Performance of recombinant chimeric proteins in the serological diagnosis of Trypanosoma cruzi infection in dogs.

Background: Dogs are considered sentinels in areas of Trypanosoma cruzi transmission risk to humans. ELISA is generally the method of choice for diagnosing T. cruzi exposure in dogs, but its performance substantially depends on the antigenic matrix employed. In previous studies, our group has developed four chimeric antigens (IBMP-8.1, 8.2, 8.3, and 8.4) and evaluated their potential for diagnosing T. cruzi exposure in humans. For human sera, these chimeric antigens presented superior diagnostic performances as compared to commercial tests available in Brazil, Spain, and Argentina. Therefore, in this study we have evaluated the potential of these antigenic proteins for detection of anti-T. cruzi IgG antibodies in dog sera. Methodology/Principal findings: The IBMP-ELISA assays were optimized by checkerboard titration. Subsequently, the diagnostic potential was validated through analysis of ROC curves and the performance of the tests was determined using double entry tables. Cross-reactivity was also evaluated for babesiosis, ehrlichiosis, dirofilariosis, anaplasmosis, and visceral leishmaniasis. Best performance was shown by IBMP-8.3 and IBMP-8.4, although all four antigens demonstrated a high diagnostic performance with 46 positive and 149 negative samples tested. IBMP-8.3 demonstrated 100% sensitivity, followed by IBMP-8.4 (96.7?100%), IBMP-8.2 (73.3?87.5%), and IBMP-8.1 (50?100%). The highest specificities were achieved with IBMP-8.2 (100%) and IBMP-8.4 (100%), followed by IBMP-8.3 (96.7?97.5%) and IBMP 8.1 (89.1?100%). Conclusions/Significance: The use of chimeric antigenic matrices in immunoassays for anti-T. cruzi IgG antibody detection in sera of infected dogs was shown to be a promising tool for veterinary diagnosis and epidemiological studies. The chimeric antigens used in this work allowed also to overcome the common hurdles related to serodiagnosis of T. cruzi infection, especially regarding variation of efficiency parameters according to different strains and cross-reactivity with other infectious diseases

Avaliação e validação de proteínas quiméricas do tryponosoma cruzi no diagnóstico da doença de chagas em cães, Brasil

Submitted by Repositório Arca ([email protected]) on 2019-10-02T14:23:37Z No. of bitstreams: 1 license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2019-10-03T11:53:51Z (GMT) No. of bitstreams: 2 Leonardo Maia Leony. Avaliação e validação...2019.pdf: 8981403 bytes, checksum: ca0c382c92842bf98a5a65a099500974 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5)Made available in DSpace on 2019-10-03T11:53:51Z (GMT). No. of bitstreams: 2 Leonardo Maia Leony. Avaliação e validação...2019.pdf: 8981403 bytes, checksum: ca0c382c92842bf98a5a65a099500974 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2019Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES) – Código de Aperfeiçoamento 001”. IGM – Fiocruz - BA FAPESB.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Salvador, BA, Brasil.Os cães domésticos são considerados animais sentinela e reservatórios para doença de Chagas (DC), devido ao seu papel na manutenção do ciclo de transmissão em ambientes domésticos e sua correlação com a prevalência da DC humana. O ELISA indireto é geralmente a metodologia escolhida para o diagnóstico, no entanto, seu desempenho depende substancialmente dos antígenos empregados. Uma estratégia para ultrapassar esta limitação é utilizando antígenos quiméricos, compostos por sequências peptídicas antigênicas, imunodominantes, conservadas e repetitivas de múltiplas proteínas do parasita. Por isto, o nosso grupo desenvolveu quatro antígenos quiméricos (IBMP-8.1, 8.2, 8.3 e 8.4), seu potencial diagnóstico foi avaliado para o diagnóstico da DC humana, no qual os antígenos quiméricos apresentaram um desempenho diagnóstico superior em comparação aos dos testes comercialmente disponíveis no Brasil, Espanha e Argentina. OBJETIVO: Este estudo teve como objetivo avaliar a capacidade sorodiagnóstica dos antígenos IBMP frente às amostras de cães naturalmente e experimentalmente infectados com Trypanosoma cruzi. MATERIAIS E MÉTODOS: Os ensaios foram otimizados por checkerboard titration. Posteriormente, o potencial diagnóstico foi validado através de curvas ROC e o desempenho dos testes foi determinado utilizando tabelas de dupla entrada. A reatividade cruzada também foi avaliada para babesiose, erliquiose, dirofilariose, anaplasmose e leishmaniose. RESULTADOS: Todos os antígenos quiméricos demonstraram um alto desempenho diagnóstico, especialmente o IBMP-8.3 e IBMP-8.4. O antígeno IBMP-8.3 demonstrou uma sensibilidade de 100%, seguido pelo IBMP-8.4 (96.7% - 100%), IBMP-8.2 (73.3% - 87.5%) e IBMP-8.1 (50% - 100%). As especificidades mais elevadas dos antígenos foram para o IBMP-8.2 (100%) e IBMP-8.4 (100%), seguido por IBMP-8.3 (96.7% - 97.5%) e IBMP 8.1 (89.1% -100%). CONCLUSÕES: Portanto, o uso de antígenos quiméricos em imunoensaios para diagnosticar a DC em cães é uma ferramenta promissora para fins veterinários e epidemiológicos. O uso de antígenos quiméricos apresentou uma maior resistência à problemas comuns observados no diagnóstico sorológico de DC, especialmente em relação à variação dos parâmetros de eficiência de acordo com a cepa do parasita e reatividade cruzada com outras doenças parasitárias.Dogs are considered a sentinel group for Chagas disease, due to their role in maintaining the transmission cycle in domestic environments and their correlation with humans’ Chagas prevalence. ELISA is the generally chosen methodology for diagnosis, however, its performance depends substantially on the employed antigenic matrix. A strategy to overcome this limitation is by utilizing chimeric antigens, consisting of antigenic, immunodominant, conserved and repeating amino acid sequences of the parasite. As such, our group developed four chimeric antigens (IBMP-8.1, 8.2, 8.3 and 8.4) and their diagnostic potential was previously evaluated for human’s diagnosis, in which the chimeric antigens presented a superior diagnostic performance compared to commercially available tests in Brazil, Spain, and Argentina. OBJECTIVE: This study aimed to evaluate the potential diagnostic ability of these antigenic proteins for Trypanosoma cruzi infection in dogs. MATERIALS AND METHODS: The assays were optimized by checkerboard titration. Subsequently, the diagnostic potential was validated through ROC curves and the tests performance was determined using double entry tables. Cross-reactivity was also evaluated for babesiosis, ehrlichiosis, dirofilariasis, anaplasmosis and leishmaniasis. RESULTS: All chimeric antigens demonstrated a high diagnostic performance, especially IBMP-8.3 and IBMP-8.4. IBMP-8.3 antigen demonstrated a 100% sensitivity, followed by IBMP-8.4 (96.7% - 100%), IBMP-8.2 (73.3% - 87.5%), and IBMP-8.1 (50% - 100%). The highest specificities of the antigens were for IBMP-8.2 (100%) and IBMP-8.4 (100%), followed by IBMP-8.3 (96.7% - 97.5%) and IBMP 8.1 (89.1%-100%). CONCLUSIONS AND STUDY CONTRIBUTIONS: Therefore, the use of chimeric antigenic matrices in immunoassays to diagnose CD in dogs is a highly promising tool for veterinary and epidemiological purposes. The use of chimeric antigens also efficiently addressed the common hurdles seen in CD serodiagnosis, especially regarding variation in efficiency parameters according to parasite’s strain and cross-reactivity with other infectious diseases

Alterations in the lipid profiles and circulating liver enzymes in individuals infected by Schistosoma mansoni

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2019-02-01T18:03:19Z No. of bitstreams: 1 SILVA, F.L. Alterations in lipid profile...2018.pdf: 2734859 bytes, checksum: 8b8fe8385076ecbba9e7c3b334824b27 (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2019-02-01T18:11:28Z (GMT) No. of bitstreams: 1 SILVA, F.L. Alterations in lipid profile...2018.pdf: 2734859 bytes, checksum: 8b8fe8385076ecbba9e7c3b334824b27 (MD5)Made available in DSpace on 2019-02-01T18:11:28Z (GMT). No. of bitstreams: 1 SILVA, F.L. Alterations in lipid profile...2018.pdf: 2734859 bytes, checksum: 8b8fe8385076ecbba9e7c3b334824b27 (MD5) Previous issue date: 2018Gonçalo Moniz Institute (Fiocruz-BA).Fundação Oswaldo Cruz. Instituto Aggeu Magalhães. Laboratório de Análise de Sistemas de Informações em Saúde. Recife, PE, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Laboratório Avançado de Saúde Pública. Salvador, BA, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Laboratório de Interação Parasito-Hospedeiro e Epidemiologia. Salvador, BA, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Laboratório Avançado de Saúde Pública. Salvador, BA, Brasil.Grupo Promedica. Laboratório Datalab. Salvador, BA, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Laboratório Avançado de Saúde Pública. Salvador, BA, Brasil.Portal hypertension and periportal fibrosis commonly occur in severe Schistosoma mansoni infection. Changes in lipid profile and elevated levels of circulating liver enzymes have also been described in infected individuals. The present study sought to assess the alterations in laboratory parameters associated with liver disorder in individuals infected by S. mansoni who visited a private routine laboratory service. Levels of circulating liver enzymes (gamma-glutamyl transferase [γ-GT], aspartate transaminase [AST], alanine transaminase [ALT], and alkaline phosphatase [ALP]) and a lipid panel (total cholesterol [COL], high-density lipoprotein [HDL], low-density lipoprotein [LDL], very low-density lipoprotein [VLDL], and triglycerides [TRI]) were evaluated in both infected and non-infected individuals and relative risk was used to measure associations. Methods: Data were collected for analysis from a total of 1,078 cases identified in 379,600 individuals who submitted samples to the Datalab Laboratory (Salvador, Bahia) between 2004 and 2008. Results: S. mansoni infection led to increased circulating levels of γ-GT in both women and men, AST (women), and ALP (men). S. mansoni infection was a protective factor against increased levels of TRI, CHO, and VLDL for individuals aged 19 years or older. The results of our analysis indicate that alterations in lipid metabolism and circulating liver enzymes in asymptomatic S. mansoni-infected individuals might be attributed to eggs lodged in the hepatic sinusoids. Conclusions: Parasitological testing for S. mansoni should be indicated in endemic areas when this pattern of alterations is detected

Alterations in the lipid profiles and circulating liver enzymes in individuals infected by Schistosoma mansoni

Abstract INTRODUCTION Portal hypertension and periportal fibrosis commonly occur in severe Schistosoma mansoni infection. Changes in lipid profile and elevated levels of circulating liver enzymes have also been described in infected individuals. The present study sought to assess the alterations in laboratory parameters associated with liver disorder in individuals infected by S. mansoni who visited a private routine laboratory service. Levels of circulating liver enzymes (gamma-glutamyl transferase [γ-GT], aspartate transaminase [AST], alanine transaminase [ALT], and alkaline phosphatase [ALP]) and a lipid panel (total cholesterol [COL], high-density lipoprotein [HDL], low-density lipoprotein [LDL], very low-density lipoprotein [VLDL], and triglycerides [TRI]) were evaluated in both infected and non-infected individuals and relative risk was used to measure associations. METHODS Data were collected for analysis from a total of 1,078 cases identified in 379,600 individuals who submitted samples to the Datalab Laboratory (Salvador, Bahia) between 2004 and 2008. RESULTS S. mansoni infection led to increased circulating levels of γ-GT in both women and men, AST (women), and ALP (men). S. mansoni infection was a protective factor against increased levels of TRI, CHO, and VLDL for individuals aged 19 years or older. The results of our analysis indicate that alterations in lipid metabolism and circulating liver enzymes in asymptomatic S. mansoni-infected individuals might be attributed to eggs lodged in the hepatic sinusoids. CONCLUSIONS Parasitological testing for S. mansoni should be indicated in endemic areas when this pattern of alterations is detected

Correction: Performance of Treponema pallidum recombinant proteins in the serological diagnosis of syphilis.

[This corrects the article DOI: 10.1371/journal.pone.0234043.]

Performance of Treponema pallidum recombinant proteins in the serological diagnosis of syphilis.

Syphilis serodiagnosis is challenging because distinct clinical forms of the infection may influence serological performance and discordant results between tests make clinical decisions difficult. Several recombinant Treponema pallidum-proteins have already been tested for syphilis diagnosis and they are critical to achieve high accuracy in serological testing. Our aim was to assess the varied from performance of T. pallidum-recombinant proteins TmpA, TpN17 and TpN47 for syphilis serodiagnosis. The proteins were evaluated using sera of 338 T. pallidum-negative, 173 T. pallidum-positive individuals and 209 sera from individuals infected with unrelated diseases. The diagnostic potential was validated by analysis of ROC curves. In the liquid microarray analyses, the ROC curve varied from 99.0% for TmpA and TpN17 to 100% for TpN47. The sensitivity score yielded values of up to 90% for TpN17, 100% for TpN47 and 80.0% for TmpA. The lowest and highest specificity values were presented by TpN47 (91.9%) and TmpA antigens (100%), respectively. TpN47 showed the highest accuracy score (95.5%) among all the recombinant proteins assayed. For the ELISA, the ROC curve was 97.2%, 91.8% and 81.6% for TpN17, TmpA and TpN47, respectively. TpN17 and TmpA yielded a sensitivity of 69.9%, while TpN47 obtained a value of 53.8%. Specificity was almost 100% for all three proteins. No cross-reaction was observed for TpN17 in the serum samples from non-bacterial infections. Regarding leptospirosis-positive samples, cross-reactivity score was varied from 8.6 to 34.6%. This is most probably due to conservation of the epitopes in these proteins across bacteria. The use of recombinant proteins in immunoassays for syphilis diagnosis was showed provide greater reliability to results of the treponemal assays. Despite the low sensitivity, the proteins showed high diagnostic capacity due to the AUC values found. However, an improvement in sensitivity could be achieved when antigenic mixtures are evaluated

Detection of anti-Trypanosoma cruzi antibodies by chimeric antigens in chronic Chagas disease-individuals from endemic South American countries

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2019-05-07T14:26:01Z No. of bitstreams: 1 Del-Rei et al., Detection ...2019.pdf: 2364416 bytes, checksum: 1fb04e34df00f90597065a5a189a69b6 (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2019-05-07T16:39:03Z (GMT) No. of bitstreams: 1 Del-Rei et al., Detection ...2019.pdf: 2364416 bytes, checksum: 1fb04e34df00f90597065a5a189a69b6 (MD5)Made available in DSpace on 2019-05-07T16:39:03Z (GMT). No. of bitstreams: 1 Del-Rei et al., Detection ...2019.pdf: 2364416 bytes, checksum: 1fb04e34df00f90597065a5a189a69b6 (MD5) Previous issue date: 2019Fundación Bunge y Born 2012 Dr. Silvia Andrea Longhi; Consejo Nacional de Investigaciones Científicas y Tecnológicas PIP/0974-2011 Dr. Alejandro Gabriel Schijman; Gonçalo Moniz Institute PROEP/IGM 400904/2013-6 Dr. Mitermayer Galvão dos Reis;Coordination of Superior Level Staff Improvement CAPES - PROEX 0720/2018 Dr. Fred Luciano Neves Santos; National Council for Scientific and Technological Development CNPq Proc. No. 312195/2015-0 Dr. Nilson Ivo Tonin Zanchin; and National Council for Scientific and Technological Development CNPq Proc. No. 307319/2016-4 Dr. Mitermayer Galvão dos Reis. The funders were not involved in the design of the study, in the collection, analysis and interpretation of the data, or in writing the manuscript.Faculty of Technology and Sciences of Bahia. Salvador, BA, Brazil.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Salvador, BA, Brasil.Molecular Biology Institute of Paraná. Curitiba, PR, Brazil.Fundação Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Salvador, BA, Brasil / Federal University of Bahia. Department of Pathology and Legal Medicine. Salvador, BA, Brazil / Yale University. School of Public Health. Department of Epidemiology of Microbial Diseases. New Haven, Connecticut, United States of America.Fundação Oswaldo Cruz. Instituto Aggeu Magalhães. Recife, PE, Brasil.Institute for Research on Genetic Engineering and Molecular Biology “Dr Héctor Torres”. Laboratory of Molecular Biology of Chagas Disease. Buenos Aires, Argentina.Institute for Research on Genetic Engineering and Molecular Biology “Dr Héctor Torres”. Laboratory of Molecular Biology of Chagas Disease. Buenos Aires, Argentina.Fundação Oswaldo Cruz. Instituto Gonçalo Moniz. Salvador, BA, Brasil.Laboratory diagnosis of chronic Chagas disease is a troubling factor due to lack of reference tests. The WHO suggests the use of two distinct commercial serological tests in parallel. The performance of commercial immunoassays might fluctuate depending on the antigenic matrices and the local strains of T. cruzi in different geographical settings. The use of antigenic matrices based on chimeric proteins can solve these limitations. Here, we evaluated the diagnostic performance of two chimeric T. cruzi antigens (IBMP-8.1 and -8.4) to diagnose chronic Chagas disease in individuals from endemic South American countries

Stability Assessment of Four Chimeric Proteins for Human Chagas Disease Immunodiagnosis

The performance of an immunoassay relies on antigen-antibody interaction; hence, antigen chemical stability and structural integrity are paramount for an efficient assay. We conducted a functional, thermostability and long-term stability analysis of different chimeric antigens (IBMP), in order to assess effects of adverse conditions on four antigens employed in ELISA to diagnose Chagas disease. ELISA-based immunoassays have served as a model for biosensors development, as both assess molecular interactions. To evaluate thermostability, samples were heated and cooled to verify heat-induced denaturation reversibility. In relation to storage stability, the antigens were analyzed at 25 °C at different moments. Long-term stability tests were performed using eight sets of microplates sensitized. Antigens were structurally analyzed through circular dichroism (CD), dynamic light scattering, SDS-PAGE, and functionally evaluated by ELISA. Data suggest that IBMP antigens are stable, over adverse conditions and for over a year. Daily analysis revealed minor changes in the molecular structure. Functionally, IBMP-8.2 and IBMP-8.3 antigens showed reactivity towards anti-T. cruzi antibodies, even after 72 h at 25 °C. Long-term stability tests showed that all antigens were comparable to the control group and all antigens demonstrated stability for one year. Data suggest that the antigens maintained their function and structural characteristics even in adverse conditions, making them a sturdy and reliable candidate to be employed in future in vitro diagnostic tests applicable to different models of POC devices, such as modern biosensors in development

Global incidence in Brazil per 100,000 inhabitants of CS (A) and SiP (B) according to SINAN.

Joinpoint regression analysis using APC (Annual Percentage Change) calculations identified three distinct periods for each form of syphilis.</p

Spatiotemporal distribution of SiP cases (2001 to 2017) in Brazil, based on cases by microregions.

Digital maps in the public domain (publicly available) were obtained from IBGE cartographic database in shapefile format (.shp), which was subsequently reformatted and analyzed using QGIS version 3.10. Authors specify that this figure is licensed under CC BY 4.0.</p