Biochemical and immunologic comparison of virus-like particles for a rotavirus subunit vaccine.

A parenterally administered rotavirus vaccine composed of virus-like particles (VLPs) is being evaluated for human use. VLPs composed of bovine VP6 and simian VP7 (SA11, G3) proteins (6/7-VLPs) or of bovine VP2, bovine VP6, and simian VP7 (SA11, G3) proteins (2/6/7-VLPs) were synthesized and purified from Sf9 insect cells co-infected with recombinant baculoviruses. 6/7- and 2/6/7-VLP administered parenterally (i.

Synthesis of rotavirus-like particles in insect cells: comparative and quantitative analysis.

When the three major structural proteins, VP2, VP6, and VP7, of rotavirus are co-expressed in insect cells infected with recombinant baculoviruses, they self-assemble into triple-layered virus-like particles (VLPs) that are similar in morphology to native rotavirus. In order to establish the most favorable conditions for the synthesis of rotavirus VLPs, we have compared the kinetics of 2/6/7-VLP synthesis in two different insect cell lines: High Five cells propagated in Excell 405 medium and Spodoptera frugiperda 9 cells in Excell 400 medium. The majority of VLPs produced in both cell lines were released into the culture medium, and these released VLPs were predominantly triple-layered and were found to be stable for the period of six or seven days examined.

Heterotypic protection from rotavirus infection in mice vaccinated with virus-like particles.

Virus-like particles (VLPs) composed of rotavirus VP2, VP6, and VP7 of G1 or G3 serotype specificity were produced in insect cells coinfected with recombinant baculoviruses expressing single rotavirus genes. The VLPs were purified and subsequently evaluated for immunogenicity and protection in the adult mouse model of rotavirus infection. Mice were vaccinated twice intramuscularly with G1 VLPs formulated with Quillaja saponaria (QS-21) or adsorbed to aluminium hydroxide (AlOH), or with G1 VLPs alone.

Virus-like particles as a rotavirus subunit vaccine.

Rotavirus subunit vaccines are being evaluated for use in humans. The virus-like particles (VLPs) for these vaccines are produced in insect cells coinfected with combinations of baculovirus recombinants expressing bovine RIF VP2 and simian SA11, VP4, VP6, or VP7 rotavirus proteins. VLPs were administered parenterally to mice and rabbits, and the immunogenicity and protective efficacy of the vaccines were evaluated.

Sequence diversity of small, round-structured viruses in the Norwalk virus group.

We have determined the nucleotide sequences of a highly conserved region of the RNA-dependent RNA polymerase of the prototype Snow Mountain agent (SMA) and of four other small, round-structured viruses (antigenically Norwalk virus [NV]-like or SMA-like) following reverse transcription-PCR amplification of viral RNA obtained from human stools. The stool samples were either from volunteers administered SMA or from sporadic cases and outbreaks of gastroenteritis that occurred in Japan and the United Kingdom between 1984 and 1992. The GLPSG and YGDD RNA polymerase motifs were in the proper locations in the sequences of the five SRSVs, but each sequence was distinct from the 8FIIa prototype NV sequence and from each other.

Norwalk virus infection of volunteers: new insights based on improved assays.

Norwalk virus infection is a common cause of gastroenteritis in humans. The clinical features and virologic and immunologic responses following oral administration of Norwalk virus to 50 volunteers were monitored. New ELISAs using recombinant virus particles as the antigen source were used to assess the pattern of virus shedding and the specific immune responses.

Subclass-specific serum antibody responses to recombinant Norwalk virus capsid antigen (rNV) in adults infected with Norwalk, Snow Mountain, or Hawaii virus.

Subclass-specific antibody responses to the Norwalk virus capsid protein in adults challenged with Norwalk, Snow Mountain, or Hawaii virus were evaluated by solid-phase enzyme immunoassay using recombinant Norwalk virus capsid antigen (rNV). Fourfold or greater serum immunoglobulin G (IgG) antibody responses to rNV were detected in 15 of 20 volunteers challenged with Norwalk virus, and serum IgA and IgM antibody responses to rNV were seen in almost all subjects who had rNV IgG responses. Serum rNV IgG antibody responses also were detected in 6 of 15 volunteers challenged with Snow Mountain virus and 2 of 12 volunteers challenged with the Hawaii virus.

Field trial of rhesus rotavirus or human-rhesus rotavirus reassortant vaccine of VP7 serotype 3 or 1 specificity in infants. The Elmwood, Panorama, and Westfall Pediatric Groups.

Orally administered live rhesus monkey rotavirus vaccine (RRV, VP7 serotype 3) and human-rhesus reassortant rotavirus vaccine (DxRRV, VP7 serotype 1) were evaluated in a placebo-controlled field trial of 223 infants 2-4 months old. Both vaccines were mildly reactogenic but were generally well tolerated in the 10 days after vaccination. RRV and DxRRV were immunogenic, inducing serum antibody responses in 78% and 71% of the vaccines, respectively.

Antigenic relatedness among the Norwalk-like agents by serum antibody rises.

The Norwalk, Snow Mountain (SMA), and Hawaii agents are etiologically associated with separate outbreaks of acute viral gastroenteritis. Previous cross-challenge of volunteers, immune electron microscopy, and/or enzyme-immunoassay analysis suggested that these agents are antigenically distinct. We examined paired sera from human volunteers challenged with these agents for the presence of homologous and heterologous serum antibody titer rises to the agents.

Comparison of three enzyme immunoassays to tissue culture for the diagnosis of rotavirus gastroenteritis in infants and young children.

Three enzyme immunoassays (EIAs), Rotazyme II, IDL, and Pathfinder, were evaluated for rotavirus detection in stool and rectal swab specimens from children with symptomatic gastroenteritis and compared with virus isolation in primary African green monkey kidney cells. Of 125 specimens tested, 49 were rotavirus positive by tissue culture isolation; of these 49, 40 were positive by Rotazyme II, 43 were positive by IDL, and 46 were positive by Pathfinder EIAs. As compared with tissue culture isolation, the Rotazyme II, IDL, and Pathfinder EIAs had sensitivities of 82, 88, and 94%, specificities of 90, 99, and 95%, and overall agreements of 86, 94, and 94%, respectively.

Foodborne Snow Mountain agent gastroenteritis with secondary person-to-person spread in a retirement community.

A variety of small round-structured viruses are being recognized with increasing frequency as a cause of gastroenteritis in the community, but have rarely been reported to cause outbreaks in hospitals or extended-care facilities. From March 20 through April 15, 1988, an outbreak of gastroenteritis occurred in a retirement facility in the San Francisco Bay area. Illness was characterized by diarrhea, nausea, and vomiting; two residents died.

Viral gastroenteritis aboard a cruise ship.

A 32-nm small round structured virus (SRSV), possibly related to the Snow Mountain agent (SMA), was implicated as the cause of recurrent outbreaks of gastroenteritis on a cruise ship. There was no identifiable relation to food or water consumption, but the risk of gastroenteritis among passengers who had shared toilet facilities was twice that of those who had a private bathroom and the rate of illness was related to the number of passengers sharing a communal restroom (ie, with one or more toilets): contaminated bathrooms may be an important vehicle for person-to-person spread of this enteric agent. In each cabin, index patients who had vomited in their cabins were more likely to have had cabinmates who subsequently became ill than were index patients who had not vomited.

Development of an enzyme immunoassay for the Hawaii agent of viral gastroenteritis.

The Hawaii agent is a Norwalk-like virus of acute gastroenteritis in humans which is antigenically distinct from the prototype Norwalk agent. We established a solid phase sandwich type microtiter enzyme immunoassay (EIA) for Hawaii antigen employing sera and stools from experimentally challenged volunteers as reagents. This assay detected the Hawaii agent in stools from 3 of 8 volunteers who were ill after oral challenge with the Hawaii agent, including one specimen which was positive to a dilution of 1/320.

Field trial of rhesus rotavirus vaccine in infants.

Orally administered rhesus rotavirus vaccine (RRV) was evaluated in a placebo-controlled study in 176 infants (ages 2 to 4 months). Eighty-eight infants received a dose of 10(4) plaque-forming units of the vaccine, and 88 received the placebo. RRV was well-tolerated but mildly reactogenic in the 10 days after vaccination.

Production of a monoclonal antibody against the Snow Mountain agent of gastroenteritis by in vitro immunization of murine spleen cells.

The Snow Mountain agent (SMA) is a 27- to 32-nm noncultivatable virus that causes acute gastroenteritis in humans. SMA is morphologically similar to but immunologically distinct from the Norwalk agent. SMA has been partially purified from the stool of experimentally infected volunteers and contains a single structural protein of Mr 62,000 as well as one or more non-virion-associated soluble proteins.

Snow Mountain agent gastroenteritis from clams.

A 1983 investigation of two clambake-related gastroenteritis outbreaks in Rochester, New York, showed that 84 (43%) of 196 persons interviewed had an acute illness characterized by watery diarrhea, vomiting, and abdominal cramps. None of the ill persons were hospitalized or had complications. Illness was associated with eating raw (p = 0.

Foodborne Snow Mountain agent gastroenteritis in a school cafeteria.

In 1984, an outbreak of gastroenteritis occurred at a school with 1,860 students in Brooklyn, NY. In a single-stage cluster sample of 375 students, 129 (34%) had illnesses that met our case definition of vomiting or diarrhea. The mean incubation period was 26 hours, and the mean illness duration was 24 hours.

Novel agents of viral enteritis in humans.

Considerable information has recently emerged regarding certain "groups" of novel viruses associated with gastroenteritis in humans. The viruses reviewed here are 20-35 nm in diameter and can be detected in the stools of acutely ill individuals with gastroenteritis. These viruses can be conveniently divided into four "groups": Norwalk-like agents, caliciviruses, astroviruses, and other small round viruses.

Enzyme-linked immunosorbent assays for Snow Mountain and Norwalk agents of viral gastroenteritis.

Enzyme-linked immunosorbent assays (ELISAs) for antigen detection and blocking ELISAs for serum antibody rises were developed for the Snow Mountain and Norwalk agents of viral gastroenteritis. The ELISAs were as sensitive as the existing radioimmunoassays and were specific for the Snow Mountain or Norwalk agent. The blocking ELISAs detected the same number of significant rises in antibodies to these agents as did the existing blocking radioimmunoassays.

Characterization of the Snow Mountain agent of viral gastroenteritis.

Snow Mountain agent (SMA) is a 27- to 32-nm virus which is the etiologic agent of outbreaks of acute gastroenteritis in Colorado and Vermont. SMA is morphologically similar to but antigenically distinct from the Norwalk and Hawaii agents of viral gastroenteritis but, like those agents, has not been cultivated in vitro. We purified and characterized SMA directly from human stool specimens containing the virus.

Radioimmunoassay for detection of the Snow Mountain Agent of viral gastroenteritis.

The Snow Mountain Agent (SMA) is a Norwalk-like viral agent of acute gastroenteritis that has been detected only by immune electron microscopy (IEM). We established a solid phase microtiter radioimmunoassay (RIA) for SMA antigen employing pre-and post-challenge sera from volunteer studies as capture antibodies. SMA was detected in 12 of 67 stool samples from volunteers who were ill after oral challenge with SMA.