UTILIZATION OF AUTOPSY RNA FOR THE SYNTHESIS OF THE NUCLEOCAPSID ANTIGEN OF A NEWLY RECOGNIZED VIRUS-ASSOCIATED WITH HANTAVIRUS PULMONARY SYNDROME

A newly recognized hantavirus was recently found to be associated with an outbreak of acute respiratory illness in the southwestern United S tates. The disease, which has become known as hantavirus pulmonary syn drome, has an unusually high mortality (64%). Virus isolation attempts have been unsuccessful thus far, resulting in a lack of homologous an tigen for use in diagnostic assays. For this reason, a molecular appro ach was initiated to produce recombinant homologous antigen. The virus nucleocapsid (N) protein was selected, since N has been shown to be a sensitive antigenic target in other hantavirus systems. The N protein open reading frame of the virus S genome segment was synthesized from frozen autopsy tissue by polymerase chain reaction amplification, fol lowed by cloning and expression in Hela cells (vaccinia-T7 RNA polymer ase system) and Escherichia coli. N protein-expressing Hela cells serv ed as excellent antigens for an improved indirect immunofluorescence a ssay. Use of the E.coli-expressed N protein in an enzyme-linked immuno sorbent assay improved the sensitivity and specificity when compared w ith heterologous antigens used previously. Preliminary analysis also i ndicates that the higher sensitivity could result in earlier detection of infected persons. These data demonstrate that even in the absence of a virus isolate, the necessary homologous antigen can be produced a nd can serve to improve the detection and diagnostic capabilities need ed to combat this newly recognized fatal respiratory illness in the Un ited States.