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

Comparison of diagnostic performance of RT-qPCR, RT-LAMP and IgM/IgG rapid tests for detection of SARS-CoV-2 among healthcare workers in Brazil

Background: COVID-19 has become a major public health problem after the outbreak caused by SARS-CoV-2 virus. Great efforts to contain COVID-19 transmission have been applied worldwide. In this context, accurate and fast diagnosis is essential. Methods: In this prospective study, we evaluated the clinical performance of three different RNA-based molecular tests – RT-qPCR (Charité protocol), RT-qPCR (CDC (USA) protocol) and RT-LAMP – and one rapid test for detecting anti-SARS-CoV-2 IgM and IgG antibodies. Results: Our results demonstrate that RT-qPCR using the CDC (USA) protocol is the most accurate diagnostic test among those evaluated, while oro-nasopharyngeal swabs are the most appropriate biological sample. RT-LAMP was the RNA-based molecular test with lowest sensitivity while the serological test presented the lowest sensitivity among all evaluated tests, indicating that the latter test is not a good predictor of disease in the first days after symptoms onset. Additionally, we observed higher viral load in individuals who reported more than 3 symptoms at the baseline. Nevertheless, viral load had not impacted the probability of testing positive for SARS-CoV-2. Conclusion: Our data indicates that RT-qPCR using the CDC (USA) protocol in oro-nasopharyngeal swabs samples should be the method of choice to diagnosis COVID-19

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

2D protein map showing the spots of the insoluble fraction (Proteins 430 to 903) 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

Number of trophozoites or cysts recovered from Swiss mice gut after infection with <i>G</i>. <i>duodenalis</i> strains BHCF1 and Portland-1.

<p>Mice were inoculated with 1 x 10⁶ trophozoites through intra-gastric route. Parasites were recovered from gut and counted in Neubauer chamber. Numbers correspond to average and standard deviation of parasite counting’s obtained for each group, from three independent experiments, with minimal number of 3 mice per group, in each experiment.</p

Classification of the identified proteins by molecular function (A), biological process (B) and protein class (C).

<p>This classification is suggested by gene ontology and Panther^® and provides a general characterization of a group of proteins. The numbers presented in each graph correspond to the number of proteins.</p