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PubMed1978

Bovine respiratory syncytial virus infection of bovine embryonic lung cultures: enhancement of infectivity with diethylaminoethyl-dextran and virus-infected cells.

The effects of incorporating diethylaminoethyl-dextran (DEAE-D) in the inoculum with bovine respiratory syncytial virus (BRSV) on the infectivity of BRSV was evaluated. A concentration of 40 microgram DEAE-D/ml provided maximal enhancement of infection as determined by the time of onset of cytopathic effect (CPE), the percentage of cells infected by the inoculum, and the amount of virus produced. When DEAE-D was used in the inoculum, the CPE appeared a day earlier, the percentage of cells infected by the inoculum, as determined by the fluorescent antibody test, was increased 11 times, and the viral titer was increased 2 times as compared to results obtained without DEAE-D. Bovine respiratory syncytial virus-infected cultures contained much cell-associated virus which could be liberated by sonication to increase the titer of virus stocks. The use of BRSV-infected cells rather than supernates from BRSV-infected cells increased the rate at which a cytopathic effect developed, although it did not substantially increase the titer of virus which was harvested. The use of DEAE-D in the inoculum and the passage of BRSV-infected cells instead of viral suspensions was found to be the quickest and most effective method of consistently obtaining BRSV with a titer of about 10(5.5) TCID50/ml.

Animals↗
PubMed1995

Bovine respiratory syncytial virus replicates minimally in bovine alveolar macrophages.

The interaction between two different bovine respiratory syncytial virus (BRSV) strains and bovine alveolar macrophages (BAMs) was studied in vitro. Bovine respiratory syncytial virus replicated minimally in BAMs and most of the virus produced remained cell-associated. Approximately 1 out of 1,000 BAMs produced infectious virus, a number that further declined during the 7 days of culture. In contrast, BAMs exposed to bovine parainfluenza 3 virus (PI3V) produced high amounts of infectious virus. The number of BAMs that contained BRSV antigen depended on the antigen load of the inoculum and not on the infectivity of the virus. Antibody mediated enhancement of infection was not detected. It is concluded that bovine alveolar macrophages exhibit a high intrinsic resistance to BRSV, but not to PI3V.

Animals↗
PubMed1992

Evaluation of three rapid enzyme immunoassays and cell culture for detection of respiratory syncytial virus.

Three rapid enzyme immunoassay techniques for the detection of respiratory syncytial virus antigen (Becton Dickinson Directigen RSV, Abbott RSV Testpack and Abbott RSV EIA) and cell culture were evaluated in a total of 250 nasal washings. The sensitivity and specificity were 62% and 76% respectively for Directigen, 64% and 86% for RSV Testpack, and 76% and 81% for RSV EIA, taking cell culture as the reference method. Agreement between cell culture and EIA techniques was 79% (70 positive and 128 negative results). All three EIA techniques gave positive results in 69 samples (52 positive and 17 negative in the cell culture). In 121 samples all three EIA techniques gave negative results (103 negative and 18 positive in the cell culture). Using the cell culture technique 46 strains other than respiratory syncytial virus were isolated.

Animals↗
PubMed2004

Comparison of clinical characteristics of influenza and respiratory syncytial virus infection in hospitalised children and adolescents.

UNLABELLED: While significant morbidity due to respiratory syncytial virus (RSV) infection in the paediatric population has been well acknowledged, little is known about the burden of influenza in primarily healthy children in Europe. In our institution, a University Children's Hospital in Switzerland, medical staff were encouraged to take nasopharyngeal specimens for multiplex polymerase chain reaction assays for influenza A and B, RSV and several other pathogens from patients hospitalised with respiratory symptoms. We took advantage of this strategy and performed a retrospective study to compare specific characteristics of influenza virus infections with those of RSV during two consecutive winter seasons. Overall, 126 patients were positive for RSV and 60 patients were positive for influenza (type A: 45; type B: 15). The median age of children with RSV, influenza A, and influenza B infection was 4 months; 2 years and 4 months; and 6 years and 2 months, respectively (P < 0.001). Fever and cough predominated in children with influenza infection whereas cough, rhinorrhoea, feeding difficulties and dyspnoea were the major symptoms in children with RSV infection. Of patients with influenza, 41% suffered from lower respiratory tract infection compared to 91% of those with RSV infection (P < 0.001). Of 60 patients hospitalised with influenza, 12 (20%) experienced febrile convulsions. None of the patients with influenza had been immunised in the respective winter season, although 27% of them had at least one underlying medical condition that would have counted as an indication for immunisation in Switzerland. CONCLUSION: influenza virus infections, like respiratory syncytial virus infections, are a major cause of hospitalisation in children with respiratory illness during the winter season. Since it is impossible to make an aetiological diagnosis on clinical grounds, it is important to apply specific diagnostic tools in children hospitalised with respiratory illness in order to better characterise the relative burden of disease caused by the respective agents.

Adolescent↗
PubMed2001

A comparison of cytokine responses in respiratory syncytial virus and influenza A infections in infants.

UNLABELLED: Respiratory syncytial virus (RSV) infection is a major cause of bronchiolitis in infants while influenza A infection usually manifests as upper respiratory tract infection. We hypothesised that the immunological responses of infants to RSV infection and influenza A infection are different. This prospective study was undertaken to compare the cytokine responses during RSV and influenza A infection. Sera and nasopharyngeal aspirates (NPA) were collected from infants with a coryzal illness with or without wheeze who were admitted to the paediatric wards during 1998. Cytokines, adhesion molecules, RANTES, IgE and eosinophil cationic protein (ECP) were measured by enzyme linked immunosorbent assay or fluorescence enzyme immunoassay. The diagnosis of RSV and influenza infections was based on direct immunofluorescence and viral culture. Of the 39 infants studied, RSV infection was confirmed in 11 patients and Influenza A in 10 patients. All RSV patients and one influenza A patient had wheeze during the infection. The serum concentrations of interleukin (IL)-4 and IL-5, regulated upon activation normal T cell expressed and secreted (RANTES) and soluble intercellular adhesion molecule 1 (sICAM-1) in infants with RSV infection were significantly higher than those with influenza A infection (all P < 0.02). The concentration of tumour necrosis factor-alpha (TNF-alpha) in NPA was significantly lower in infants with RSV infection (P < 0.01). CONCLUSION: A predominant T helper cell type 2 cytokine and related immunological response was observed in infants with respiratory syncytial virus infection whereas a predominant pro-inflammatory cytokine response was observed in infants with influenza A infection. This may explain the different clinical manifestations of the two viral infections in infants.

Cell Adhesion Molecules↗
PubMed1983

Respiratory syncytial virus glycoproteins.

The proteins of respiratory syncytial (RS) virus were analyzed by SDS-polyacrylamide gel electrophoresis. Eight virion structural proteins with molecular weights of 180,000, 89,000, 48,000, 42,000, 34,000, 28,000, 25,000, and 21,000 were identified. These proteins were given tentative designations of L (180,000), G (89,000), F1 (48,000), NP (42,000), P (34,000), M (28,000), Vp25 (25,000), and F2 (21,000). The 89,000-, 48,000-, and 21,000-dalton polypeptides were glycosylated and could be purified on lentil-lectin sepharose columns. All three glycoproteins could be immunoprecipitated from extracts of infected cells but not from uninfected cells, suggesting that they are viral specified. The host cell affected the apparent molecular weights of the largest and smallest glycosylated polypeptides possibly by differences in glycosylation. The 48,000- and 21,000-dalton glycopolypeptides were disulfide linked subunits of a 68,000-dalton glycoprotein that was seen on unreduced gels. The 68,000-dalton glycoprotein was thus similar to the fusion (F) protein of paramyxoviruses. Treatment of infected cultures with tunicamycin, a drug that blocks glycosylation, inhibited syncytial formation and resulted in over a 1000-fold reduction of extracellular infectious virus. Virions purified from tunicamycin-treated cells had reduced amounts of all three glycosylated proteins. No new forms of these proteins were conclusively identified, suggesting that unglycosylated forms of RS glycoproteins were not incorporated into virion membranes.

Chromatography, Affinity↗
PubMed1991

Nucleotide sequence analysis of the bovine respiratory syncytial virus fusion protein mRNA and expression from a recombinant vaccinia virus.

Bovine respiratory syncytial (BRS) virus is an important cause of serious respiratory illness in calves. The disease caused in calves is similar to that caused by human respiratory syncytial (HRS) virus in children. The two viruses, however, have distinct host ranges and the attachment glycoproteins, G, have no antigenic cross-reactivity. The fusion glycoproteins, F, of the HRS and BRS viruses, however, have some antigenic cross-reactivity. To further compare the BRS virus and HRS virus fusion proteins, we determined the nucleotide sequence of cDNA clones to the BRS virus F protein mRNA, deduced the amino acid sequence, and compared these sequences with the HRS virus F protein sequences. The BRS virus F mRNA was 1899 nucleotides in length and had a single major open reading frame which could code for a polypeptide of 574 amino acids with an estimated molecular weight of 63.8 kDa. Structural features predicted from the amino acid sequence included an NH2-terminal signal sequence (residues 1-26), a site for proteolytic cleavage (residues 131-136) to generate the disulfide-linked F1 and F2 subunits, and a hydrophobic transmembrane anchor sequence (residues 522-549). The nucleic acid identity between the BRS virus and the HRS virus F mRNA sequences was 71.5%. The predicted BRS virus F protein shared 80.5% overall amino acid identity with the HRS virus F protein with 89% identity in the F1 polypeptide but only 68% identity in the F2 polypeptide. The position and number of the cysteine residues in the F1 and F2 polypeptides were conserved among all F proteins. However, BRS virus F protein had only three potential N-linked carbohydrate acceptor sites in comparison to four or five for the HRS viruses. A difference in the extent of glycosylation between the BRS and HRS virus F2 polypeptides was shown to be responsible for differences observed in the electrophoretic mobility of these proteins. A cDNA containing the complete open reading frame of the BRS virus F mRNA was inserted into the thymidine kinase gene of vaccinia virus and following homologous recombination, a recombinant virus containing the BRS virus F gene was isolated. The BRS virus F protein was expressed in recombinant virus infected cells as demonstrated by immunoprecipitation and was transported to and expressed on the surface of infected cells as shown by indirect immunofluorescence.

Amino Acid Sequence↗
PubMed1988

Bovine respiratory syncytial virus-specific monoclonal antibodies.

Five hybridomas were produced which secreted monoclonal antibodies to bovine respiratory syncytial virus (BRSV). Two antibodies (8G12, 15C7) neutralized the virus and inhibited syncytia formation in vitro. These monoclonal antibodies also stained, by indirect fluorescent assay, an external envelope protein of living virus-infected cells, and recognized the 48k subunit of the viral fusion protein by Western blot analysis of bovine respiratory syncytial virus-infected cell lysates. Three other monoclonals (6A12, 14D3, 14E3) stained, by indirect fluorescent assay, acetone-fixed virus-infected cells but not living cells. Three hybridomas (6A12, 8G12, 15C7) secreted monoclonal antibody of isotype IgG1, k; two hybridomas secreted monoclonal antibody of isotype IgG2a, k. This apparently is the first report of monoclonal antibodies specific for BRSV glycoproteins.

Animals↗
PubMed1989

Detection of IgE antibodies to bovine respiratory syncytial virus.

The role of IgE antibodies against respiratory syncytial virus has attracted attention for both human and bovine disease. To detect such antibodies, we have developed an enzyme-linked immunosorbent assay (ELISA) specific for bovine respiratory syncytial virus (BRSV). Firstly, antiserum strongly positive for BRSV-specific IgE was produced by immunizing a levamisole-treated calf with BRSV. The presence and specificity of BRSV-specific IgE in this animal was confirmed with the Praunitz-Kustner (PK) technique. Potential interference in an ELISA by other BRSV-specific immunoglobulin isotypes was eliminated by preferential precipitation of serum samples with 27.5% saturated ammonium sulfate. The correlation between the PK and the assay was greater than 93% and the ELISA was found to be more specific than the PK. Indeed, in a pilot experimental infection study, the serum levels of BRSV-specific IgE were found to correlate with the symptom expression following repetitive live virus aerosolization. This may prove to be a useful rapid test to study both herd immunity and the potential pathogenic influence of IgE.

Animals↗
PubMedDate not supplied

Purification of human respiratory syncytial virus: superiority of sucrose gradient over percoll, renografin, and metrizamide gradients.

A method was devised for producing and purifying human respiratory syncytial virus (HRSV) preparations with high titers. Previous attempts to obtain substantial amounts of purified human respiratory syncytial virus have been unsuccessful due to the extreme lability of this virus, its close association with the host cell membrane, and its tendency to aggregate during concentration procedures. We describe a comparative study of various purification media as well as a novel approach for obtaining high titers of HRSV. Virus was produced in HEp-2 cells grown on gelatin beads and concentrated either by precipitation with polyethylene glycol or by centrifugation over a sucrose cushion, the latter yielding 100% recovery. These procedures employed EDTA to disaggregate the virus and MgSO4 as a virus stabilizing agent. Attempts to purify HRSV over percoll, renografin and metrizamide gradients lead to loss of infectivity. Sucrose was found to represent the best purification medium with a yield of up to 60%.

Carcinoma↗
PubMed1990

The efficacy of intranasal interferon alpha-2a in respiratory syncytial virus infection in volunteers.

In a double-blind, placebo-controlled study, self-administered intranasal interferon alpha-2a or placebo was given both before and after challenge with respiratory syncytial virus. The incidence of colds and the severity of signs and symptoms were reduced in those receiving interferon alpha-2a as compared with those given placebo. In a further double-blind, placebo-controlled study, self-administered interferon alpha-2a or placebo was given only to those volunteers who developed colds following challenge with respiratory syncytial virus. There was no evidence that interferon alpha-2a reduced the severity of the signs and symptoms or shortened the duration of the illness. The similarity of these results to the effect of interferon alpha-2a in rhinovirus infections in volunteers is discussed.

Administration, Intranasal↗
PubMed1991

Respiratory syncytial virus and supraventricular tachycardia in an infant.

A 7-week infant admitted with bronchiolitis proven to be caused by respiratory syncytial virus developed a supraventricular tachycardia which responded to digitalization. She has remained well and no longer requires medication 6 months later, her electrocardiogram now being normal. The course of the illness suggests an association between infection with respiratory syncytial virus and the development of the tachycardia.

Bronchiolitis↗
PubMed1992

Gene junction sequences of bovine respiratory syncytial virus.

The nucleotide sequences of seven gene junctions (N-P, P-M, M-SH, SH-G, G-F, F-M2 and M2-L) of bovine respiratory syncytial virus (BRSV) strain A51908 were determined by dideoxynucleotide sequencing of cDNAs from polytranscript mRNAs and from genomic RNA. By comparison with the consensus sequences derived from human respiratory syncytial virus (HRSV) mRNAs, gene-start and gene-end sequences were found in all BRSV mRNAs. There was a perfect match between the BRSV and HRSV in all gene-start sequences, except for the sequence of the SH gene which contained one nucleotide difference compared to HRSV A2; and the gene-start sequence of the L gene, which was one nucleotide shorter than the corresponding sequence of HRSV. Analysis of the intergenic regions showed a high degree of divergence in the nucleotide sequence between BRSV and HRSV. However, the length of the nucleotides in the intergenic sequences was similar for a given gene junction. As in the case of HRSV, the M2 and L genes of BRSV overlap by 68 nucleotides, suggesting a similar transcription attenuation mechanism. The sequences of the overlap, corresponding to the 3' end of the L gene, were almost identical between BRSV and HRSV.

Animals↗
PubMed1995

Production and characterization of monoclonal antibodies to bovine respiratory syncytial virus.

Monoclonal antibodies to the SmithKline Beecham Animal Health BRSVR vaccine strain (375 isolate of Bovine Respiratory Syncytial Virus) were produced and then characterized by radioimmunoprecipitation followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, virus neutralization, inhibition of viral-induced fusion and isotype. Nineteen hybridomas produced antibodies that were reactive with the F0, F1, N or P viral proteins of bovine respiratory syncytial virus. One hybridoma (1E72C4) produced antibodies that immunoprecipitated the F0 glycoprotein, neutralized virus (1:8) in the presence of complement, but did not inhibit fusion. Another hybridoma (8B21E7) produced antibodies that immunoprecipitated the F0, F1 and F2 glycoproteins, neutralized virus (1:4) with and without complement and inhibited fusion. Antibodies from 11 hybridomas immunoprecipitated N and F1 proteins, one hybridoma immunoprecipitated the N and P protein, and 5 hybridomas immunoprecipitated the N protein. All monoclonal antibodies were of the IgG2b subtype with either kappa or lambda light chains.

Animals↗
PubMed1998

The effects of bovine respiratory syncytial virus on the phagocytic and antigen-presenting capacity of peripheral blood monocytes and monocytic cell lines derived from lambs and calves.

Human respiratory syncytial virus and bovine respiratory syncytial virus (BRSV) suppress lymphocyte responses to mitogens. In the present study, the possible effects of BRSV on some functions of antigen-presenting cells (APC) were investigated by exposing ovine monocytic cells to the virus before their use as APC. The depletion of monocytes from peripheral blood mononuclear cells resulted in the near total abrogation of proliferative responses to phytohaemagglutinin (PHA). Reactivity was restored by the addition of homologous monocytic cells derived from ovine peripheral blood monocytes as APC. The exposure of these monocytic cells to BRSV for 48 h before their use as APC significantly reduced the proliferative responses of uninfected ovine lymphocytes to PHA. Furthermore, the exposure of bovine peripheral blood monocytes and bovine and ovine monocytic cell lines to BRSV for 48 h reduced their capacity to phagocytize latex beads.

Animals↗
PubMed1990

Variation in severity of respiratory syncytial virus infections with subtype.

Two major subtypes of respiratory syncytial virus have been identified. This study assessed the hypothesis that A-subtype infections were more severe than B-subtype infections among the 157 infants hospitalized in two hospitals in Rochester, N.Y., during two winters. Severity was measured both by specific clinical observations and by a severity index that was derived empirically. Among all subjects, several clinical observations suggested that A-subtype infections were more severe. For example, mechanical ventilation was required in 12.6% of those with A-subtype compared with 1.6% of those with B-subtype infection (relative risk = 7.88; p = 0.01). Among high-risk infants (infants with underlying conditions or age 3 months or less at admission), carbon dioxide tension greater than 45 mm Hg was found in 37.0% of those with A-subtype compared with 12.0% of those with B-subtype infection (relative risk = 3.08; p = 0.04). In discrete multivariate (logit) analysis, effects of subtype (odds ratio = 6.59; p less than 0.01) on severity remained after adjustment for other statistically significant effects of age less than 3 months, underlying condition, and premature birth. The finding that A-subtype infections were more severe might have important implications for vaccine development, studies of the virulence of respiratory syncytial virus, clinical management (e.g., selection for antiviral therapy), and long-term prognosis.

Age Factors↗
PubMed1982

The role of respiratory syncytial virus and other viral pathogens in acute otitis media.

We utilized recently developed enzyme immunoassay techniques to examine the role of selected viruses in the etiology of acute otitis media. Viral pathogens were found in middle ear fluids obtained from 13 (24%) of 53 children with acute otitis media; respiratory syncytial virus accounted for ten of the 13 viral agents identified. In addition, respiratory syncytial viral antigen was found in nasopharyngeal washings obtained from 15 of the 53 children. Seven of these children had RSV identified as the sole middle ear pathogen, whereas six children had otitis caused by Streptococcus pneumoniae as either the sole middle ear pathogen or in combination with RSV. Similarly, all three children with respiratory infections caused by influenza virus had ear infections caused by bacterial pathogens, either alone or in combination with influenza virus. These findings suggest that, in patients with viral respiratory infection, coexisting acute otitis media may be associated with the recovery of either viruses or bacteria from the middle ear exudates.

Acute Disease↗
PubMed1999

Reducing the rate of nosocomially transmitted respiratory syncytial virus.

BACKGROUND: A large number (17) of nosocomial respiratory syncytial virus cases led to the development of control measures to prevent transmission of respiratory syncytial virus (RSV) within the Johns Hopkins Hospital's Children's Center. METHODS: The control plan is based on a 2-stage process. In stage 1, the staff are notified that RSV is in the community, and information is distributed through a communication tree. Stage 2 requires that nasopharyngeal aspirates be obtained from all children <3 years of age who have respiratory symptoms. The aspirates are tested directly for RSV antigen and cultured for RSV. The children are placed on pediatric droplet precautions pending those results. RESULTS: The proportion of nosocomial RSV cases dropped from 16.5% before the use of RSV control measures to 7.2% after the initiation of the control program. A case of RSV identified in the hospital was 2.6 times more likely to be nosocomially acquired before the intervention compared with after the intervention. Approximately 14 cases of RSV are prevented each year, which results in a savings of 56 hospital-days and more than $84,000 in direct hospital-related charges alone. CONCLUSIONS: The nosocomial spread of RSV can be reduced by a specific and feasible control plan that includes early identification and rapid isolation of potential RSV cases.

Baltimore↗