Functional roles of the V3 hypervariable region of HIV-1 gp160 in the processing of gp160 and in the formation of syncytia in CD4+ cells.

To study the roles of the V3 hypervariable region (amino acid residues 301-336) of the HIV-1 envelope glycoprotein (gp160) during infection, we constructed recombinant vaccinia viruses that expressed either wild-type gp160 (v-env10) or mutant gp160 in which the V3 region was deleted (v-dl29.1 and v-dl29.2). In v-dl29.1 the V3 loop, formed by disulfide bonding between cysteine residues 301 and 336, was deleted from cys301 to cys336 (inclusive) and replaced by one serine residue. In v-dl29.2 the V3 loop was deleted from arg303 to ala334 and replaced by three residues: gly-ala-gly. Cells infected with all three recombinant vaccinia viruses expressed gp160 on the cell surface, but v-dl29.1-derived gp160 was not cleaved into gp120 and gp41 and did not bind the CD4 glycoprotein. In contrast, gp160 produced by recombinant v-dl29.2 was cleaved normally, and the mutant gp120 produced was secreted and retained binding activity to CD4+ cells. However, both mutants failed to induce syncytia in HeLa CD4+ cells. Thus a disulfide loop at the V3 portion of gp160 is required for cleavage into gp120 and gp41, presumably because the loop is required for proper tertiary structure. The sequence within the V3 loop, however, is not required for cleavage and secretion of gp160, or for binding to CD4+, but this region is essential for gp120-mediated syncytia formation.

Rapid diagnosis of LaCrosse encephalitis: detection of specific immunoglobulin M in cerebrospinal fluid.

An immunoglobulin M (IgM) antibody capture enzyme immunoassay (MAC-EIA) was developed for the rapid and early diagnosis of LaCrosse (LAC) virus infections. The MAC-EIA was a sensitive and specific technique for the detection of IgM antibodies to LAC virus in cerebrospinal fluid specimens and in acute-phase serum specimens. In a retrospective study, cerebrospinal fluid and acute-phase serum paired samples from 108 patients were tested by the MAC-EIA and by an IgM immunofluorescence assay. The results were compared with the original diagnosis, which was made by using a variety of classical serological tests including serum neutralization, hemagglutination inhibition, and complement fixation. Thirty patients were confirmed as having LAC virus infections; of these, 30 (100%) were diagnosed as positive by serum MAC-EIA, and 27 (90%) were positive by cerebrospinal fluid MAC-EIA. The MAC-EIA was more sensitive than the IgM immunofluorescence assay. Two patients who were not previously confirmed as positive cases were diagnosed as having LAC virus infections by the MAC-EIA. One patient who was subsequently diagnosed as having a Jamestown Canyon virus infection and two patients who were previously infected with Jamestown Canyon virus were not falsely identified as having LAC virus infections by the MAC-EIA.