Pooling of sera for human T-cell lymphotropic virus (HTLV) screening in a time of increasing health care expenditure and limited resources.

Identifying the true prevalence of human T-cell lymphotropic virus, mostly type 1 (HTLV-1), and the number of patients with HTLV-1-associated diseases, in addition to introducing HTLV-1/2 serology during the prenatal of pregnant women and in individuals infected with other viruses that share transmission routes with HTLV-1, are actions that could help to recognize the importance of this virus by WHO and national health organizations, and to control its transmission/dissemination. As Brazil is endemic to HTLV and there is an increase in health care expenditure, but resources are limited, any strategy that could reduce the cost of HTLV screening is needed and welcomed. This study aimed to determine whether the strategy of pooling sera for HTLV antibody determination is feasible and reduces the costs. Two enzyme immunoassays (EIA Murex HTLV-I+II, Diasorin, UK, and Gold ELISA HTLV-1+2, REM Ind. Com. Ltda., SP, Brazil), and serum samples that resulted in different levels of HTLV-1/2 antibodies by EIA and of which a volume allowed assay validation were employed for analysis. The diagnostic sensitivity and specificity and Cohen's Kappa value, as well as the accuracy and precision were analyzed. After validating the five-sample pool using the EIA Murex (Cohen's Kappa = 1.0), the technique was employed for individual cost comparison in 2,625 serum samples from populations at risk of HTLV infections (HBV, HCV, and HIV-infected individuals). The results from individual and pooled samples confirmed the diagnostic sensitivity (100%) and specificity (100%) of the pooling and a cost minimization varying from 60.7% to 73.6%. In conclusion, the results of this study suggest the use of pooling sera in sero-epidemiological surveillance studies and possibly in prenatal care screening programs in Brazil.

Surveillance of human retroviruses in blood samples from patients with hepatitis B and C in São Paulo, Brazil.

INTRODUCTION: Human retroviruses and the hepatitis B and C viruses (HBV and HCV, respectively) share routes of transmission; thus, coinfections occur and could alter subsequent disease outcomes. A preliminary study on human T-lymphotropic virus types 1 and 2 (HTLV-1/2) in serum samples from HBV- and HCV-infected individuals in São Paulo revealed 1.3% and 5.3% rates of coinfection, respectively. These percentages were of concern since they were detected in HTLV-endemic regions and in high-risk individuals in Brazil. The present study was conducted to extend and confirm these data. METHODS: HTLV-1/2 and human immunodeficiency virus (HIV) infection status were identified in 1,984 sera for HBV and HCV viral load quantification - 1,290 samples from HBV-infected individuals (53.3% men, mean age: 47.1 years) and 694 samples from HCV-infected individuals (56.3% men, mean age: 50.1 years). HTLV-1/2 antibodies were detected by enzyme immunoassay, followed by western blotting and line immunoassay; HIV infection was detected by enzyme immunoassay. RESULTS: HTLV-1/-2 infection was detected in 1.9% HBV-infected individuals (0.7% HTLV-1 and 1.2% HTLV-2) and in 4.0% (2.4% HTLV-1 and 1.6% HTLV-2) HCV-infected individuals; HIV infection was detected in 9.2% and 14.5%, respectively. Strong associations with HTLV and HIV, male sex, and older age were found in HBV/HTLV and HCV/HTLV-coinfected individuals (p<0.05). CONCLUSIONS: HTLV-1 and HTLV-2 were confirmed to be prevalent in individuals with HBV and HCV in São Paulo; coinfected individuals deserve further clinical and laboratory investigation.

Pooling of sera for human t-cell lymphotropic virus (HTLV) screening in a time of increasing health care expenditure and limited resouces

Identifying the true prevalence of human T-cell lymphotropic virus, mostly type 1 (HTLV-1), and the number of patients with HTLV-1-associated diseases, in addition to introducing HTLV-1/2 serology during the prenatal of pregnant women and in individuals infected with other viruses that share transmission routes with HTLV-1, are actions that could help to recognize the importance of this virus by WHO and national health organizations, and to control its transmission/dissemination. As Brazil is endemic to HTLV and there is an increase in health care expenditure, but resources are limited, any strategy that could reduce the cost of HTLV screening is needed and welcomed. This study aimed to determine whether the strategy of pooling sera for HTLV antibody determination is feasible and reduces the costs. Two enzyme immunoassays (EIA Murex HTLV-I+II, Diasorin, UK, and Gold ELISA HTLV-1+2, REM Ind. Com. Ltda., SP, Brazil), and serum samples that resulted in different levels of HTLV-1/2 antibodies by EIA and of which a volume allowed assay validation were employed for analysis. The diagnostic sensitivity and specificity and Cohen's Kappa value, as well as the accuracy and precision were analyzed. After validating the five-sample pool using the EIA Murex (Cohen's Kappa = 1.0), the technique was employed for individual cost comparison in 2,625 serum samples from populations at risk of HTLV infections (HBV, HCV, and HIV-infected individuals). The results from individual and pooled samples confirmed the diagnostic sensitivity (100%) and specificity (100%) of the pooling and a cost minimization varying from 60.7% to 73.6%. In conclusion, the results of this study suggest the use of pooling sera in sero-epidemiological surveillance studies and possibly in prenatal care screening programs in Brazil.

Avaliação do emprego de pool de soros na triagem de infecção por HTLV-1/2 / Evaluation of the use of serum pool in screening for HTLV-1/2 infection

Os vírus linfotrópicos de células T humanas dos tipos um e dois (HTLV-1 e HTLV-2) são endêmicos no Brasil. A triagem para HTLV-1/2 é obrigatória em bancos de sangue no país desde 1993, e a partir de 2014 é recomendada ao menos uma vez no acompanhamento de pacientes com HIV/aids, mas não em outras populações consideradas de risco para adquirir/transmitir esta infecção, como por exemplo, gestantes e pacientes com hepatites virais dos tipos B e C. Como o número de indivíduos em risco para adquirir/transmitir os HTLV a serem testados anualmente no Brasil é alto, qualquer estratégia que reduza o custo da triagem sorológica é necessária e bem vinda. O presente estudo avaliou o desempenho e o custo-minimização do uso de pool de soros na triagem sorológica de infecção por HTLV-1/2. Oitenta e uma amostras de soro sabidamente positivas para HTLV-1/2 foram retestadas utilizando dois ensaios imunoenzimáticos na triagem (EIA Murex HTLV I+II, Diasorin, e GOLD ELISA HTLV-1/2, REM) e dois testes confirmatórios [Western blot (WB), HTLV BLOT 2.4, MP Biomedicals e imunoensaio de linha (LIA), INNO-LIA HTLV I/II Score, Fujirebio], e separadas de acordo com os valores de DO/cut-off em: fortemente reagentes (DO/cut-off >12), e moderadamente reagentes (DO/cut-off >2,0 a 12,0). Posteriormente, estas amostras foram diluídas na razão dois em bolsa de plasma negativa para marcadores de banco de sangue até a perda de reatividade, e em seguida diluídas em diferentes pools de soros positivos e negativos para HIV, HBV e HCV, com vistas a determinar, respectivamente, a maior diluição de soro sem perda de sensibilidade e garantir a especificidade da reação. Subsequentemente, amostras com volume suficiente para ensaios de validação de métodos de diagnóstico segundo os parâmetros estabelecidos pelo Instituto Adolfo Lutz (P-SG-0022) foram selecionadas e testadas quanto à estratégia de pool de soros; 40 pools foram utilizados nos testes de acurácia e sete pools nos de precisão. Para o cálculo de redução de custo (custo-minimização), 2.000 amostras de soro/plasma de pacientes com hepatites virais B e C, e 625 amostras de plasma de pacientes com HIV/aids que haviam sido testadas individualmente e cujos resultados haviam sido publicados foram avaliadas em pool. Os resultados obtidos mostraram que o kit Murex foi mais sensível podendo ser diluído na razão 1:5 sem perda de sensibilidade e especificidade diagnóstica, com resultados de exatidão, precisão, sensibilidade, especificidade, valor preditivo positivo e negativo de 100% (coeficiente de correlação Kappa = 1). Em populações de risco, o uso da estratégia de pool de soros mostrou a mesma sensibilidade da análise individual, e uma redução de custo de 70,4% no grupo HBV, 60,7% no grupo HCV e 73,6% no grupo HIV/aids; estando o custo-minimização relacionado à prevalência da infecção nas populações de estudo: 1,9% (HBV), 4,0% (HCV), e 1,1% (HIV/aids). Concluindo, os resultados obtidos permitem sugerir a introdução da triagem sorológica para HTLV-1/2 utilizando pool de cinco soros e o kit Murex em inquéritos epidemiológicos, no acompanhamento de pacientes com outras infecções virais e possivelmente em gestantes no pré-natal. (AU)

The human T-cell lymphotropic viruses type 1 and type 2 (HTLV-1 and HTLV-2) are endemic in Brazil. HTLV-1/2 screening is mandatory in blood banks in the country since 1993, and from 2014 it is recommended at least once in the follow-up of patients with HIV/AIDS, but not in other populations considered at risk for acquiring/transmitting these infections, such as pregnant women and patients with viral hepatitis B and C. As the number of individuals at risk for acquiring/transmitting HTLV to be tested annually in Brazil is high, any strategy that reduces the cost of screening is necessary and welcomed. The present study evaluated the performance and costminimization of the use of pooling sera in serological screening for HTLV-1/2 infections. Eighty-one HTLV-1/2 truly positive serum samples were retested using two screening enzyme-linked immunoassays (EIA Murex HTLV I + II, Diasorin, and GOLD ELISA HTLV-1/2, REM) and two confirmatory assays [Western blot (WB), HTLV BLOT 2.4, MP Biomedicals and Line Immunoassay (LIA), INNO-LIA HTLV I / II Score, Fujirebio], and separated according to OD/cut-off values into: strongly reagents (OD/cut-off> 12), and moderately reagents (DO/ cut-off> 2.0 to 12.0). Subsequently, these samples were diluted at ratio two in serum from negative blood bag for infectious blood-borne diseases until the loss of reactivity to determine the highest serum dilution without loss of sensitivity, and next diluted in different pools of positive and negative HIV, HBV and HCV sera to ensure the specificity of the pooling strategy. Afterwards, samples with sufficient volume for diagnostic method validation assays according to the parameters established by the Adolfo Lutz Institute (P-SG-0022) were selected and tested for the pooling strategy: 40 pools for accuracy and 7 pools for precision analysis. For mostminimization calculation, 2,000 serum/plasma samples from patients with viral hepatitis B and C and 625 plasma samples from patients with HIV/AIDS that had been individually tested and the results had been published elsewhere were tested in pooling. The results obtained showed that the Murex kit was more sensitive and could be used at 1:5 dilution without loss of sensitivity. The accuracy, precision, sensitivity, specificity, and positive and negative predictive values were 100% (Kappa correlation coefficient = 1). In at-risk populations, the use of the serum pooling strategy reduced the cost of screening (cost-minimization) by 70.4% in the HBV, 60.7% in the HCV, and 73.6% in HIV/AIDS groups of patients, which depended on the prevalence of HTLV-1/2 in such populations: 1.9% (HBV), 4.0% (HCV), e 1.1% (HIV/aids). In conclusion, the results obtained suggest the introduction of HTLV-1/2 serological screening using pooling of five sera and the Murex kit in epidemiological surveys, in the follow-up of patients with other viral infections and possibly in prenatal screening programs. (AU)

Surveillance of human retroviruses in blood samples from patients with hepatitis B and C in São Paulo, Brazil

Abstract INTRODUCTION Human retroviruses and the hepatitis B and C viruses (HBV and HCV, respectively) share routes of transmission; thus, coinfections occur and could alter subsequent disease outcomes. A preliminary study on human T-lymphotropic virus types 1 and 2 (HTLV-1/2) in serum samples from HBV- and HCV-infected individuals in São Paulo revealed 1.3% and 5.3% rates of coinfection, respectively. These percentages were of concern since they were detected in HTLV-endemic regions and in high-risk individuals in Brazil. The present study was conducted to extend and confirm these data. METHODS HTLV-1/2 and human immunodeficiency virus (HIV) infection status were identified in 1,984 sera for HBV and HCV viral load quantification - 1,290 samples from HBV-infected individuals (53.3% men, mean age: 47.1 years) and 694 samples from HCV-infected individuals (56.3% men, mean age: 50.1 years). HTLV-1/2 antibodies were detected by enzyme immunoassay, followed by western blotting and line immunoassay; HIV infection was detected by enzyme immunoassay. RESULTS HTLV-1/-2 infection was detected in 1.9% HBV-infected individuals (0.7% HTLV-1 and 1.2% HTLV-2) and in 4.0% (2.4% HTLV-1 and 1.6% HTLV-2) HCV-infected individuals; HIV infection was detected in 9.2% and 14.5%, respectively. Strong associations with HTLV and HIV, male sex, and older age were found in HBV/HTLV and HCV/HTLV-coinfected individuals (p<0.05). CONCLUSIONS HTLV-1 and HTLV-2 were confirmed to be prevalent in individuals with HBV and HCV in São Paulo; coinfected individuals deserve further clinical and laboratory investigation.