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Biomedical subjects

R A Bolin

Publications and source records attributed to R A Bolin.

14 recordsLinked to original sources
PubMed1992

The Murray Valley encephalitis virus prM protein confers acid resistance to virus particles and alters the expression of epitopes within the R2 domain of E glycoprotein.

To study the role of the precursor to the membrane protein (prM) in flavivirus maturation, we inhibited the proteolytic processing of the Murray Valley encephalitis (MVE) virus prM to membrane protein in infected cells by adding the acidotropic agent ammonium chloride late in the virus replication cycle. Viruses purified from supernatants of ammonium chloride-treated cells contained prM protein and were unable to fuse C6/36 mosquito cells from without. When ammonium chloride was removed from the cells, both the processing of prM and the fusion activity of the purified viruses were partially restored. By using monoclonal antibodies (MAbs) specific for the envelope (E) glycoprotein of MVE virus, we found that at least three epitopes were less accessible to their corresponding antibodies in the prM-containing MVE virus particles. Amino-terminal sequencing of proteolytic fragments of the E protein which were reactive with sequence-specific peptide antisera or MAb enabled us to estimate the site of the E protein interacting with the prM to be within amino acids 200 to 327. Since prM-containing viruses were up to 400-fold more resistant to a low pH environment, we conclude that the E-prM interaction might be necessary to protect the E protein from irreversible conformational changes caused by maturation into the acidic vesicles of the exocytic pathway.

Acids↗
PubMed1991

Synthetic peptides of the E2 glycoprotein of Venezuelan equine encephalomyelitis virus. II. Antibody to the amino terminus protects animals by limiting viral replication.

A peptide composed of the amino-terminal 25 amino acids of the E2 glycoprotein of the virulent Trinidad donkey (TRD) strain of Venezuelan equine encephalomyelitis virus was found to protect peptide-immunized mice from lethal TRD virus challenge (Hunt et al., 1990). Viral growth in peptide-immunized animals was found to be limited in comparison to that in nonimmunized controls. Although both treated and control groups of mice responded to virus challenge by producing neutralizing antibody, only immunized mice with preexisting antipeptide antibody survived. Polyclonal antipeptide sera as well as a monoclonal antipeptide antibody were able to passively protect naive mice from TRD virus challenge, despite the fact that these antibodies were nonneutralizing. Passive transfer of antipeptide antibody to immunosuppressed recipients was not protective, thus indicating that survival of TRD virus challenge required an in situ immune response as well as preexisting antipeptide antibody. Binding studies of both polyclonal and monoclonal antipeptide antibodies indicated that they recognize only epitopes present on virus-infected cells or denatured virus.

Animals↗
PubMed1991

Use of a new synthetic-peptide-derived monoclonal antibody to differentiate between vaccine and wild-type Venezuelan equine encephalomyelitis viruses.

We have prepared a murine monoclonal antibody (MAb) capable of distinguishing between wild-type Venezuelan equine encephalomyelitis (VEE) virus and the TC-83 vaccine derivative. This MAb, 1A2B-10, was derived from immunization with a synthetic peptide corresponding to the first 19 amino acids of the E2 glycoprotein of Trinidad donkey VEE virus. The MAb reacts with prototype viruses from all naturally occurring VEE subtypes except subtype 6 in an enzyme-linked immunosorbent assay. It does not react with TC-83 virus or members of the western and eastern equine encephalitis virus complex or with Semliki Forest virus. This antibody will also differentiate between TC-83 and Trinidad donkey VEE virus in indirect immunofluorescence assays with virus-infected Vero cells.

Amino Acid Sequence↗
PubMed1990

Antibodies to dengue 2 virus E-glycoprotein synthetic peptides identify antigenic conformation.

Eighteen synthetic peptides have now been prepared from dengue (DEN) 2 virus, representing 80% of the extra-membranal domain of the DEN envelope (E)-glycoprotein. These peptides were selected based upon our previous results with Murray Valley encephalitis (MVE) virus, computer analysis of the DEN virus sequence, and the predicted E-glycoprotein secondary structure. Six of these peptides were "structural" peptides derived from either nonlinear amino acid sequences or predicted disulfide bridge-stabilized loop structures. As with MVE virus, some peptides could be recognized by antivirus hyperimmune ascitic fluids, but monoclonal antiflavivirus antibodies failed to react with any peptide. Immunogenicity of these peptides was tested in BALB/c or NIH-Swiss mice. Fourteen of these peptides elicited antipeptide antibody in one or both mouse strains. Eleven of these antipeptides reacted with virus. Peptides 35 (amino acids 35-55) and 17 (amino acids 352-368) elicited virus-neutralizing antibody. Four antipeptides were more efficient at binding to pH 5.0-treated virus. These antipeptides precisely defined a flavivirus group common sequence (amino acids 98-110) that has biochemical characteristics similar to previously defined viral fusion sequences.

Amino Acid Sequence↗
PubMed1990

Identification of monoclonal antibodies capable of differentiating antigenic varieties of eastern equine encephalitis viruses.

We have isolated and characterized 3 monoclonal antibody (Mab) reagents useful in the serological identification of varieties of eastern equine encephalitis (EEE) viruses. These antibodies were specific for the E1 glycoprotein of their homologous viruses. One Mab, 1B5C-3, reacted specifically with all North American (NA) EEE viruses isolated over a 50 year period. This antigenic stability of NA isolates was genetically confirmed by oligonucleotide fingerprinting. Evolutionary stability is a unique feature among alphaviruses. The Mab, 1C1J-4 reacted specifically with 1 South American isolate of EEE virus. A third Mab, 1B1C-4, was EEE virus complex reactive. While none of these antibodies had virus neutralizing activity, the identified reactivities could be demonstrated in the more rapid serological tests of enzyme-linked immunosorbent assay and indirect immunofluorescence.

Alphavirus↗
PubMed1989

Surveillance of St. Louis encephalitis virus vectors in Grand Junction, Colorado, in 1987.

Grand Junction, Colorado, was the site of a St. Louis encephalitis (SLE) outbreak in 1985. Epidemiologic and ecologic investigations in 1985 and 1986 suggested that Culex tarsalis may not have been the exclusive vector in the outbreak and that Cx. pipiens may have contributed to transmission as an accessory vector. A limited field study in 1987 generally confirmed observations from 1986 that Cx. pipiens was more abundant than Cx. tarsalis in late summer when SLE virus transmission normally occurs. In both years, infection rates in Cx. tarsalis were higher than in Cx. pipiens, but in 1987 the only SLE virus isolate from Cx. pipiens was obtained early in the season. Truck trap collections showed that Cx. pipiens was the principal vector species collected, comprising 86% of the total. Light trap collections underestimated the population of Cx. pipiens; gravid trap collections gave a closer approximation of the relative proportions of Cx. pipiens and Cx. tarsalis in the vector mosquito population after midsummer.

Animals↗
PubMed1988

Stability of St. Louis encephalitis viral antigen detected by enzyme immunoassay in infected mosquitoes.

The use of enzyme immunoassay to detect St. Louis encephalitis (SLE) viral antigen in vector mosquitoes enhances the effectiveness of surveillance because infected mosquitoes can be identified more rapidly than with conventional virus isolation systems and because it is a simple and accessible procedure. Infectivity among mosquitoes experimentally infected with SLE virus was lost within 24 h after the mosquitoes were stored at 27 degrees C and 80% relative humidity; however, viral antigen remained stable under these conditions and could be detected by enzyme immunoassay 2 weeks later. Desiccation further extended the period during which antigen could be detected to 6 weeks. Absorbances were higher in infected mosquitoes stored at 27 degrees C than in mosquitoes frozen continuously. Absorbances in infected mosquitoes also increased after repeated freezing and thawing and sonication. Both phenomena may be related to the release of antigen from decaying or disrupted cells. The relative stability of SLE viral antigen at ambient temperatures lends flexibility to schemes which use direct antigen detection to identify vectors. Surveillance systems can be designed without regard to collecting living mosquitoes, and a cold chain in unnecessary to preserve specimens, thus reducing the cost of surveillance and expanding the geographic areas to which it is accessible.

Animals↗
PubMed1987

Epidemiologic aspects of a St. Louis encephalitis outbreak in Mesa County, Colorado.

An epidemic of St. Louis encephalitis in 1985 in Mesa County, Colorado, led to 17 cases, including one fatality. Risk was associated with advanced age and residence in Grand Junction, the county's principal city. A trend was observed toward higher risk in females. However, increased risk in females was not associated with higher infection rates (increased exposure). Capture enzyme immunoassays detected specific immunoglobulin M (IgM) and immunoglobulin A after infection. A serosurvey of Grand Junction residents disclosed an infection rate of 4.0%, indicating that 1,123 epidemic St. Louis encephalitis infections may have occurred in the city. Evidence of previous St. Louis encephalitis virus infection was found in 11.2% of survey respondents who had neutralizing antibody to the virus without specific IgM. The prevalence of St. Louis encephalitis virus antibody was similar to rates observed in serosurveys undertaken 30 years earlier, indicating that the level of endemic St. Louis encephalitis transmission in the city had not changed appreciably in that interval.

Adolescent↗
PubMed1987

Detection of St. Louis encephalitis virus antigen in mosquitoes by capture enzyme immunoassay.

Surveillance methods that measure St. Louis encephalitis (SLE) virus activity in nature may provide forewarning of its epidemic occurrence in humans. An antigen capture enzyme immunoassay was developed to detect SLE virus in infected mosquitoes. The assay detected purified SLE viral antigen at a concentration of 62 pg/0.1 ml when antigen was incubated overnight; 250 pg/0.1 ml was detected in a single-day assay (antigen incubated for 3 h). The assay detected 67.9 and 70.8% of laboratory-prepared pools of infected mosquitoes after 3 h and overnight incubation, respectively. The sensitivity of the procedure was 90.5% in identifying pools with infectious titers greater than dex 3.0. The specificity of the assay was controlled by retesting positive pools preincubated with SLE virus and normal antibodies, which led to a diminution of signal in the pools containing viral antigen. The procedure was suitably specific in discriminating between SLE and related flaviviruses, detecting only high infectious doses of heterologous antigens.

Animals↗
PubMed1981

The ecology of Colorado tick fever in Rocky Mountain National Park in 1974. II. Infection in small mammals.

Field studies of Colorado tick fever (CTF) in small mammals in Rocky Mountain National Park (RMNP) in 1974 established that Eutamias minimus and Spermophilus lateralis were the most important hosts for CTF virus and were the source of virus for immature stages of the tick vector, Dermacentor andersoni. Other species (Peromyscus maniculatus, Spermophilus richardsonii, Eutamias umbrinus) are secondary hosts. The intensity of viral activity in rodents varied greatly from locality to locality. Highest rodent infection rates were found to occur in the Moraine Park area of RMNP. Lowest infection rates occurred above 3,290 meters in altitude at Rainbow Curve and on the tundra. The prevalence of infection in rodents was constant from April--July (5--6% of animals captured were viremic) and then declined to 1.7--2.5% in August and September coincident with a decline in nymphal tick ectoparasitism. Many animals were captured which were simultaneously viremic and antibody-positive. Under field conditions, neutralizing antibody seroconversion does not always occur.

Animals↗
PubMed1981

Transovarial transmission of St. Louis encephalitis virus by Culex pipiens complex mosquitoes.

Experiments were conducted to determine whether transovarial transmission (TOT) of St. Louis encephalitis (SLE) virus occurs in Culex pipiens complex mosquitoes, the principal vectors of SLE virus in the central-eastern United States. In 1978, field-collected mosquitoes from Memphis, Tennessee, and McLeansboro, Illinois, were used; during 1979, colonized mosquitoes from Chicago, Illinois, and Memphis, Tennessee, were used. Mosquitoes were infected by feeding on viremic chicks inoculated with an SLE virus strain isolated from Cx. pipiens complex mosquitoes collected from Memphis, Tennessee, in 1976. During the 1979 experiments, progeny larval and adult mosquitoes were held at two temperatures, 18 and 25 degrees C. Progeny were tested for virus by plaque assay in duck embryo cell cultures and by inoculation of Aedes albopictus C6/36 cells and examination by immunofluorescence. In 1978, most of the progeny tested were from the first ovarian cycle, and a single occurrence of TOT was documented. In 1979, a single TOT occurred from 46,856 first ovarian cycle progeny, whereas 7 of 9,234 progeny of the second ovarian cycle were infected. The rate of TOT was higher for progeny of Memphis than Chicago mosquitoes, and for mosquitoes held at 18 degrees C than at 25 degrees C; however, these differences were not statistically significant. Four positive pools were females, and three were fed on chicks for transmission attempts. The positive Chicago mosquito pool failed to transmit, but both Memphis pools successfully transmitted virus. The overall rates of TOT of SLE virus in progeny of the first and second ovarian cycle were, respectively, 1/45, 151 and 1/1,460. The significance of these results as they relate to the natural history of SLE virus is discussed.

Animals↗