Rapid and Sensitive Qualitative Duoplex Real-Time PCR Method for Discriminatory and Confirmatory Diagnosis of HTLV-1 and HTLV-2 Infections: Brazilian Multicentric Study.

Human T cell lymphotropic virus (HTLV) is the caustive agent of two main conditions i. e., the HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and the adult T-cell leukemia/lymphoma (ATLL). HTLV diagnosis is based on serological and molecular approaches; however, an accurate and validated method is still needed. The objective of this study was to establish a rapid and sensitive molecular test to confirm and discriminate HTLV 1/2 types. The test validation was performed as a multicentric study involving HTLV confirmation centers throughout Brazil. Proviral DNA was extracted from whole blood and the amplification was performed using in-house designed primer and probe sets targeting the pol genomic region. An internal control to validate the extraction and amplification was also included. The limit of detection (LoD) of the assay was four copies/reaction for HTLV-1 and 10.9 copies/reaction for HTLV-2. The diagnostic sensitivity of the platform was 94.6% for HTLV-1, 78.6% for HTLV-2, and the specificity was 100% for both viruses. Cross-reactions of the test with human viruses including HAV, HBV, HCV, HIV-1/2, and parvovirus B19 were not observed. During the multicentric validation, the test was used to screen a total of 692 blood samples obtained from previously confirmed HTLV-positive individuals. From these, 91.1% tested positive being concordant with the previously obtained results. In conclusion, our duoplex-RT-PCR-HTLV1 /2 presented adequate efficiency for HTLV-1/2 differentiation showing high sensitivity and specificity. Therefore, it can be a suitable tool for confirmation of suspected and inconclusive HTLV cases, prenatal and pre-transplant diagnosis, in Brazil and in other countries HTLV-endemic countries.

New diagnostic criteria for neurocysticercosis: Reliability and validity.

OBJECTIVE: The diagnosis of neurocysticercosis (NCC) remains problematic because of the heterogeneity of its clinical, immunological, and imaging characteristics. Our aim was to develop and assess a new set of diagnostic criteria for NCC, which might allow for the accurate detection of, and differentiation between, parenchymal and extraparenchymal disease. METHODS: A group of Latin American NCC experts developed by consensus a new set of diagnostic criteria for NCC. A multicenter, retrospective study was then conducted to validate it. The reference standard for diagnosis of active NCC was the disappearance or reduction of cysts after anthelmintic treatment. In total, three pairs of independent neurologists blinded to the diagnosis evaluated 93 cases (with NCC) and 93 controls (without NCC) using the new diagnostic criteria. Mixed-effects logistic regression models were used to estimate sensitivity and specificity. RESULTS: Inter-rater reliability (kappa) of diagnosis among evaluators was 0.60. For diagnosis of NCC versus no NCC, the new criteria had a sensitivity of 93.2% and specificity of 81.4%. For parenchymal NCC, the new criteria had a sensitivity of 89.8% and specificity of 80.7% and for extraparenchymal NCC, the new criteria had a sensitivity of 65.9% and specificity of 94.9%. INTERPRETATION: These criteria have acceptable reliability and validity and could be a new tool for clinicians and researchers. An advantage of the new criteria is that they consider parasite location (ie, parenchymal or extraparenchymal), which is an important factor determining the clinical, immunological, and radiological presentation of the disease, and importantly, its treatment and prognosis. Ann Neurol 2016;80:434-442.

HLA-G 14-bp insertion/deletion polymorphism is a risk factor for HTLV-1 infection.

About 95% of HTLV-1 infected patients remain asymptomatic throughout life, and the risk factors associated with the development of related diseases, such as HAM/TSP and ATL, are not fully understood. The human leukocyte antigen-G molecule (HLA-G), a nonclassical HLA class I molecule encoded by MHC, is expressed in several pathological conditions, including viral infection, and is related to immunosuppressive effects that allow the virus-infected cells to escape the antiviral defense of the host. The 14-bp insertion/deletion polymorphism of exon 8 HLA-G gene influences the stability of the transcripts and could be related to HTLV-1-infected cell protection and to the increase of proviral load. The present study analyzed by conventional PCR the 14-bp insertion/deletion polymorphism of exon 8 HLA-G gene in 150 unrelated healthy subjects, 82 HTLV-1 infected patients with symptoms (33 ATL and 49 HAM), and 56 asymptomatic HTLV-1 infected patients (HAC). In addition, the proviral load was determined by quantitative real-time PCR in all infected groups and correlated with 14-bp insertion/deletion genotypes. The heterozygote genotype frequencies were significantly higher in HAM, in the symptomatic group, and in infected patients compared to control (p < 0.05). The proviral load was higher in the symptomatic group than the HAC group (p < 0.0005). The comparison of proviral load and genotypes showed that -14-bp/-14-bp genotype had a higher proviral load than +14-bp/-14-bp and +14-bp/+14-bp genotypes. Although HLA-G 14-bp polymorphism does not appear to be associated with HTLV-1 related disease development, it could be a genetic risk factor for susceptibility to infection.

[Cloning and expression of the transmembranic glycoprotein from human T cell lymphotropic virus in a prokaryotic system]. / Clonagem e expressão da glicoproteína transmembrana do vírus linfotrópico de células T humanas em sistema procarioto.

HTLV-1 is the virus that causes T cell lymphoma/leukemia in adults and a neurological disorder known as HTLV-associated myelopathy or tropical spastic paraparesis. One of the transmission means is through contaminated blood and its byproducts. Because of the risk of HTLV-associated infections, screening for HTLV was introduced for Brazilian blood donors in 1993. Most of the diagnostic kits used in the national blood banks are bought from foreign companies. Brazil does not have the technology to produce this material and there is a need to produce diagnostic systems with national technology. In this study, we show the expression of gp21/HTLV-1 in Escherichia coli and its reactivity towards monoclonal antibodies and the antibodies of infected patients. Expressing these proteins is the first step towards obtaining diagnostic kits with Brazilian biotechnology.

Clonagem e expressão da glicoproteína transmembrana do vírus linfotrópico de células T humanas em sistema procarioto / Cloning and expression of the transmembranic glycoprotein from human T cell lymphotropic virus in a prokaryotic system

O HTLV-1 é o vírus causador da leucemia/linfoma de célula T no adulto e de uma desordem neurológica conhecida por mielopatia associada ao HTLV ou paraparesia espástica tropical. Um dos modos de transmissão é pelo sangue contaminado e seus subprodutos e, devido ao risco de infecções associadas ao HTLV sua pesquisa na triagem de doadores de sangue foi introduzida no Brasil a partir de 1993. Os kits diagnósticos utilizados nos bancos de sangue nacionais são na sua maioria comprados de empresas estrangeiras. O Brasil não detém a tecnologia para produção deste material e há a necessidade de produção de sistemas de diagnóstico com tecnologia nacional. Neste trabalho, mostramos a expressão da gp21/HTLV-1 em Escherichia coli e sua reatividade frente a anticorpos monoclonais e de pacientes infectados. Expressar tais proteínas é o primeiro passo para obtenção de conjuntos diagnósticos com tecnologia brasileira.

HTLV-1 is the virus that causes T cell lymphoma/leukemia in adults and a neurological disorder known as HTLV-associated myelopathy or tropical spastic paraparesis. One of the transmission means is through contaminated blood and its byproducts. Because of the risk of HTLV-associated infections, screening for HTLV was introduced for Brazilian blood donors in 1993. Most of the diagnostic kits used in the national blood banks are bought from foreign companies. Brazil does not have the technology to produce this material and there is a need to produce diagnostic systems with national technology. In this study, we show the expression of gp21/HTLV-1 in Escherichia coli and its reactivity towards monoclonal antibodies and the antibodies of infected patients. Expressing these proteins is the first step towards obtaining diagnostic kits with Brazilian biotechnology.

[Cerebrospinal fluid puncture, consent inform, and ethics in research: the Brazilian Academy of Neurology recommendations]. / Coleta do líquido cefalorraquidiano, termo de consentimento livre e esclarecido e aspectos éticos em pesquisa: recomendações do Departamento Científico de LCR da Academia Brasileira de Neurologia.

The Cerebrospinal Fluid (CSF) Scientific Department of the Brazilian Academy of Neurology (SD-BAN) suggests the use of consent inform for patients submitted to lumbar puncture. It should be explained to the patients the possible complications related to CSF puncture. The laws related to the research in human beings have also been discussed by the CSF SD-BAN.

Coleta do líquido cefalorraquidiano, termo de consentimento livre e esclarecido e aspectos éticos em pesquisa: recomendaçöes do Departamento Científico de LCR da Academia Brasileira de Neurologia / Cerebrospinal fluid puncture, consent inform, and ethics in research: the Brazilian Academy of Neurology recommendations

O Departamento Científico de LCR (DC-LCR) da Academia Brasileira de Neurologia (ABN) recomenda a adoçäo do Termo de Consentimento Livre e Esclarecido (TCLE) como procedimento prévio à punçäo para coleta de LCR, tendo como finalidade o adequado esclarecimento dos pacientes quanto aos riscos do procedimento e às medidas de prevençäo de complicaçöes do exame. O documento final do TCLE foi resultado de consenso entre os membros de DC-LCR. O DC-LCR da ABN considerou também pertinente e importante a divulgaçäo das normas legais relacionadas à pesquisas em seres humanos em nosso país

HTLV-I/HTLV-II / HTLV-I/HTLV-II

O presente volume é uma atualizaçäo sobre os vírus linfotrópicos humanos I e II (HTLV-I/II) e aborda aspectos da biologia, epidemiologia, diagnóstico e aconselhamento de doadores positivos.

HTLV-I/HTLV-II / HTLV-I/HTLV-II

O volume de número III é sobre os vírus linfotrópicos humanos I e II (HTLV-I/II) e aborda aspectos relevantes da biologia, epidemiologia, diagnóstico e aconselhamento de doadores positivos.