Enzymatic activities and protein profile of latex from Calotropis procera.

The laticifer fluid of Calotropis procera is rich in proteins and there is evidence that they are involved in the pharmacological properties of the latex. However, not much is known about how the latex-containing proteins are produced or their functions. In this study, laticifer proteins of C. procera were pooled and examined by 1D and 2D electrophoresis, masses spectrometry (MALDI-TOF) and characterized in respect of proteolytic activity and oxidative enzymes. Soluble laticifer proteins were predominantly composed of basic proteins (PI>6.0) with molecular masses varying between 5 and 95 kDa. Proteins with a molecular mass of approximately 26,000 Da were more evident. Strong anti-oxidative activity of superoxide dismutase (EC 1.15.1.1) (1007.74+/-91.89 Ug(-1)DM) and, to a lesser extent ascorbate peroxidase (EC 1.11.1.1) (0.117(d)+/-0.013 microMol H(2)O(2)g(-1)min(-1)), were detected. However, catalase (EC 1.11.1.6) was absent. The strong proteolytic activities of laticifer proteins from C. procera were shown to be shared by at least four distinct cysteine proteinases (EC 3.4.22.16) that were isolated by gel filtration chromatography. Serine and metaloproteinases were not detected and aspartic proteinase activities were barely visible. Chitinases (EC 3.2.1.14) were also isolated in a chitin column and their activities quantified. The presence of these enzymatic activities in latex from C. procera may confirm their involvement in resistance to phytopathogens and insects, mainly in its leaves where the latex circulates abundantly.     

Caatinga Ethnobotany: Anthropogenic Landscape Modification and Useful Species in Brazil’s Semi-Arid Northeast

Caatinga Ethnobotany: Anthropogenic Landscape Modification and Useful Species in Brazil’s Semi-Arid Northeast This study explores the contribution of anthropogenic landscapes in providing useful botanical resources to a Caatinga community in Pernambuco, Brazil. Ethnobotanical data were collected through semi-structured interviews using the checklist-interview method and by means of a “field herbarium” of the most abundant species in the anthropogenic zones. We recorded 119 species distributed in 36 families, of which 79 were found to be useful. Forage was the most prominent use category, containing 84% of the citations, followed by medicinals (36.70%), foods (10.12%), and wood (8.86%). Herbaceous species predominated (63.29%), followed by shrubs (3.79%), sub-shrubs (21.51%), trees (8.86%), and creepers (2.53%). Trees exhibited a greater number of uses than other life-forms (p < 0.05). Significant differences in richness were found, with the highest richness of species (χ2 = 60.28, p < 0.05), genera (χ2 = 49.03, p < 0.05), and families (χ2 = 20.16, p < 0.05) appearing in the rainy season. We concluded that fodder use was the most important category in our anthropogenic research areas, accounting for a higher number of species, genera, and families. The next most important categories were medicinal, timber, and food plants, respectively.     

Antifungal effects of the flavonoids kaempferol and quercetin: a possible alternative for the control of fungal biofilms

This study aimed to determine the minimum inhibitory concentration (MIC) of kaempferol and quercetin against planktonic and biofilm forms of the Candida parapsilosis complex. Initially, nine C. parapsilosis sensu stricto, nine C. orthopsilosis and nine C. metapsilosis strains were used. Planktonic susceptibility to kaempferol and quercetin was assessed. Growing and mature biofilms were then exposed to the flavonoids at MIC or 10xMIC, respectively, and theywere also analyzed by confocal laser scanning microscopy. The MIC ranges were 32-128 µg ml^?1 for kaempferol and 0.5-16 µg ml^?1 for quercetin. Kaempferol and quercetin decreased (P?<?0.05) the metabolic activity and biomass of growing biofilms of the C. parapsilosis complex. As for mature biofilms, the metabolic effects of the flavonoids varied, according to the cryptic species, but kaempferol caused an overall reduction in biofilm biomass. Microscopic analyses showed restructuring of biofilms after flavonoid exposure. These results highlight the potential use of these compounds as sustainable resources for the control of fungal biofilms.     

MyD88-dependent activation of dendritic cells and CD4(+) T lymphocytes mediates symptoms, but is not required for the immunological control of parasites during rodent malaria

We investigated the role of different TLRs and MyD88 in host resistance to infection and malaria pathogenesis. TLR2(-/-), TLR4(-/-), TLR6(-/-), TLR9(-/-) or CD14(-/-) mice showed no change in phenotypes (parasitemia, body weight and temperature) when infected with Plasmodium chabaudi chabaudi (AS). MyD88(-/-) mice displayed comparable ability to wild type animals in controlling and clearing parasitemia. Importantly, MyD88(-/-) mice exhibited impaired production of TNF-alpha and IFN-gamma as well as attenuated symptoms, as indicated by changes in body weight and temperature during parasitemia. Consistently, CD11b(+) monocytes and CD11c(+) dendritic cells from infected MyD88(-/-) mice were shown impaired for production of pro-inflammatory cytokines, and in initiating CD4(+) T cell responses. Importantly, the inhibition of T cell activation with anti-CD134L, mostly inhibited IFN-gamma, partially inhibited TNF-alpha production, and protected the animals from malaria symptoms. Our findings suggest that MyD88 and possibly its associated TLRs expressed by dendritic cells play an important role in pro-inflammatory responses, T cell activation, and pathogenesis of malaria, but are not critical for the immunological control of the erythrocytic stage of P. chabaudi.     

Driftnet fishery threats sea turtles in the Atlantic Ocean

Fisheries are recognised as a major threat to sea turtles worldwide. Oceanic driftnets are considered the main cause of the steep decline in Pacific Ocean populations of the leatherback sea turtle Dermochelys coriacea. The world’s largest leatherback population nests in West Africa and migrates across the Atlantic Ocean to feed off the South American coast. There, the turtles encounter a range of fisheries, including the Brazilian driftnet fishery targeting hammerhead sharks. From 2002 to 2008, 351 sea turtles were incidentally caught in 41 fishing trips and 371 sets. Leatherbacks accounted for 77.3% of the take (n = 252 turtles, capture rate = 0.1405 turtles/km of net), followed by loggerheads Caretta caretta (47 individuals, capture rate = 0.0262 turtles/km of net), green turtles Chelonia mydas (27 individuals, capture rate = 0.0151 turtles/km of net) and unidentified hard-shelled turtles (25 individual, capture rate = 0.0139 turtles/km of net) that fell off the net during hauling. Immediate mortality (i.e., turtles that were dead upon reaching the vessel, excluding post-release mortality) was similar among the species and accounted for 22.2 to 29.4% of turtles hauled onboard. The annual catch by this fishery ranged from 1,212 to 6,160 leatherback turtles, as estimated based on bootstrap procedures under different fishing effort scenarios in the 1990s. The present inertia in law and enforcement regarding gillnet regulations in Brazil could result in the reestablishment of the driftnet fishery, driving rates of leatherback mortality to levels similar to those observed in previous decades. This development could potentially lead to the collapse of the South Atlantic leatherback population, mirroring the decline of the species in the Pacific. In light of these potential impacts and similar threats to other pelagic mega fauna, we recommend banning this type of fishery in the region.     

The major chromoblastomycosis fungal pathogen, Fonsecaea pedrosoi, extracellularly releases proteolytic enzymes whose expression is modulated by culture medium composition: implications on the fungal development and cleavage of key's host structures

We investigated the possible secretion of peptidases by F. pedrosoi, when conidial cells were cultured in two distinct media. Aspartyl proteolytic activity was detected on the Czapeck-Dox-derived supernatant, which was blocked by pepstatin, and only active in extremely acidic conditions. The supernatant obtained after conidia growth in Kauffman medium presented metallopeptidase activity, which was active over a broad pH range and sensitive to 1,10-phenanthroline and EGTA. Additionally, both culture supernatants were able to cleave a wide range of proteinaceous substrates, including important human serum proteins (e.g. albumin and immunoglobulin G) and extracellular matrix components (e.g. fibronectin and laminin). As peptidases participate in different cellular metabolic pathways, we also tested the influence of proteolytic inhibitors on the F. pedrosoi conidia development in vitro. The metallopeptidase inhibitors, 1,10-phenanthroline, EGTA and EDTA, strongly abrogated the growth of conidial forms by approximately 95%, 85% and 60%, respectively. Moreover, 1,10-phenanthroline blocked the differentiation process from conidia to mycelia, an essential step during the F. pedrosoi life cycle. Phenylmethanesulfonyl fluoride, a serine peptidase inhibitor, slightly reduced the conidial growth, whereas proteolytic inhibitors of cysteine (E-64) and aspartic (pepstatin) type peptidases did not alter conidial developmental behavior. In summary, our results showed for the first time the expression of extracellular proteolytic activity by F. pedrosoi conidial cells.     

Human colostrum contains IgA antibodies reactive to colonization factors I and II of enterotoxigenic Escherichia coli

Diarrhea is an important cause of morbidity and mortality amongst infants of low socio-economic levels in developing countries and in travelers who visit such areas. Enterotoxigenic E. coli strains express two sets of virulence-associated factors: enterotoxins (heat-stable toxins or heat-labile toxins) and colonization factors. Studies have shown that breast-feeding protects infants against infectious diseases, such as diarrhea, as it presents a great variety of immunological components. The aim of this study was to analyze the reactivity of immunoglobulin A from human colostrum to colonization factor antigens I and II. The colostrum ability in preventing enterotoxigenic E. coli adhesion to Caco-2 cells was also evaluated. Colostrum samples were collected from 32 healthy women, and a human colostrum pool was prepared. Enterotoxigenic E. coli strains expressing colonization factor antigens I and II were utilized. The colostrum pool and individual samples showed variable antienterotoxigenic E. coli immunoglobulin A titers, that were reactive with colonization factor antigen I and CS1/CS3 (colonization factor antigen II). The human colostrum pool and individual samples inhibited enterotoxigenic E. coli colonization factor antigen I and II adhesion to Caco-2 cells, at variable levels, and this ability was a result of immunoglobulin A antibodies reactive to these colonization factors. The immunoglobulin A-depleted pool lost this inhibitory ability. As bacterial adhesion is the initial mechanism of enterotoxigenic E. coli infection, breast-feeding could protect the offspring against diarrhea caused by this agent.