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Evaluation of two commercially available test methods to determine the feasibility of testing for respiratory syncytial virus in a community hospital laboratory.

The feasibility of performing respiratory syncytial virus (RSV) testing in a community hospital laboratory was assessed. The Abbott RSV EIA Kit, an enzyme immunoassay (EIA), and the Ortho Respiratory Syncytial Virus Identification Reagent, a direct fluorescent antibody (DFA) technique, were evaluated in terms of technologist time, result time, cost, and agreement of results. There was little difference in the amount of technologist time required for either method. The DFA technique was shown to be much less expensive than the EIA method and also required at least 50% less time to generate a final result. When compared with an indirect fluorescent antibody technique, the EIA technique was more sensitive (87%) than the DFA method (81%). Both methods showed 100% specificity. Based on the findings of this study, the DFA technique was determined to be the more feasible method for a community hospital laboratory to use for the rapid detection of RSV.

Child↗

Isolation of respiratory syncytial and influenza viruses from the sputum of patients hospitalized with pneumonia.

Routinely collected sputum specimens from 100 adults hospitalized with pneumonia were frozen at -70 C until inoculation into Madin-Darby canine kidney, fetal tonsil, and esophageal epithelial cells. Six influenza A (H3N2) viruses, two respiratory syncytial viruses, three rhinoviruses, and nine herpes simplex viruses were recovered. Four patients with influenza virus and one with respiratory syncytial virus isolated had nosocomial pneumonia. Viral isolation from sputum specimens may aid the diagnosis of pneumonia of unclear etiology and merits further evaluation as a diagnostic tool and as an adjunct to influenza surveillance.

Aged↗

Immunization with glycoprotein subunits of respiratory syncytial virus to protect cotton rats against viral infection.

The cotton rat model of respiratory syncytial virus infection was used to study immunization with viral glycoproteins. Animals immunized with the purified attachment protein (G) or the fusion protein (F) developed complete pulmonary resistance, but only partial nasal resistance, to challenge with respiratory syncytial virus. Antibody produced to the G protein neutralized virus, whereas antibody to the F protein neutralized virus and also inhibited fusion of infected cells. There was no evidence of enhanced pulmonary pathology in any immunized group.

Animals↗

Respiratory syncytial virus epidemics: variable dominance of subgroups A and B strains among children, 1981-1986.

We examined the distribution of subgroups A and B strains from respiratory syncytial virus during five epidemic years from 1981 to 1986 in Huntington, West Virginia. Of 235 infants and children with respiratory syncytial virus infection, 211 had virus reisolated from frozen throat swab specimens for subgroup characterization by reactivity with a panel of monoclonal antibodies to the G, F, NP, M, and P proteins by using an enzyme immunoassay. We identified 160 (75.8%) strains as subgroup A and 51 (24.2%) as subgroup B. Strains of both subgroups were isolated in all years. Small, but approximately equal, numbers of subgroup B strains were isolated each epidemic year. By contrast, subgroup A strains occurred at least three times as often in all years except 1984-1985. The very low number of subgroup A strains isolated during the 1984-1985 epidemic gave dominance to subgroup B strains.

Antibodies, Monoclonal↗

Immunization of cotton rats with the human respiratory syncytial virus F glycoprotein produced using a baculovirus vector.

The F glycoprotein of respiratory syncytial virus in insect cells was produced using a baculovirus expression vector to examine its potential as a subunit vaccine. Two different forms of the F glycoprotein were expressed: the intact F (F) glycoprotein and the truncated (Ft) glycoprotein, in which the COOH-terminal anchor region was deleted. The F glycoprotein remained cell associated, whereas the Ft glycoprotein was secreted into the media of infected cells. In contrast to the processing of the F0 precursor into its F1 and F2 subunits that was observed in mammalian cells, a second cleavage site within the F1 subunit was recognized by the insect cell proteases and resulted in the formation of two F1 subunits. The baculovirus-expressed Ft glycoprotein induced neutralizing antibodies in cotton rats and protected vaccinated animals from challenge with respiratory syncytial virus.

Animals↗

Risk factors for respiratory syncytial virus-associated lower respiratory illnesses in the first year of life.

The relation of breast feeding and other factors to the incidence of respiratory syncytial virus-associated lower respiratory tract illness (RSV-LRI) in the first year of life is examined. The study population is 1,179 healthy infants enrolled at birth between May 1980 and January 1984 into the Tucson Children's Respiratory Study, Tucson, Arizona. Each subject's data were assessed at each month of age during the first year of life, during those months when respiratory syncytial virus was isolated. A number of significant relations were observed, particularly between 1 and 3 months of age. At this age, the risk of having a RSV-LRI increased in association with less than 1-month or no breast feeding, with being male, and with increasing numbers of others sharing the child's bedroom. In multivariate analysis, only sex and the number of others sharing the room remained as significant direct effects. However, a significant interaction demonstrated that breast feeding has a protective role in relation to RSV-LRIs for those infants of mothers with a lower education level. The risk of having a RSV-LRI increases with combinations of risk factors. Being in day care was a significant risk factor in the 7- to 9-month age range. The RSV-LRI rate also varies by birth month. A separate case-control study assessed relations of RSV-LRIs with cord serum RSV antibody. Those with lower cord serum RSV antibody, who also have minimal breast feeding, were found to be especially at risk for RSV-LRIs in the first 5 months of life.

Antibodies, Viral↗

A technique for the administration of ribavirin to mechanically ventilated infants with severe respiratory syncytial virus infection.

Fifteen infants with respiratory syncytial virus pulmonary infection admitted to our pediatric ICU from December 1, 1985 through April 30, 1986, required mechanical ventilation. These patients were placed on an open trial of ribavirin therapy. We describe a technique for the safe delivery of aerosolized ribavirin to these infants while on the ventilator. The agent was delivered for 16 h/day for 7 days. Modifications of the ventilator circuit were needed to prevent the condensation of the drug in the ventilator tubing and to allow for the safe and effective operation of the ventilator. A common ventilator strategy was used for all patients. The highest positive inspiratory pressure generated was 42 +/- 9.5 (SD) cm H2O, the highest PEEP was 5.9 +/- 3.2 cm H2O, the duration of ventilation was 10.7 +/- 8.5 days, and exposure to fraction of inspired oxygen was greater than or equal to 0.6 for 55.3 h. Ribavirin levels were measurable in two patients, thereby demonstrating that the drug was in fact delivered and absorbed. Our preliminary results demonstrate that ribavirin can be delivered to the patients with respiratory syncytial viral infections who require mechanical ventilation; however, further studies are indicated to evaluate the efficacy and dose responsiveness, alterations in pulmonary dynamics, and safety of ribavirin in delivery to infants requiring ventilation.

Administration, Inhalation↗

Respiratory syncytial virus-induced acute lung injury in adult patients with bone marrow transplants: a clinical approach and review of the literature.

Acute lung injury induced by respiratory syncytial virus (RSV) is a major cause of morbidity and mortality in patients who have undergone bone marrow transplantation. Twenty-nine of the 74 patients who received bone marrow transplants at the University of Minnesota during a 1-year period developed evidence of acute lung injury, and RSV was identified as the cause in 8. We discuss the clinical course of these 8 patients and offer a clinical approach to RSV infection occurring after bone marrow transplantation. We also review the immune response to infection with RSV and relate this information to the nature and degree of immunosuppression present in patients undergoing this type of transplantation. We found bronchoalveolar lavage with rapid antigen detection to be particularly useful for the prompt diagnosis of this serious infection. The virus was obtained from the lower respiratory tract of each patient and was identified in lavage effluent by culture and by antigen detection (ELISA). The mean time to a positive culture was 6 days, while detection of antigens of respiratory syncytial virus by ELISA was completed within 18 hours in all cases. The clinical progression of the illness in immunocompromised patients appears to be the same as in non-immunocompromised persons: upper respiratory tract infection and illness precede lower respiratory tract infection and acute lung injury. Seven of our 8 patients had upper respiratory tract symptoms or abnormal sinus radiographs, and upper respiratory specimens (cultures and ELISA from nasopharynx, throat, and sputum) were positive in 5 of 8 patients. Six patients developed RSV-induced lung injury before marrow engraftment; 4 of them had respiratory failure requiring mechanical ventilation and died, including 3 in whom RSV was eliminated from the lower respiratory tract following treatment with ribavirin aerosol. Two additional pre-engraftment patients had only relatively mild lung injury 4 days after beginning treatment with ribavirin for RSV infection in the upper respiratory tract. Their recovery suggests that early treatment may ameliorate RSV-induced lung injury. The remaining 2 patients developed lung injury after marrow engraftment. Both of these patients had clear chest radiographs, responded clinically to ribavirin, and survived. RSV is a potentially treatable cause of life-threatening lung injury, if the physician is aggressive in identifying the virus in the upper respiratory tract before evidence of lung injury appears. Rapid detection methods are essential when bone marrow transplant patients have fever along with signs, symptoms, or radiographic indications of nasal or sinus disorders.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗

Comparison of nasal brush and nasopharyngeal aspirate techniques in obtaining specimens for detection of respiratory syncytial viral antigen by immunofluorescence.

Diagnosis of respiratory syncytial virus by antigen detection is dependent on obtaining adequate respiratory epithelial cells. Two specimen collection methods, nasopharyngeal aspiration (NPA) and nasal brushing (NB), were compared. Thirty-two pediatric patients with presumed viral pneumonia or bronchiolitis (34 episodes) had both NPA and NB performed. Of 34 specimens 15 were culture-positive for respiratory syncytial virus. Of these 12 NPA samples and 10 NB samples had viral inclusions by immunofluorescent antibody staining (IFA). Of culture-negative samples, 1 of 17 NB was positive by IFA. One specimen obtained by NB had too few cells to read by the IFA method. Sensitivity and specificity were 80 and 100% for NPA and 67 and 94% for NB. Total respiratory cells and IFA-positive cells (classified as few, moderate, or many) were greater with NPA; however, NB was also an effective procedure and was better tolerated by children, less expensive and easier to perform.

Antigens, Viral↗

Historical cohort evaluation of ribavirin efficacy in respiratory syncytial virus infection.

Evaluation of ribavirin therapy for respiratory syncytial virus infection of the lower respiratory tract is problematic because of multiple risk factors and variable severity of illness in respiratory syncytial virus-infected patients. To address these difficulties we used multivariate analysis and performed a historical concurrent cohort study in two children's hospitals one of which had used ribavirin since licensing and one that had not utilized ribavirin therapy. The medical records of 158 patients who satisfied the American Academy of Pediatrics inclusion criteria for receiving ribavirin were analyzed for three seasons (1988 to 1991). No significant benefit of ribavirin therapy could be appreciated for the whole group in length of stay (median, 5.0 vs. 6.5 days), days on oxygen therapy (median, 5 vs. 3), progression to ventilator status (3.8 vs. 3.9%) or mortality (1.9% vs. 1.9%) for ribavirin treatment vs. supportive care. Multivariate analysis failed to uncover a beneficial effect of ribavirin when all risk factors were considered. No significant differences were noted when ventilated and nonventilated patients were examined separately. Our data raise questions about the efficacy of ribavirin when used in common practice and suggest that further prospective study, with appropriate analysis, is needed to justify the continued widespread use of this drug.

Cohort Studies↗

Ribavirin effect on pulmonary function in young infants with respiratory syncytial virus bronchiolitis.

To assess the effect of ribavirin on pulmonary function in infants with respiratory syncytial virus bronchiolitis, we performed a randomized (nonmatched), double blinded, placebo-controlled study of 19 infants with RSV bronchiolitis. Infants with underlying respiratory, cardiac or immunologic disease were excluded. Patients were given ribavirin (10) or placebo (9) via an aerosol generator for 18 hours/day for 3 days. Pulmonary function (dynamic compliance, total lung resistance) was calculated using a pneumotachographic method on Days 1, 2 and 7. Differences between groups on clinical criteria were not found. Approximately one-half of each group showed increased compliance and decreased lung resistance after 24 to 48 hours of therapy. By Day 7 compliance had increased 30% in the placebo group and 210% in the ribavirin-treated infants (P = 0.05). Significant differences in the rate of change of lung resistance were not seen by Day 7. We conclude that previously noted improvements in the early course of respiratory syncytial virus bronchiolitis treated with ribavirin do not appear to be a result of measurable changes in pulmonary function. However, paradoxical increases in airway resistance were not found in patients treated with ribavirin.

Aerosols↗

Presumed respiratory syncytial virus pneumonia in an adolescent compromised host.

A 15-year-old boy with nephrotic syndrome, renal insufficiency, and cutaneous anergy had severe pneumonia with pleural effusion. There was serologic evidence of respiratory syncytial virus infection, and extensive microbiologic, histologic, and serologic studies failed to identify any other etiologic agent. Respiratory syncytial virus as a possible cause of severe pneumonia in immunocompromised hosts has not been previously reported.

Adolescent↗

Management of respiratory syncytial virus with lower respiratory tract infection in infants and children.

Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infection in infants and children requiring pediatric hospitalizations. Infants with chronic lung, cardiac, or neuromuscular conditions are at increased risk for RSV infection. Early RSV is associated with subsequent diagnosis of reactive airway disease. The management of RSV with lower respiratory track infection in infants and children remains controversial. Bronchodilators may have some short-term benefit, but are not recommended as standard practice for infants and children. Antiviral therapy may be used for high-risk and severely ill patients. Corticosteroids may be effective in cases of moderate to severe RSV with lower respiratory track infection. Monoclonal antibodies have shown some promise in achieving passive immunity for those at greatest risk, including preterm infants younger than 1 year or infants younger than 2 years with chronic lung disease. Emergency management remains primarily supportive, with vigilant monitoring of oxygenation and hydration status. Interventions include supplemental oxygen therapy, ventilation, and fluid and nutrition therapy. Respiratory syncytial virus prophylaxis for high-risk patients includes intramuscular injections of palivizumab (Synagis) each month during RSV season, from November through April. Prevention strategies include washing hands, cleaning environment surfaces, and isolating infants and children with RSV in the emergency care area.

Adrenal Cortex Hormones↗

Evaluation of the efficacy of prednisolone in early wheezing induced by rhinovirus or respiratory syncytial virus.

BACKGROUND: The role of systemic corticosteroids in the treatment of early childhood wheezing in children is not clear. OBJECTIVE: We sought to determine whether prednisolone is effective in rhinovirus-induced early wheezing. METHODS: We conducted a controlled trial comparing oral prednisolone (2 mg/kg per day in three divided doses for 3 days) with placebo in 78 hospitalized children (mean age, 1.1 year; standard deviation, 0.7) experiencing their first or second episode of wheezing induced by rhinovirus or respiratory syncytial virus. Mixed viral infections were excluded. Our primary end point was the time until the patient was ready for discharge; secondary end points included oxygen saturation during hospitalization, duration of symptoms, occurrence of relapses during the next 2 months and blood eosinophil counts at discharge and 2 weeks later. RESULTS: In multivariate regression analysis, prednisolone did not influence the time until ready for discharge, but it decreased relapses during the subsequent 2-month period in rhinovirus-affected children (prednisolone versus placebo, 22% versus 56%; odds ratio, 19.06; 95% confidence interval, 2.52-144.03; P = 0.004) and in children with blood eosinophils > or = 0.2 x 10/L (respectively, 24% versus 71%; odds ratio, 10.57; 95% confidence interval, 1.99-56.22; P = 0.006). Rhinovirus-affected children had more blood eosinophils on admission (mean, 0.44 versus 0.086 x 10/L), had a higher prevalence of atopy (44% versus 8%) and were older (mean, 1.4 versus 0.9 years, P < 0.001 for all) than respiratory syncytial virus-infected children. CONCLUSION: Prednisolone reduced relapses during a 2-month period after first episodes of wheezing associated with rhinovirus infection or blood eosinophils > or = 0.2 x 10/L.

Acute Disease↗

Virus development in enucleate cells: echovirus, poliovirus, pseudorabies virus, reovirus, respiratory syncytial virus and Semliki Forest virus.

A group of RNA viruses, echovirus, poliovirus, reovirus, respiratory syncytial virus and Semliki Forest virus have been examined for ability to grow in enucleate African green monkey kidney (BSCi) cells. Semliki Forest virus produced an almost normal yield of virus but poliovirus, echovirus, reovirus and respiratory syncytial virus, although showing clear evidence of virus replication when compared with a nuclear DNA virus (pseudorabies virus) gave much lower yields than those from nucleate cells. Analysis of enucleate cells infected with echovirus and reovirus showed no evidence of a specific block in the synthesis of any virus-specified polypeptide. Infection with vesicular stomatitis virus at intervals after enucleation demonstrated a diminishing ability to support virus growth with increasing time. It is suggested that the yield of virus obtained from an enucleate cell is related to the length of the growth cycle of the virus, the reduced yield obtained with some viruses reflecting the declining ability of the enucleate cell to support virus growth.

Autoradiography↗

Sequence comparison between the fusion protein of human and bovine respiratory syncytial viruses.

The nucleotide sequence was determined for the fusion (F) protein-coding mRNA of the bovine respiratory syncytial virus (strain RB 94) and the amino acid sequence of the F protein was deduced for comparison with the sequence of human respiratory syncytial virus subtypes A and B (RSS-2 and 18537 strains). The human and bovine RS virus F proteins (excluding the cleaved signal peptide) share 83 to 84% homology. The greatest divergence occurred within the F2 subunit in the region preceding the cleavage activation site.

Amino Acid Sequence↗

Defective synthesis of envelope proteins by temperature-sensitive mutants representing complementation groups B and D of respiratory syncytial virus.

The phenotypes of two complementing temperature-sensitive (ts) mutants of respiratory syncytial (RS) virus indicate that the mutational lesions involve the attachment (G) and matrix (M) proteins of the viral envelope. Synthesis of the G protein was affected in cells infected with mutant tsA2 (complementation group B); the p50 precursor of the G protein was synthesized normally, but further maturation to the fully glycosylated form was defective at 39 degrees C. A non-ts alteration in the efficiency of cleavage of the F0 precursor to the F1 and F2 subunits of the fusion protein was also observed in tsA2-infected cells, which is consistent with the aberrant non-syncytial plaque morphology induced by tsA2 in certain cells. In cells infected with mutant tsN1 (complementation group D) the M protein disappeared from the soluble cytoplasmic fraction soon after synthesis at 39 degrees C and had a slightly decreased electrophoretic mobility. The M protein of non-ts revertants was stable at 39 degrees C, which links the defect in M protein stability with the tsN1 phenotype. However, the aberrant mobility phenotype remained, suggesting pseudoreversion. These results assign two of the eight complementation groups of ts mutants of RS virus.

Cell Line↗

Bovine respiratory syncytial virus nucleocapsid protein: mRNA sequence analysis and expression from recombinant vaccinia virus vectors.

The nucleotide sequence of the mRNA encoding the nucleocapsid (N) protein of bovine respiratory syncytial (BRS) virus, strain 391-2, was determined. Recombinant vectors containing a cDNA of the complete N gene were constructed, and expression of the N protein in eukaryotic cells was demonstrated using two different vector systems. The BRS virus N mRNA was 1197 nucleotides in length, exclusive of poly(A), and had a single major open reading frame that encoded a polypeptide of 391 amino acids with a calculated M(r) of 42.6K. The nucleotide and amino acid sequences of the BRS virus N gene were compared to those of human respiratory syncytial (HRS) virus strains A2 and 18537, and to BRS virus strain A51908. The level of nucleic acid identity between the N mRNA of BRS virus 391-2 and both HRS virus subtypes was 80 to 81%, whereas the identity between the two BRS virus strains was 97%. A 93 to 94% level of identity was observed between the deduced amino acid sequences of the N protein of BRS virus 391-2 and the corresponding sequences of the two HRS virus strains. The two BRS virus N proteins differed in amino acid sequence at only three positions. Recombinant BRS virus N protein was expressed using two different vector systems: in cells from a plasmid using the vaccinia virus/T7 polymerase expression system or from a recombinant vaccinia virus. N proteins synthesized by the two vector systems migrated with an electrophoretic mobility identical to that of authentic BRS virus N protein, and were precipitated by anti-BRS virus antibodies.

Amino Acid Sequence↗