Development of quantitative multiplex RT-qPCR one step assay for detection of hepatitis delta virus.

Hepatitis Delta is a disease caused by exposure to hepatitis B (HBV) and hepatitis D (HDV) viruses, usually with a more severe clinical outcome when compared to an HBV monoinfection. To date, the real prevalence of HDV infection is underestimated and detection methods are poorly available, especially in more endemic regions. Therefore, a one-step RT-qPCR method for quantification of HDV-RNA was developed. Biological samples were selected between 2017 and 2023 from patients at the Ambulatório Especializado em Hepatites Virais of the Centro de Pesquisa em Medicina Tropical de Rondônia and Serviço de Assistência Especializada and underwent the test developed by this study and a second quantitative RT-qPCR assay. The slope of the initial quantitative assay was - 3.321 with an efficiency of 100.04% and amplification factor equal to 2. Analysis of the repeatability data revealed a Limit of Quantification of 5 copies/reaction and Limit of Detection (95%) of 2.83 copies per reaction. In the diagnostic sensitivity tests, there was an accuracy of 97.37% when compared to the reference test. This assay proved to be highly efficient and reproducible, making it a valuable tool to monitor hepatitis Delta patients and assess the risk of disease progression, as well as the effectiveness of treatment.

A Multiplex Nanopore Sequencing Approach for the Detection of Multiple Arboviral Species.

The emergence and continued geographic expansion of arboviruses and the growing number of infected people have highlighted the need to develop and improve multiplex methods for rapid and specific detection of pathogens. Sequencing technologies are promising tools that can help in the laboratory diagnosis of conditions that share common symptoms, such as pathologies caused by emerging arboviruses. In this study, we integrated nanopore sequencing and the advantages of reverse transcription polymerase chain reaction (RT-PCR) to develop a multiplex RT-PCR protocol for the detection of Chikungunya virus (CHIKV) and several orthoflaviviruses (such as dengue (Orthoflavivirus dengue), Zika (Orthoflavivirus zikaense), yellow fever (Orthoflavivirus flavi), and West Nile (Orthoflavivirus nilense) viruses) in a single reaction, which provides data for sequence-based differentiation of arbovirus lineages.

Levels of hepatitis B antibody titers are affected by age and doses gap time in children from a high endemic area of the western Amazon.

BACKGROUND: Despite completion of the vaccine schedule for hepatitis B virus (HBV), children may display levels of HBV surface antibodies (anti-HBs) that are considered inadequate for sufficient protection (<10 IU/L). AIMS: Our aim was to investigate if age and gap time between HBV vaccine doses may negatively affect the levels of anti-HBs in children, and if these relationships are modified by sex. METHODS: In a high-endemic HBV region of the western Brazilian Amazon we enrolled children who had completed the HBV vaccine schedule. All children underwent analysis of anti-HBs and a clinical examination. RESULTS: We included 522 children (mean age 4.3 ± 0.8 years; 50% male). Median anti-HBs was 28.4 [interquartile range (IQR) 5.4 to 128.6] IU/L and 32% had anti-HBs <10 IU/L. The median gap time from last to preceding dose was 2.4 [IQR 2.1 to 3.3] months. Levels of anti-HBs decreased with higher age (-42% per year increase [95%CI -56% to -24%], p<0.001), but not with longer gap time (+23% per month increase [95%CI -16% to +62%], p = 0.249). After adjusting for relevant confounders, gap time became significant (p = 0.032) and age remained a significant predictor of anti-HBs (p<0.001). CONCLUSION: One third of assessed children displayed anti-HBs <10 IU/L. Levels of anti-HBs decreased with higher age and increased with longer gap time between the last two doses.