A cysteine protease from the latex of Ficus benjamina has in vitro anthelmintic activity against Haemonchus contortus

Abstract Haemonchus contortus is a gastrointestinal nematode that is responsible for high mortality rates in ruminant herds. The resistance of nematodes to synthetic anthelmintics is widespread and requires a continuous search for new bioactive molecules, such as proteins. The objective of this study was to evaluate the anthelmintic potential of a protease purified from the latex of Ficus benjamina against H. contortus . Fresh latex was collected from plants via small incisions in the green stems, the rubber was removed by centrifugation, and the latex protein extract (LPE) was obtained. After LPE fractionation with ammonium sulfate and chromatography of the fraction containing the highest proteolytic activity on CM-cellulose, a cysteine protease (FbP) was purified. FbP has a molecular mass of approximately 23.97 kDa, and its proteolytic activity was stable between pH 6.0 and pH 10 and over a broad temperature range, with optimum activity at 60 °C. FbP inhibited both the development and exsheathment of H. contortus larvae, with 50% effective concentrations of 0.26 and 0.79 mg/mL, respectively. We conclude that this cysteine protease from F. benjamina latex with anthelmintic activity against H. contortus could be a promising alternative for the development of products for use in parasite control programmes

Identification of the alpha-enolase P46 in the extracellular membrane vesicles of Bacteroides fragilis

<div><p> BACKGROUND Members of the Bacteroides fragilis group are the most important components of the normal human gut microbiome, but are also major opportunistic pathogens that are responsible for significant mortality, especially in the case of bacteraemia and other severe infections, such as intra-abdominal abscesses. Up to now, several virulence factors have been described that might explain the involvement of B. fragilis in these infections. The secretion of extracellular membrane vesicles (EMVs) has been proposed to play a role in pathogenesis and symbiosis in gram-negative bacteria, by releasing soluble proteins and other molecules. In B. fragilis, these vesicles are known to have haemagglutination and sialidosis activities, and also contain a capsular polysaccharide (PSA), although their involvement in virulence is still not clear. OBJECTIVE The aim of this study was to identify proteins in the EMV of the 638R B. fragilis strain by mass spectrometry, and also to assess for the presence of Bfp60, a surface plasminogen (Plg) activator, previously shown in B. fragilis to be responsible for the conversion of inactive Plg to active plasmin, which can also bind to laminin-1. METHODS B. fragilis was cultured in a minimum defined media and EMVs were obtained by differential centrifugation, ultracentrifugation, and filtration. The purified EMVs were observed by both transmission electron microscopy (TEM) and immunoelectron microscopy (IM). To identify EMV constituent proteins, EMVs were separated by 1D SDS-PAGE and proteomic analysis of proteins sized 35 kDa to approximately 65 kDa was performed using mass spectrometry (MALDI-TOF MS). FINDINGS TEM micrographs proved the presence of spherical vesicles and IM confirmed the presence of Bfp60 protein on their surface. Mass spectrometry identified 23 proteins with high confidence. One of the proteins from the B. fragilis EMVs was identified as an enolase P46 with a possible lyase activity. MAIN CONCLUSIONS Although the Bfp60 protein was not detected by proteomics, α-enolase P46 was found to be present in the EMVs of B. fragilis. The P46 protein has been previously described to be present in the outer membrane of B. fragilis as an iron-regulated protein.</p></div

Four Conventional Soybean [Glycine max (L.) Merrill] Seeds Exhibit Different Protein Profiles As Revealed by Proteomic Analysis

Soybeans have several functional properties due to their composition and may exert beneficial health effects that are attributed to proteins and their derivative peptides. The present study aimed to analyze the protein profiles of four new conventional soybean seeds (BRS 257, BRS 258, BRS 267, and Embrapa 48) with the use of proteomic tools. Two-dimensional (2D) and one-dimensional (1D) gel electrophoreses were performed, followed by MALDI-TOF/TOF and ESI-Q-TOF mass spectrometry analyses, respectively. These two different experimental approaches allowed the identification of 117 proteins from 1D gels and 46 differentially expressed protein spots in 2D gels. BRS 267 showed the greatest diversity of identified spots in the 2D gel analyses. In the 1D gels, the major groups were storage (25–40%) and lipid metabolism (11–25%) proteins. The differences in protein composition between cultivars could indicate functional and nutritional differences and could direct the development of new cultivars

Genotyping and descriptive proteomics of a potential zoonotic canine strain of Giardia duodenalis, infective to mice

The zoonotic potential of giardiasis, as proposed by WHO since the late 70's, has been largely confirmed in this century. The genetic assemblages A and B of Giardia duodenalis are frequently isolated from human and canine hosts. Most of the assemblage A strains are not infective to adult mice, which can limit the range of studies regarding to biology of G. duodenalis, including virulence factors and the interaction with host immune system. This study aimed to determine the infectivity in mice of an assemblage A Giardia duodenalis strain (BHFC1) isolated from a dog and to classify the strain in sub-assemblages (Al, All, AIII) through the phylogenetic analysis of beta-giardin (bg), triose phosphate isomerase (tpi) and glutamate dehydrogenase (gdh) genes. In addition, the proteomic profile of soluble and insoluble protein fractions of trophozoites was analyzed by 2D-electrophoresis. Accordingly, trophozoites of BHFC1 were highly infective to Swiss mice. The phylogenetic analysis of tpi and gdh revealed that BHFC1 clustered to sub-assemblage Al. The proteomic map of soluble and insoluble protein fractions led to the identification of 187 proteins of G. duodenalis, 27 of them corresponding to hypothetical proteins. Considering both soluble and soluble fractions, the vast majority of the identified proteins (n = 82) were classified as metabolic proteins, mainly associated with carbon and lipid metabolism, including 53 proteins with catalytic activity. Some of the identified proteins correspond to antigens while others can be correlated with virulence. Besides a significant complementation to the proteomic data of G. duodenalis, these data provide an important source of information for future studies on various aspects of the biology of this parasite, such as virulence factors and host and pathogen interactions1110CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE MINAS GERAIS - FAPEMIGsem informaçã

Genotyping and Descriptive Proteomics of a Potential Zoonotic Canine Strain of Giardia duodenalis, Infective to Mice

Submitted by Sandra Infurna ([email protected]) on 2017-03-02T18:57:50Z No. of bitstreams: 1 alex_chapeaurouge_etal_IOC_2016.pdf: 1813108 bytes, checksum: d2d4db74321df407e1868919c06835d6 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2017-03-02T19:11:30Z (GMT) No. of bitstreams: 1 alex_chapeaurouge_etal_IOC_2016.pdf: 1813108 bytes, checksum: d2d4db74321df407e1868919c06835d6 (MD5)Made available in DSpace on 2017-03-02T19:11:30Z (GMT). No. of bitstreams: 1 alex_chapeaurouge_etal_IOC_2016.pdf: 1813108 bytes, checksum: d2d4db74321df407e1868919c06835d6 (MD5) Previous issue date: 2016Universidade Federal de Minas Gerais. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Belo Horizonte, MG, Brasil.Fundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Belo Horizonte, MG, Brasil.Universidade de Campinas. Centro de Biologia Molecular e Engenharia Genética. Campinas, SP, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Toxinologia. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Toxinologia. Rio de Janeiro, RJ, Brasil.Universidade Federal de Minas Gerais. Departamento de Biologia Geral. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Departamento de Parasitologia. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Belo Horizonte, MG, Brasil.Georgetown University. Biology Department. Washington, USA.Fundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Belo Horizonte, MG, Brasil.The zoonotic potential of giardiasis, as proposed by WHO since the late 70's, has been largely confirmed in this century. The genetic assemblages A and B of Giardia duodenalis are frequently isolated from human and canine hosts. Most of the assemblage A strains are not infective to adult mice, which can limit the range of studies regarding to biology of G. duodenalis, including virulence factors and the interaction with host immune system. This study aimed to determine the infectivity in mice of an assemblage A Giardia duodenalis strain (BHFC1) isolated from a dog and to classify the strain in sub-assemblages (AI, AII, AIII) through the phylogenetic analysis of beta-giardin (bg), triose phosphate isomerase (tpi) and glutamate dehydrogenase (gdh) genes. In addition, the proteomic profile of soluble and insoluble protein fractions of trophozoites was analyzed by 2D-electrophoresis. Accordingly, trophozoites of BHFC1 were highly infective to Swiss mice. The phylogenetic analysis of tpi and gdh revealed that BHFC1 clustered to sub-assemblage AI. The proteomic map of soluble and insoluble protein fractions led to the identification of 187 proteins of G. duodenalis, 27 of them corresponding to hypothetical proteins. Considering both soluble and soluble fractions, the vast majority of the identified proteins (n = 82) were classified as metabolic proteins, mainly associated with carbon and lipid metabolism, including 53 proteins with catalytic activity. Some of the identified proteins correspond to antigens while others can be correlated with virulence. Besides a significant complementation to the proteomic data of G. duodenalis, these data provide an important source of information for future studies on various aspects of the biology of this parasite, such as virulence factors and host and pathogen interactions

2D protein map showing the spots of the soluble protein fraction (Proteins 1 to 429) and the numerical distribution of localized proteins.

<p>The protein identification correspondent to each number is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0164946#pone.0164946.s003" target="_blank">S3 Table</a>.</p