DETECTION AND SUBSEQUENT SEQUENCING OF PUUMALA-VIRUS FROM HUMAN SPECIMENS BY PCR

A sensitive method based on PCR was developed for the detection of Puu mala virus (PUU) in human samples. The assay was found to be specific for PUU-like strains and distinguished between these and hantaviruses of other serotypes. The detection limit was found to be 10(-5) focus-f orming units. Clinical samples were collected from patients with nephr opathia epidemica in Sweden and western Russia. Five whole blood sampl es collected from patients in Russia with the acute phase of disease w ere found to be positive by the PCR. AII samples were negative for PUU antigen when examined by enzyme-linked immunosorbent assay. Virus iso lation on Vero E6 cells from several of the acute-phase samples, inclu ding,the 5 PCR-positive samples, was not successful. The amplified sam ples were subjected to direct nucleic acid sequencing for confirmation of identity. The sequences differed from each other and were closely related to the Russian bank vole isolate CG-1820, thereby indicating t he origin of nephropathia epidemica. The PCR was used for amplificatio n and subsequent nucleotide sequencing of eight PUU-like isolates with different geographic origins. The Swedish strains were more closely r elated to the Finnish PUU prototype strain, Sotkamo, than to the Russi an isolates. Interestingly, a Belgian isolate, CG-13891, differed mark edly from all other PUU strains.