Dengue diagnostics: serious inaccuracies are likely to occur if pre-analytical conditions are not strictly followed

BACKGROUND The heat-labile nature of Dengue virus (DENV) in serum samples must be considered when applying routine diagnostic tests to avoid issues that could impact the accuracy of test results with direct implications for case management and disease reporting. OBJECTIVES To check if pre-analytical variables, such as storage time and temperature, have an impact on the accuracy of the main routine diagnostic tests for dengue. METHODS Virus isolation, reverse transcription real-time polymerase chain reaction (RT-PCR) and NS1 enzyme-linked immunosorbent assay (ELISA) were evaluated using 84 samples submitted to different pre-analytical conditions. FINDINGS Sensitivity and negative predictive value were directly affected by sample storage conditions. RT-PCR and virus isolation showed greater dependence on well-conserved samples for an accurate diagnosis. Interestingly, even storage at -30ºC for a relatively short time (15 days) was not adequate for accurate results using virus isolation and RT-PCR tests. On the other hand, NS1 ELISA showed no significant reduction in positivity for aliquots tested under the same conditions as in the previous tests. MAIN CONCLUSIONS Our results support the stability of the NS1 marker in ELISA diagnosis and indicate that the accuracy of routine tests such as virus isolation and RT-PCR is significantly affected by inadequate transport and storage conditions of serum samples.

Laboratórios sentinelas - uma proposta para o monitoramento das infecçöes pelos vírus das hepatites A e B / Sentinel laboratories - a proposal for monitoring viral A and B hepatitis infections

Considerando que as hepatites virais (HV) são representativas da categoria de infecções freqüentemente assintomáticas, propõe-se a utilização de um sistema de laboratórios sentinelas (SLS) como método de monitoramento das infecções pelos vírus das hepatites A (VHA) e B (VHB), através da pesquisa de marcadores virais em material sorológico excedente coletado por outros motivos. Por intermédio de revisão bibliográfica e de avaliação da aplicabilidade da estratégia de laboratórios sentinelas no acompanhamento da tendência destas infecções, desenvolvem-se as bases técnicas para um SLS, discutem-se suas potencialidades e limitações, sua importância na complementação das informações da Vigilância Epidemiológica (VE) das hepatites virais e os instrumentos para criteriosa avaliação do sistema, possibilitando correções e redirecionamentos. Suas principais vantagens sobre o sistema de VE passivo são a simplicidade, melhor qualidade das informações e maior abrangência, gerando aceitabilidade e efetividade. De menor custo que os inquéritos de soroprevalência, um SLS apresenta sensibilidade adequada para o monitoramento do VHA e VHB e possibilita o gerenciamento de programas de prevenção.

Viral hepatitis (VH) is representative of a category of infections with a significant fraction of asymptomatic cases. In this paper, the use of a system of sentinel laboratories (SSL) in the monitoring of infections for hepatitis A (HAV) and hepatitis B viruses (HBV) is considered, arguing the importance of complementing the information of Epidemiological Surveillance (ES) and the development of the technical bases of this system, its potential, limitations and evaluation criteria. Through bibliographical review and evaluation of the applicability of the laboratory sentinel technique to follow the trend of these infections, we consider the use of exceeding serologic material, collected for other reasons, for the research of viral markers with the aim of supplementing the information of the ES for prevention programs. The main advantages of this system over the passive system of ES, are a better quality of information, increased simplicity, and enhanced effectiveness and acceptability. It is less costly than serumprevalence surveys,and provides good sensitivity for the monitoring of HAV and HBV

HBV DNA assay using PCR-microfluidic chips for blood screening / 中国输血杂志

Objective To test HBV DNA by using PCR-microfluidic chip assay. Methods Pooled sera ( 5?50ul ) negative for ELISA serological tests were tested for HBV DNA using PCR-microfluidic chips assay. Individual donor samples were tested if the pooled sera were positive. The sensitivity of PCR-microfluidic chips assay was determined by serial dilutions of the standard control serum. The specificity of PCR-microfludic chips assay was also determined by testing 56 various serum samples. Serial dilutions of the standard control sera were tested repeatedly for understanding the inter- and intra-assay variation of this method. Results Seven of 545 nonrenumerated donors (1.28%) were found positive for HBV DNA. The sensitivity of PCR-microfluidic chips assay was 4.81?102copies/ml. The HBV DNA was positive for all 37 samples from HBeAg positive patients. The HBV DNA tests of samples from HCV RNA positive patients, anti-HAV IgM positive patients were all negative. The inter- and intra assay CV ranges were 15.6%~40.2% and 11.9%~30.6% respectively. Conclusion It is necessary to test HBV DNA for improving blood safety and it is feasible to test pooled serum samples for HBV DNA by PCR-microfluidic chips assay, because it is convenient, time-saving, sensitive, specific and the results are reproducible.