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Master's Dissertation
DOI
Document
Author
Full name
Angélica Silva
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
Ribeirão Preto, 2018
Supervisor
Committee
Figueiredo, Luiz Tadeu Moraes (President)
Lima, Danielle Malta
Machado, Cleni Mara Marzocchi
Arruda Neto, Eurico de
Title in Portuguese
Ensaio imunoenzimático utilizando células infectadas como antígeno para diagnóstico de infecções causadas pelos vírus Zika e Chikungunya
Keywords in Portuguese
Arbovírus
Chikungunya
Diagnóstico
Vírus
Zika
Abstract in Portuguese
No Brasil os vírus Zika e Chikungunya vem causando enormes epidemias com grande morbidade e significante mortalidade, tornou-se importante desenvolver métodos diagnósticos eficazes para discrimar suas infecções. No presente trabalho, desenvolvemos ensaios imunoenzimáticos utilizando células de cultura, aedes albopictus pseudo C6/36, infectadas e utilizadas como antígeno viral (EIA-ICC), para diagnósticos de infecções provocadas pelos vírus Zika (ZIKV) e Chikungunya (CHIKV). As placas contendo células infectadas tiveram suas antigenicidades verificadas nos ensaios utilizando anticorpos policlonais oriundo de camundongos hiperimunizados com ZIKV, CHIKV e outros flavivirus e alphavirus. O EIAICC ZIKV detectou anticorpos murinos de forma homotípica, porém reações cruzadas foram encontradas com Dengue 1 (DENV 1), Saint Louis Vírus (SLEV), Rocio Vírus (ROCV) e West Nilo Vírus (WNV). O EIA-ICC CHIKV também detectou anticorpos murinos de forma homotípica e reações cruzadas foram observadas com os Mayaro Vírus (MAYV) e Mucambo Vírus (MUCV). Resultados comparativos de 39 amostras foram realizados no EIA-ICC ZIKV, que demonstrou sensibilidade de 87,5% (IgM) e 100% (IgG) e especificidade de 95,65% (IgM) e 83,33% (IgG). O EIA-ICC CHIKV, teste comparativos entre 59 amostras, demostrou sensibilidade de 64,7% (IgM) e 92,3% (IgG) e especificidade de 40% (IgM) e 96,96% (IgG). Os EIA-ICCs também foram usados para diagnósticos das arboviroses ZIKV e CHIKV em 174 soros de pacientes com doença febril aguda, no EIA-ICC ZIKV foi encontrado 8,13% de IgM, 20,34% IgG e 6,97% de IgG e IgM na mesmo soro, no EIA-ICC CHIKV encontramos IgG em 8,13% dos soros e IgG em 16,86%, soros com anticorpos IgG e IgM nos mesmos foram vistos em 1,74%. Também encontramos em 11 soros anticorpos tanto para ZIKV quanto para CHIKV. Portanto o EIA-ICC ZIKV mostrou para detecção de IgM e IgG, adequadamente sensível e específico para ser validado em estudos com maiores números de amostras e ser aplicados no diagnóstico. O EIA-ICC CHIKV mostrou para detecção de IgG, adequadamente sensível e específico para ser validado em estudos com maiores números de amostras e ser aplicados no diagnóstico. Porém não é indicado para a detecção de IgM
Title in English
Enzymme immunoassay using infected cells as antigen for diagnosis of infections by Zika vírus and Chikungunya vírus
Keywords in English
Arbovirus
Chikungunya
Diagnosis
Virus
Zika
Abstract in English
In Brazil, Zika and Chikungunya viruses have been causing huge epidemics with great morbidity and significant mortality. It's important to develop effective diagnostic methods to discriminate against their infections. In the present study, we developed immunoenzymatic assays using cultured cells from aedes albopictus C6 / 36 infected and used as viral antigen (EIA-ICC), for diagnosis of infections caused by the Zika (ZIKV) and Chikungunya (CHIKV) viruses. Plates containing infected cells had their antigenicities verified in the assays using polyclonal antibodies from mice hyperimmunized with ZIKV, CHIKV and other flaviviruses and alphaviruses. The EIA-ICC ZIKV detected murine antibodies in a homotypic manner, but cross-reactions were found with Dengue 1 virus (DENV 1), Saint Louis virus (SLEV), Rocio virus (ROCV) and West Nile virus (WNV). The EIA-ICC CHIKV also detected murine antibodies in a homotypic manner and cross-reactions were observed with the Mayaro virus (MAYV) and Mucambo virus (MUCV). Comparative results of 39 samples were performed in the EIA-ICC ZIKV, which showed sensitivity of 87.5% (IgM) and 100% (IgG) and specificity of 95.65% (IgM) and 83.33% (IgG). The EIA-ICC CHIKV, a comparative test among 59 samples, showed a sensitivity of 64.7% (IgM) and 92.3% (IgG) and a specificity of 40% (IgM) and 96.96% (IgG). The EIA-ICCs were also used for diagnosis of ZIKV and CHIKV arboviruses in 174 sera from patients with acute febrile illness. In the EIA-ICC, ZIKV found 8.13% IgM, 20.34% IgG and 6.97% IgG and IgM in the same serum, in the EIA-ICC CHIKV we found IgG in 8.13% of the sera and IgG in 16.86%, sera with IgG and IgM antibodies in them were seen in 1.74%. We also found 11 samples positive for both ZIKV and CHIKV. Therefore, the EIA-ICC ZIKV showed for detection of IgM and IgG, adequately sensitive and specific to be validated in studies with larger numbers of samples and to be applied in the diagnosis. The EIA-ICC CHIKV showed for IgG detection, adequately sensitive and specific to be validated in studies with larger numbers of samples and to be applied in the diagnosis. However, it is not indicated for the detection of IgM
 
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Publishing Date
2019-06-26
 
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