Molecular and viral research on dengue hemorrhagic fever

为了阐明与登革热出血热发生相关的各种病毒和宿主因素，我们开展了一项为期三年的研究项目。我们在孟加拉国、菲律宾和越南登革热流行期间分离了一些登革热病毒株，并测定了病毒株的核苷酸序列。并与常规数据库中得到的序列进行了比较。我们目睹了21世纪初菲律宾登革热病毒2型的基因型转变，从本地菌株到世界性菌株。在孟加拉国和越南，登革热病毒在当地进化，并在这些国家引起周期性流行病。在与登革出血热和登革热病毒的比较中，我们没有发现两组之间有任何显著差异。在流式细胞术分析中，我们清楚地证明了登革热病毒可以感染外周B细胞和NK细胞，这表明登革热病毒在人体内的目标并不总是在特定的细胞群上，病毒可以根据其对每个病毒群的亲和力选择目标细胞。后来，通过使用不同的细胞系从一个病人身上分离出两种不同的病毒株，这一事实进一步加强了这一想法。一种病毒染色从蚊子细胞中分离出来，在细胞中生长良好，但不能在人B细胞系中生长。另一种病毒由人单核细胞培养细胞株K562分离得到，在B细胞株中生长良好。这两种病毒在E蛋白中具有氨基酸交替，表明这是细胞趋向性的决定因素。登革病毒种群差异很可能对确定登革病毒在人感染中的发病机制有一定影响。有必要进一步澄清这种登革热病毒种群差距的重要性，以了解确定人类登革热病毒感染严重程度的机制。

In order to elucidate various virus and host factors that are relevant to the development of dengue hemorrhagic fever, we conducted a three-year research project. We isolated a number of dengue virus strains during dengue epidemic in Bangladesh, the Philippines, and Vietnam and determined nucleotide sequences of the virus strains. The data was compared with each other as well as the sequences obtained in conventional data base. We witnessed the genotype shift of dengue virus 2 in the Philippines during the early 2000s from local strain to cosmopolitan strain. While in Bangladesh and Vietnam, dengue viruses evolved locally and caused periodic epidemics in the countries. In the comparisons made with viruses from dengue hemorrhagic fever and dengue fever, we could not detect any significant differences between the two groups. In the flow cytometry analysis, we clearly demonstrated that dengue virus can infect peripheral B cells and NK cells suggesting the target of dengue virus in humans is not always on a particular cell group but the virus can choose the target cells according to its affinity to each virus group.This idea was further strengthened later by the fact that two distinct virus strains were isolated from one patient by using different cell lines for isolation. One virus stain was isolated by mosquito cells and grow well in the cells but not human B cell line. While the other virus was isolated by K562, a culture cell line form human monocyte, grow well in B cell lines. Those two viruses possessed an amino acid alternation in E protein, suggesting a determinant of cell tropism. It is very likely that dengue virus population gap may have some effect to determine dengue virus pathogenesis in human infection. Further clarification of the importance of this dengue virus population gap is necessary to understand the mechanism to determine the severity in dengue virus infections in humans.