Viral metagenomics in blood donations with post-donation illness reports from Brazil.

BACKGROUND: Post-donation illness can be described as appearance of clinical symptoms in blood donors after donation. The consequent call back of the donor to report these symptoms to the blood collection institution is considered a post-donation illness report (PDIR). The most suitable way to examine whether PDIR is related to infection is to apply next-generation sequencing (NGS) and viral metagenomics. Investigation into a PDIR can reveal its importance for transfusion safety and help elaborate strategies for donor education in order to prevent the transfusion transmission of infections which are not routinely tested by the blood collection services. MATERIALS AND METHODS: We applied NGS and viral metagenomics on blood donations which were deferred due to a PDIR. Thirty-three PDIR donations obtained in the Blood Center of Ribeirão Preto, Southeast Brazil, were evaluated. Sequencing was performed using Illumina NextSeq 550 (Illumina Inc, San Diego, CA, USA) equipment and the reads obtained for each sample were analysed by specific bioinformatic pipeline for the classification and discovery of emerging viruses. The identified viral agents by metagenomics were directly confirmed by molecular methods. RESULTS: In all PDIR donations, we found abundant reads of commensal viruses belonging to the Anelloviridae family as well as human pegivirus-1. However, we were also able to identify blood donations positive for clinically important viruses like dengue serotype-2 (DENV-2) of the Asian-American genotype and parvovirus B19 (B19V). Both viruses were also confirmed by real-time polymerase chain reaction, detecting DENV-2 RNA in a significant number of cases (7 samples, 21.2%), compared to B19V which was confirmed in 1 case (3.0%). DISCUSSION: Our study applies for the first time viral metagenomics to evaluate the significance of PDIRs. We confirm the crucial importance of the donor providing a timely PDIR for the prevention of transfusion transmission of viral infections which are not routinely tested in the blood banks worldwide.

A DNA vaccine against tuberculosis based on the 65 kDa heat-shock protein differentially activates human macrophages and dendritic cells.

BACKGROUND: A number of reports have demonstrated that rodents immunized with DNA vaccines can produce antibodies and cellular immune responses presenting a long-lasting protective immunity. These findings have attracted considerable interest in the field of DNA vaccination. We have previously described the prophylactic and therapeutic effects of a DNA vaccine encoding the Mycobacterium leprae 65 kDa heat shock protein (DNA-HSP65) in a murine model of tuberculosis. As DNA vaccines are often less effective in humans, we aimed to find out how the DNA-HSP65 stimulates human immune responses. METHODS: To address this question, we analysed the activation of both human macrophages and dendritic cells (DCs) cultured with DNA-HSP65. Then, these cells stimulated with the DNA vaccine were evaluated regarding the expression of surface markers, cytokine production and microbicidal activity. RESULTS: It was observed that DCs and macrophages presented different ability to uptake DNA vaccine. Under DNA stimulation, macrophages, characterized as CD11b+/CD86+/HLA-DR+, produced high levels of TNF-alpha, IL-6 (pro-inflammatory cytokines), and IL-10 (anti-inflammatory cytokine). Besides, they also presented a microbicidal activity higher than that observed in DCs after infection with M. tuberculosis. On the other hand, DCs, characterized as CD11c+/CD86+/CD123-/BDCA-4+/IFN-alpha-, produced high levels of IL-12 and low levels of TNF-alpha, IL-6 and IL-10. Finally, the DNA-HSP65 vaccine was able to induce proliferation of peripheral blood lymphocytes. CONCLUSION: Our data suggest that the immune response is differently activated by the DNA-HSP65 vaccine in humans. These findings provide important clues to the design of new strategies for using DNA vaccines in human immunotherapy.