation of chitosan approved for use in winemaking over the growth of the spoilage yeast species Dekkera anomala, Saccharomycodes ludwigii, Zygosaccharomyces bailii, and Pichia anomala was assessed. At the legally allowed concentration of 0.1 g/L, chitosan inhibited the growth of all spoilage yeasts, except for the tested Pichia anomala strains. Interestingly, the highly SO2-tolerant yeasts S. ludwigii and Z. bailii were highly susceptible to chitosan. The growth of commercial Saccharomyces cerevisiae was also impacted by chitosan, in a strain-dependent manner, albeit at higher concentrations. To dissect this differential inhibitory potential and gain further insight into the interaction of chitosan over fungal cells, we explored a chemogenomic analysis to identify all of the S. cerevisiae genes conferring protection against or increasing susceptibility to the commercial formulation of chitosan. Among the genes found to confer protection against chitosan, a high proportion was found to encode proteins required for the assembly and structuring of the cell wall, enzymes involved in the synthesis of plasma membrane lipids, and components of signaling pathways that respond to damages in the plasma membrane (e.g., the Rim101 pathway). The data obtained also suggest that the fungal ribosome and the vacuolar V-ATPase could be directly targeted by chitosan, since the deletion of genes encoding proteins required for the structure and function of these organelles was found to increase tolerance to chitosan. We also demonstrated, for the first time, that the deletion of ITR1, AGP2 and FPS1, encoding plasma membrane transporters, prominently increased the tolerance of S. cerevisiae to chitosan, suggesting that they can serve as carriers for chitosan. Besides providing new insights into the mode of action of chitosan against wine yeasts, this study adds relevant information for its rational use as a substitute/complementary preservative to SO2

Additional file 1: Figure S1. of Selection strategy of phage-displayed immunogens based on an in vitro evaluation of the Th1 response of PBMCs and their potential use as a vaccine against Leishmania infantum infection

Parasitological and immunological evaluations obtained in the infected and/or vaccinated animals in the second vaccine experiment. BALB/c mice (n = 16, per group) were vaccinated subcutaneously in their left hind footpad with Wild-type (WT), Random, B1 or D11 phage clones. Additional mice received only saline (n = 16). Three doses were administered at 14-day intervals. 30 days after the last vaccine, animals (n = 8, per group) were infected in the right hind footpad with 1 × 107 stationary promastigotes of L. infantum. Before and 60 days after challenge, the anti-phage and anti-parasite IgG2a and IgG1 isotype antibody levels were obtained, and the ratios between IgG2a and IgG1 results were calculated and are shown before (a) and after (b) infection. In addition, spleen cells were collected to evaluate the cytokine response, when they were incubated in complete RPMI 1640 medium (negative control) or in vitro stimulated with SLA (25 μg/ml) or with the respective clone (1 × 1010 phages), for 48 h at 37 °C in 5% CO2. IFN-γ, IL-12, GM-CSF, IL-4 and IL-10 levels were then measured by ELISA in the culture supernatants before (c) or after (d) infection. 6 days after challenge, the parasite burden was determined in the liver, spleen, draining lymph nodes and bone marrow of the animals, by a limiting dilution assay (e). Using the cell supernatants employed to evaluate cytokines, the nitrite production was also evaluated in this time (f). +indicates a statistically significant difference in relation to the B1 and D11 phages groups (P < 0.0001). ***indicates a statistically significant difference in relation to the saline, WT and Random groups (P < 0.0001). (TIFF 139 kb

Selection strategy of phage-displayed immunogens based on an in vitro evaluation of the Th1 response of PBMCs and their potential use as a vaccine against Leishmania infantum infection

Abstract Background The development of a vaccine for the prevention of visceral leishmaniasis (VL) still represents a significant unmet medical need. A human vaccine can be found if one takes into consideration that many people living in endemic areas of disease are infected but do not develop active VL, including those subjects with subclinical or asymptomatic infection. Methods In this study, a phage display was used to select phage-exposed peptides that were specific to immunoglobulin G (IgG) antibodies from asymptomatic and symptomatic VL patients, separating them from non-infected subjects. Phage clones presenting valid peptide sequences were selected and used as stimuli of peripheral blood mononuclear cells (PBMCs) obtained from both patients’ groups and controls. Those with higher interferon-gamma (IFN-γ)/interleukin (IL)-10 ratios were further selected for vaccination tests. Results Among 17 evaluated clones, two were selected, B1 and D11, and used to immunize BALB/c mice in an attempt to further validate their in vivo protective efficacy against Leishmania infantum infection. Both clones induced partial protection against the parasite challenge, which was evidenced by the reduction of parasitism in the evaluated organs, a process mediated by a specific T helper (Th)1 immune response. Conclusions To the best of our knowledge, this study is the first to use a rational strategy based on in vitro stimulation of human PBMCs with selected phage-displayed clones to obtain new immunogens against VL