Isolation of a strain of Hantaan virus from peritoneal exudate cells of a patient with hemorrhagic fever with renal syndrome.

A new strain of Hantaan virus (HTNV), GH716, was isolated from the peritoneal exudate cells (PEC) of a patient with severe hemorrhagic fever with renal syndrome (HFRS). The isolate was propagated in Vero E6 cells. At each passage the virus-infected cells were examined for HTNV with immunofluorescence technique using monoclonal antibodies (McAb) 25-1 and 84-1 against HTNV. From passage 5 on, fluorescent intensity tended to stabilize at (+++) and infectivity titer reached 10(5) TCID50/ml. Using 14 McAb to 76-118 and BI strains of HTNV, we found that strain GH716 was antigenically similar to strain 76-118 (Apodemus type) and different from strain R22 (Rattus type), suggesting that GH716 may fall into the Apodemus type of HTNV. This is the first isolation of an HTNV from human PEC collected on the tenth day of illness. The successful isolation of strain GH716 may provide an alternative source of obtaining HTNV during the later stages of HFRS.

Hemorrhagic fever with renal syndrome. Separation of human peripheral blood T and B cells and detection of viral antigen.

In this study, total blood lymphocytes were prepared from patients with hemorrhagic fever with renal syndrome (HFRS) by a density gradient on Ficoll-Hypaque. T and B cells were then purified by passing the total lymphocytes over a nylon wool column. The purities of total lymphocytes, T cells and B cells were 97.8 +/- 2.3%, 91.6 +/- 4.5% and 74.2 +/- 12.1%, respectively. Also, after modification of cell fixation and smear drying, the number of cells were increased and the time needed for slide preparation was shortened. Detection of viral antigen by immunofluorescence assay using monoclonal antibodies to Hantavirus (HTNV) showed that the total lymphocytes. T cells and B cells were infected by HTNV during the early stages of HFRS and no specific fluorescence was seen in the cells from the late diuretic phase to convalescent phase. The results suggest that virus replication in blood lymphocytes may partly contribute in the early stages to the impairment of cell immune response and in vivo spread of HTNV to its target sites.

Hemorrhagic fever with renal syndrome: separation of human peripheral blood T and B cells and detection of viral antigen.

Total lymphocytes in peripheral blood were prepared from patients with hemorrhagic fever with renal syndrome by a density gradient on Ficoll-Hypaque, and then T and B cells purified by passing the lymphocytes over a nylon column with modifications. Also, after modification of cell fixation and use of a spinner for smear drying, the number of cells were increased and the duration for slide preparation was shortened, thus resulting in quality slides. Detection of viral antigen by immunofluorescence assay using monoclonal antibodies to Hantavirus (HTNV) showed that the total lymphocytes, T cells and B cells were infected with HTNV during the early stages of the illness and no specific fluorescence was seen in the cells from the late diuretic phase to convalescent phase.