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  1. Respiratory Syncytial Virus

    MedlinePlus

    ... Palsy: Shannon's Story" 5 Things to Know About Zika & Pregnancy Respiratory Syncytial Virus KidsHealth > For Parents > Respiratory Syncytial Virus Print A ... often get it when older kids carry the virus home from school and pass it to ... often happen in epidemics that last from late fall through early spring. ...

  2. Respiratory Syncytial Virus Infections

    MedlinePlus

    Respiratory syncytial virus (RSV) causes mild, cold-like symptoms in adults and older healthy children. It can cause serious problems in ... tests can tell if your child has the virus. There is no specific treatment. You should give ...

  3. Diagnosing and treating respiratory syncytial virus bronchiolitis.

    PubMed

    Napierkowski, Daria B

    2016-09-22

    Respiratory syncytial virus (RSV) is one of the major causes of respiratory tract illness in children and can lead to significant infection and death. This article discusses the incidence, clinical presentation, diagnosis, current treatment, and prevention options to successfully diagnose and treat infections caused by RSV. PMID:27552683

  4. Respiratory syncytial virus infection in cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine respiratory syncytial virus (bRSV) is a cause of respiratory disease in cattle world-wide. It has an integral role in enzootic pneumonia in young dairy calves and summer pneumonia in nursing beef calves. Furthermore, bRSV infection can predispose calves to secondary bacterial infection by org...

  5. Reverse Genetics of Respiratory Syncytial Virus.

    PubMed

    Stobart, Christopher C; Hotard, Anne L; Meng, Jia; Moore, Martin L

    2016-01-01

    Respiratory syncytial virus (RSV) is a negative-strand RNA virus that is associated with severe lower respiratory tract infections in young infants and the elderly. RSV remains a leading cause worldwide of infant mortality, and despite the high clinical and economic burden of the virus there are currently no available vaccines. Here, we describe the methods for recovery of recombinant RSV viruses using a bacterial artificial chromosome and methods related to procurement and expansion of stocks of RSV mutants. PMID:27464692

  6. The changing landscape of respiratory syncytial virus.

    PubMed

    Polack, Fernando P

    2015-11-25

    Recognition of the acute and chronic burden of respiratory syncytial virus (RSV) lower respiratory tract infections (LRTI) sparked a wave of initiatives to develop preventive and therapeutic products against the pathogen in recent years. RSV is a leading cause of hospitalization in infants in industrialized and developing countries, has been causally linked to recurrent wheezing during childhood, associated with pediatric asthma, and is an important cause of mortality in the first months of life in the developing world. Significant changes in the epidemiology, clinical manifestations, and severe consequences of LRTI may emerge in the next decade with the advent of novel preventive strategies against RSV. This manuscript outlines some of these changes and discusses potential scenarios based on the current literature and experiences with other pathogens.

  7. Clinical experience with respiratory syncytial virus vaccines.

    PubMed

    Piedra, Pedro A

    2003-02-01

    Respiratory syncytial virus (RSV) infection is at times associated with life-threatening lower respiratory tract illness in infancy. Severe infection during the first year of life may be an important risk factor or indicator for the development of asthma in early childhood. Severe infections primarily occur in healthy infants, and young infants and children with specific risk factors. However, RSV causes respiratory infections in all age groups. Indeed it is now recognized that RSV disease is responsible for significant morbidity and mortality in the geriatric population. RSV infection remains difficult to treat, and prevention is a worldwide goal. For this reason there has been an intensive effort to develop an effective and safe RSV vaccine. Initial infection with RSV affords limited protection to reinfection, yet repeated episodes decrease the risk for lower respiratory tract illness. In the 20 years from 1960 to 1980, trials of several candidate RSV vaccines failed to attain the desired safety and protection against natural infection. Some vaccine types either failed to elicit immunogenicity, as with the live subcutaneous vaccine, or resulted in exaggerated disease on natural exposure to the virus, as with the formalin-inactivated (FI) type. Currently vaccine candidates are being developed based on the molecular virology of RSV. Recent formulations of candidate RSV vaccines have focused on subunit vaccines [such as purified fusion protein (PFP)], subunit vaccines combined with nonspecific immune activating adjuvants, live attenuated vaccines (including cold passaged, temperature-sensitive or cpts mutants), genetically engineered live attenuated vaccines and polypeptide vaccines. PMID:12671459

  8. Human Respiratory Syncytial Virus: An Introduction.

    PubMed

    Jorquera, Patricia A; Anderson, Lydia; Tripp, Ralph A

    2016-01-01

    Human respiratory syncytial virus (RSV) is understood to be a significant human pathogen in infants, young children, and the elderly and the immunocompromised. Over the last decade many important mechanisms contributing to RSV infection, replication, and disease pathogenesis have been revealed; however, there is still insufficient knowledge which has in part hampered vaccine development. Considerable information is accumulating regarding how RSV proteins modulate molecular signaling and immune responses to infection. Understanding how RSV interacts with its host is crucial to facilitate the development of safe and effective vaccines and therapeutic treatments.In this chapter, we provide a brief introduction into RSV replication, pathogenesis, and host immune response, and summarize the state of RSV vaccine and antiviral compounds in clinical stages of development. This chapter frames features of this book and the molecular methods used for understanding RSV interaction with the host. PMID:27464683

  9. Gold nanorod vaccine for respiratory syncytial virus

    NASA Astrophysics Data System (ADS)

    Stone, John W.; Thornburg, Natalie J.; Blum, David L.; Kuhn, Sam J.; Wright, David W.; Crowe, James E., Jr.

    2013-07-01

    Respiratory syncytial virus (RSV) is a major cause of pneumonia and wheezing in infants and the elderly, but to date there is no licensed vaccine. We developed a gold nanorod construct that displayed the major protective antigen of the virus, the fusion protein (F). Nanorods conjugated to RSV F were formulated as a candidate vaccine preparation by covalent attachment of viral protein using a layer-by-layer approach. In vitro studies using ELISA, electron microscopy and circular dichroism revealed that conformation-dependent epitopes were maintained during conjugation, and transmission electron microscopy studies showed that a dispersed population of particles could be achieved. Human dendritic cells treated with the vaccine induced immune responses in primary human T cells. These results suggest that this vaccine approach may be a potent method for immunizing against viruses such as RSV with surface glycoproteins that are targets for the human immune response.

  10. Respiratory Syncytial Virus Infection (RSV): Transmission and Prevention

    MedlinePlus

    ... CDC Cancel Submit Search The CDC Respiratory Syncytial Virus Infection (RSV) Note: Javascript is disabled or is ... school or childcare. They can then transmit the virus to other members of the family. RSV can ...

  11. Diagnosis of Respiratory Syncytial Virus Infection

    PubMed Central

    Popow-Kraupp, Therese; Aberle, Judith H

    2011-01-01

    Respiratory syncytial virus (RSV) is one of the most important pathogen causing severe lower respiratory tract infections in all age groups often requiring hospitalization. Rapid laboratory diagnosis of RSV infection significantly decreases the use of antibiotics, additional laboratory testing and is associated with shorter hospitalization periods. The specific diagnosis of RSV infection is based on the detection of virus or viral antigens or virus specific nucleic acid sequences in respiratory secretions. The kind and quality of the clinical specimen exerts a considerable influence on the results of all currently used viral detection assays. Antigen based tests are widely available, easy to perform and the results are available in a short time but their reduced sensitivity and specificity represent a considerable shortcoming. Among the methods available isolation in cell culture was considered the gold standard for the sensitive identification of RSV but is gradually replaced by highly sensitive and specific nucleic acid amplification assays that provide more rapid results. Of these reverse transcription polymerase chain reaction (PCR) was the first and is still the most frequently used nucleic acid-based assay. New methodologies, as for example the real-time PCR methods allow the quantification of viral nucleic acids in the clinical sample. Disadvantages of the nucleic acid based assays are their high costs and their limited standardization. Future research on new methodologies for the diagnosis of viral respiratory tract infections should focus on the development of sensitive, rapid and cost effective test systems allowing the screening for all probable causative agents. In addition varying testing protocols for summer and winter months based on epidemiologic data are needed to direct their practical use. PMID:22262985

  12. [Infantile meningitis caused by respiratory syncytial virus].

    PubMed

    Shirota, Go; Morozumi, Miyuki; Ubukata, Kimiko; Shiro, Hiroyuki

    2011-11-01

    Respiratory syncytial (RS) virus commonly causes infantile respiratory tract infection causing significant morbidity and mortality, but rarely meningitis. We report a case of meningitis caused by RS virus subgroup B in a 56-day-old boy admitted for high fever who underwent blood examination and lumbar puncture. Empirical chemotherapy was started with intravenous ampicillin, gentamicin, and cefotaxime based on laboratory data on CSF cells (84/microL) and serum CRP (13.8mg/dL) data. RS virus subgroup B was only detected using real-time PCR comprehensive reverse transcription from the first CSF, but no bacterial gene was detected. No bacteria grew from his CSF, urine, or blood. Fever and serum CRP dropped in a few days. He had neither seizures nor disturbance of consciousness and was discharged on day 11 after admission. No evidence of encephalopathy was detected in brain MRI or electroencephalography. RS virus rarely causes meningitis, but a percentage of RS-virus-infected infants exhibit symptoms such as seizure and disturbance of consciousness. We should recognize that the RS virus may cause neurological complications associated with high morbidity and mortality.

  13. Bovine respiratory syncytial virus protects cotton rats against human respiratory syncytial virus infection.

    PubMed

    Piazza, F M; Johnson, S A; Darnell, M E; Porter, D D; Hemming, V G; Prince, G A

    1993-03-01

    Human respiratory syncytial virus (HRSV) is the most frequent cause of severe respiratory infections in infancy. No vaccine against this virus has yet been protective, and antiviral drugs have been of limited utility. Using the cotton rat model of HRSV infection, we examined bovine respiratory syncytial virus (BRSV), a cause of acute respiratory disease in young cattle, as a possible vaccine candidate to protect children against HRSV infection. Cotton rats were primed intranasally with graded doses of BRSV/375 or HRSV/Long or were left unprimed. Three weeks later, they were challenged intranasally with either BRSV/375, HRSV/Long (subgroup A), or HRSV/18537 (subgroup B). At intervals postchallenge, animals were sacrificed for virus titration and histologic evaluation. Serum neutralizing antibody titers were determined at the time of viral challenge. BRSV/375 replicated to low titers in nasal tissues and lungs. Priming with 10(5) PFU of BRSV/375 effected a 500- to 1,000-fold reduction in peak nasal HRSV titer and a greater than 1,000-fold reduction in peak pulmonary HRSV titer upon challenge with HRSV/Long or HRSV/18537. In contrast to priming with HRSV, priming with BRSV did not induce substantial levels of neutralizing antibody against HRSV and was associated with a delayed onset of clearance of HRSV upon challenge. Priming with BRSV/375 caused mild nasal and pulmonary pathology and did not cause exacerbation of disease upon challenge with HRSV/Long. Our findings suggest that BRSV may be a potential vaccine against HRSV and a useful tool for studying the mechanisms of immunity to HRSV.

  14. Animal Models of Respiratory Syncytial Virus Infection and Disease

    PubMed Central

    Sacco, Randy E.; Durbin, Russell K.; Durbin, Joan E.

    2015-01-01

    The study of human respiratory syncytial virus pathogenesis and immunity has been hampered by its exquisite host specificity, and the difficulties encountered in adapting this virus to a murine host. The reasons for this obstacle are not well understood, but appear to reflect, at least in part, the inability of the virus to block the interferon response in any but the human host. This review addresses some of the issues encountered in mouse models of respiratory syncytial virus infection, and describes the advantages and disadvantages of alternative model systems. PMID:26176495

  15. Activation of Complement by Cells Infected with Respiratory Syncytial Virus

    PubMed Central

    Smith, Thomas F.; Mcintosh, Kenneth; Fishaut, Mark; Henson, Peter M.

    1981-01-01

    The ability of respiratory syncytial virus (RSV)-infected HEp-2 cells in culture to activate complement was investigated. After incubation of cells with various complement sources and buffer, binding of C3b to surfaces of infected cells was demonstrated by immunofluorescence with a double-staining technique. Nonsyncytial and syncytial (i.e., fused, multinucleated) cells were separately enumerated. Also, lysis of RSV-infected cells was assessed by lactic dehydrogenase release. In this system only RSV-infected cells stained for C3b, and they did so only after incubation with functionally active complement. Blocking of classical pathway activation with ethylenediaminetetraacetic acid diminished the number of infected nonsyncytial cells positively stained for C3b, but had no effect on staining of syncytial cells. Blocking of alternative pathway activation with either zymosan incubation or heat treatment decreased the number of both syncytial and nonsyncytial cells stained for C3b. Decreasing immunoglobulin concentration of the serum used as the complement source also decreased numbers of both cell types stained for C3b. Eliminating specific anti-RSV antibody diminished numbers of both cell types stained for C3b, but staining was not eliminated. Lastly, incubation with functionally active complement markedly increased lactic dehydrogenase release from infected cells. This study demonstrated that RSV-infected nonsyncytial and syncytial cells are able to activate complement by both classical and alternative pathways. Activation of complement by syncytial cells appears to be less dependent on the classical pathway than is activation by nonsyncytial cells, and activation by syncytial cells may require immunoglobulin but not specific antibody. These experiments suggest the possibility of complement activation during respiratory tract infection by RSV. Implications of this are discussed. Images PMID:7263071

  16. Animal models of respiratory syncytial virus infection and disease

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The study of human respiratory syncytial virus pathogenesis and immunity has been hampered by its exquisite host specificity, and the difficulties encountered in adapting this virus to a murine host. The reasons for this obstacle are not well understood, but appear to reflect, at least in part, the ...

  17. 21 CFR 866.3480 - Respiratory syncytial virus serological reagents.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Respiratory syncytial virus serological reagents. 866.3480 Section 866.3480 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents §...

  18. 21 CFR 866.3480 - Respiratory syncytial virus serological reagents.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Respiratory syncytial virus serological reagents. 866.3480 Section 866.3480 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents §...

  19. 21 CFR 866.3480 - Respiratory syncytial virus serological reagents.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Respiratory syncytial virus serological reagents. 866.3480 Section 866.3480 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents §...

  20. 21 CFR 866.3480 - Respiratory syncytial virus serological reagents.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Respiratory syncytial virus serological reagents. 866.3480 Section 866.3480 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents §...

  1. 21 CFR 866.3480 - Respiratory syncytial virus serological reagents.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Respiratory syncytial virus serological reagents. 866.3480 Section 866.3480 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents §...

  2. Immunoprophylaxis of respiratory syncytial virus infection in the infant ferret.

    PubMed

    Suffin, S C; Prince, G A; Muck, K B; Porter, D D

    1979-07-01

    Infant ferrets can be protected from respiratory syncytical virus challenge at 3 days of age by gestational infection of their mothers. Ferrets acquire their immunity to respiratory syncytial virus postpartum via immunizing products of lactation. The level of protection against viral replication correlates with the maternal serum neutralizing titer or a concomitant factor. Passive administration of adult ferret serum with a neutralizing titer of 1:1024 or greater, either i.p. or orally does not confer immunity. A nonantibody-mediated protective mechanism appears to play an important role in protecting the infant ferret from respiratory syncytial virus replication. Our findings allow the testing of the efficacy of future human vaccines before human clinical trial. PMID:448139

  3. Hot topics in the prevention of respiratory syncytial virus disease.

    PubMed

    Habibi, Maximillian S; Patel, Sanjay; Openshaw, Peter

    2011-03-01

    The 7th International Respiratory Syncytial Virus Symposium took place in Hotel Blijdorp, Rotterdam, The Netherlands. The series has been running since 1996; this meeting took place after a 3-year gap, and was attended by approximately 200 clinicians, scientists and industry representatives from all over the world. The conference covered all aspects of respiratory syncytial virus disease, including virology, cell biology, pathogenesis, clinical presentation, diagnosis, immunology, vaccines, antivirals and other therapeutic approaches. Reviews by invited keynote speakers were accompanied by oral and poster presentations, with ample opportunity for discussion of unpublished work. This article summarizes a small selection of hot topics from the meeting, focused on pathogenesis, therapeutics and vaccine development. PMID:21434796

  4. Production of interferon in respiratory syncytial virus bronchiolitis.

    PubMed Central

    Isaacs, D

    1989-01-01

    Production of interferon alfa in vitro was significantly reduced during acute respiratory syncytial virus bronchiolitis but subsequently returned to normal. Nasopharyngeal and endotracheal interferon alfa were detected intermittently and in low concentrations. The degree of impairment of in vitro production and poor in vivo production of interferon alfa suggest the need for a therapeutic trial of nebulised or systemic interferon in acute bronchiolitis. PMID:2923479

  5. Structural basis of respiratory syncytial virus neutralization by motavizumab

    SciTech Connect

    McLellan, Jason S.; Chen, Man; Kim, Albert; Yang, Yongping; Graham, Barney S.; Kwong, Peter D.

    2010-04-13

    Motavizumab is {approx}tenfold more potent than its predecessor, palivizumab (Synagis), the FDA-approved monoclonal antibody used to prevent respiratory syncytial virus (RSV) infection. The structure of motavizumab in complex with a 24-residue peptide corresponding to its epitope on the RSV fusion (F) glycoprotein reveals the structural basis for this greater potency. Modeling suggests that motavizumab recognizes a different quaternary configuration of the F glycoprotein than that observed in a homologous structure.

  6. Respiratory Syncytial Virus Hospitalizations in Healthy Preterm Infants

    PubMed Central

    Margulis, Andrea V.; Samuel, Miny; Lohr, Kathleen N.

    2016-01-01

    Background: Studies have explored the risk for and impact of respiratory syncytial virus (RSV) infection requiring hospitalization among healthy preterm infants born at 29–35 weeks of gestational age not given RSV immunoprophylaxis. We performed a systematic review and qualitative synthesis of these studies. Methods: Two experienced reviewers used prespecified inclusion/exclusion criteria to screen titles/abstracts and full-text studies using MEDLINE, Embase, BIOSIS and Cochrane Library (January 1, 1985, to November 6, 2014). We abstracted data on risk factors for RSV hospitalization, incidence and short- and long-term outcomes of RSV hospitalization. Using standard procedures, we assessed study risk of bias and graded strength of evidence (SOE). Results: We identified 4754 records and reviewed 27. Important risk factors for RSV hospitalization included young age during the RSV season, having school-age siblings and day-care attendance, with odds ratios >2.5 in at least one study (high SOE). Incidence rates for RSV hospitalizations ranged from 2.3% to 10% (low SOE). Length of hospital stays ranged from 3.8 to 6.1 days (low SOE). Recurrent wheezing rates ranged from 20.7% to 42.8% 1 to 2 years after RSV hospitalization (low SOE). Conclusions: Young chronological age and some environmental risk factors are important clinical indicators of an increased risk of RSV hospitalization in healthy preterm infants 32 to 35 weeks of gestational age. SOE was low for estimates of incidence of RSV hospitalizations, in-hospital resource use and recurrent wheezing in this population. Studies were inconsistent in study characteristics, including weeks of gestational age, age during RSV season and control for confounding factors. PMID:27093166

  7. Mouse and Cotton Rat Models of Human Respiratory Syncytial Virus.

    PubMed

    Rudd, Penny A; Chen, Weiqiang; Mahalingam, Suresh

    2016-01-01

    Human respiratory syncytial virus (hRSV) is a common respiratory virus that is usually no cause for alarm. Symptoms of hRSV usually resemble those of the common cold and can go undiagnosed. However, infants as well as the elderly are at risk for developing severe cases, which can lead to high morbidity and mortality rates especially if there are underlying health issues. Despite many years of effort, no vaccine or specific treatments exist and RSV is still the leading cause of infant hospitalizations worldwide. Here, we describe methods to infect two widely used small animal models: laboratory mice and cotton rats. PMID:27464697

  8. The pathogenesis of respiratory syncytial virus infection in infant ferrets.

    PubMed Central

    Prince, G. A.; Porter, D. D.

    1976-01-01

    The infant ferret is susceptible to respiratory syncytial virus infection in both the upper and lower respiratory tracts. In the nose, viral replication is restricted to the surface respiratory epithelium in the nasal passages and turbinates. In the lungs, viral replication is of a lower order of magnitude and is localized in the alveolar cells. The pattern of viral replication in nasal tissues is independent of the age of the animal at infection, whereas the pattern in lung tissues shows a striking age dependence, with viral replication progressively decreasing as a function of age. Thes age dependence appears to be due to an intrinsic age-related mechanism yet to be defined. We feel that the infant ferret is an acceptable model for the study of respiratory syncytial virus disease and that the study of age dependence observed in ferrets may allow elucidation of the mechanisms involved in the age dependence seen in humans. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:1251889

  9. Existence of periodic solutions in a model of respiratory syncytial virus RSV

    NASA Astrophysics Data System (ADS)

    Arenas, Abraham J.; González, Gilberto; Jódar, Lucas

    2008-08-01

    In this paper we study the existence of a positive periodic solutions for nested models of respiratory syncytial virus RSV, by using a continuation theorem based on coincidence degree theory. Conditions for the existence of periodic solutions in the model are given. Numerical simulations related to the transmission of respiratory syncytial virus in Madrid and Rio Janeiro are included.

  10. Architecture of respiratory syncytial virus revealed by electron cryotomography

    PubMed Central

    Liljeroos, Lassi; Krzyzaniak, Magdalena Anna; Helenius, Ari; Butcher, Sarah Jane

    2013-01-01

    Human respiratory syncytial virus is a human pathogen that causes severe infection of the respiratory tract. Current information about the structure of the virus and its interaction with host cells is limited. We carried out an electron cryotomographic characterization of cell culture-grown human respiratory syncytial virus to determine the architecture of the virion. The particles ranged from 100 nm to 1,000 nm in diameter and were spherical, filamentous, or a combination of the two. The filamentous morphology correlated with the presence of a cylindrical matrix protein layer linked to the inner leaflet of the viral envelope and with local ordering of the glycoprotein spikes. Recombinant viruses with only the fusion protein in their envelope showed that these glycoproteins were predominantly in the postfusion conformation, but some were also in the prefusion form. The ribonucleocapsids were left-handed, randomly oriented, and curved inside the virions. In filamentous particles, they were often adjacent to an intermediate layer of protein assigned to M2-1 (an envelope-associated protein known to mediate association of ribonucleocapsids with the matrix protein). Our results indicate important differences in structure between the Paramyxovirinae and Pneumovirinae subfamilies within the Paramyxoviridae, and provide fresh insights into host cell exit of a serious pathogen. PMID:23776214

  11. The nosocomial spread of respiratory syncytial viral infections.

    PubMed

    Hall, C B

    1983-01-01

    Respiratory syncytial virus is a regular winter visitor that is highly contagious among persons of all ages. It is the major nosocomial pathogen on infant and toddler wards, and has recently been recognized as also causing appreciable nosocomial illness in the elderly. Control of the spread of this virus has been difficult, but transmission appears to require close contact via large-particle aerosols or via fomites. Environmental conditions affect the survival of the virus on varying surfaces and skin, but self-inoculation after touching contaminated surfaces appears to be an important mode of transmission.

  12. Vaccines against respiratory syncytial virus: The time has finally come.

    PubMed

    Graham, Barney S

    2016-06-24

    Respiratory syncytial virus causes a significant public health burden, particularly in very young infants and the frail elderly. The legacy of enhanced RSV disease (ERD) from a whole formalin-inactivated RSV vaccine, and the complex biology of the virus and the neonate have delayed the development of effective vaccines. However, new insights into factors associated with ERD and breakthroughs in understanding the antigenic structure of the fusion (F) glycoprotein have increased optimism that vaccine development is possible. This has led to investment of time and resources by industry, regulatory authorities, governments, and nonprofit organizations to develop the infrastructure needed to make the advanced clinical development of RSV vaccine candidates a reality.

  13. Preventing respiratory syncytial virus in homebound premature infants.

    PubMed

    Austin, Jennifer

    2007-01-01

    This article explores the home health nurse's role in preventing respiratory syncytial virus (RSV) among premature infants. Thousands of children infected with RSV require hospitalization each year. Consistent contact with the infant alerts the nurse to subtle signs and symptoms of RSV infection, which may include nasal congestion, cough, low-grade fever, and malaise. By developing patient and caregiver trust, the home health nurse can implement an RSV prevention plan, leading to a decrease in hospitalization episodes of premature infants with RSV. Identification of patient risk factors contributing to RSV together with caregiver education is addressed in this article.

  14. Screening for Respiratory Syncytial Virus and Isolation Strategies in Children Hospitalized With acute Respiratory Tract Infection

    PubMed Central

    Pfeil, Johannes; Tabatabai, Julia; Sander, Anja; Ries, Markus; Grulich-Henn, Jürgen; Schnitzler, Paul

    2014-01-01

    Abstract Nosocomial infection with respiratory syncytial virus (RSV) is an important health risk in pediatric care but is largely preventable by efficient infection control measures. Commonly applied rapid antigen detection tests (RADTs) miss a considerable number of RSV-infected patients. The objective of our analysis was to evaluate whether readily available host parameters are associated with false-negative RADT, and to assess how these parameters could be applied in an optimized RSV isolation strategy. We retrospectively analyzed a cohort of 242 children under the age of 2 years hospitalized with acute respiratory tract infection to identify host parameters associated with false-negative RADT test result. We subsequently simulated the outcome of different isolation strategies based on RADT result and host parameters in view of the overall isolation efficacy. Out of 242 hospitalized patients, 134 (55%) patients were found RSV-positive by RT-PCR, whereas 108 (45%) patients were tested negative. The performance of the RADT was compared with the result obtained by reverse transcription polymerase chain reaction on the identical nasopharyngeal wash. Overall, we found that 85 patients (35%) were tested true positive, 108 (45%) were tested true negative, whereas a false-negative test result was obtained in 49 patients (20%). Duration of respiratory symptoms for >3 days and a respiratory admission diagnosis are associated with false-negative RADT result. In comparison with RADT alone, consideration of these clinical parameters and RADT result can decrease the rate of nonisolated RSV-infected patients from approximately 24% to 8% (65% RSV pretest probability). Consideration of both RADT and clinical parameters associated with false-negative RADT can result in an optimized RSV infection control policy. PMID:25437026

  15. Animal models of human respiratory syncytial virus disease

    PubMed Central

    Domachowske, Joseph B.; Rosenberg, Helene F.

    2011-01-01

    Infection with the human pneumovirus pathogen, respiratory syncytial virus (hRSV), causes a wide spectrum of respiratory disease, notably among infants and the elderly. Laboratory animal studies permit detailed experimental modeling of hRSV disease and are therefore indispensable in the search for novel therapies and preventative strategies. Present animal models include several target species for hRSV, including chimpanzees, cattle, sheep, cotton rats, and mice, as well as alternative animal pneumovirus models, such as bovine RSV and pneumonia virus of mice. These diverse animal models reproduce different features of hRSV disease, and their utilization should therefore be based on the scientific hypothesis under investigation. The purpose of this review is to summarize the strengths and limitations of each of these animal models. Our intent is to provide a resource for investigators and an impetus for future research. PMID:21571908

  16. Characterization of an Experimental Vaccine for Bovine Respiratory Syncytial Virus

    PubMed Central

    Hu, Kefei; Blodörn, Krister; Makabi-Panzu, Boby; Gaillard, Anne-Laure; Ellencrona, Karin; Chevret, Didier; Hellman, Lars; Bengtsson, Karin Lövgren; Riffault, Sabine; Taylor, Geraldine; Valarcher, Jean François; Eléouët, Jean-François

    2014-01-01

    Bovine respiratory syncytial virus (BRSV) and human respiratory syncytial virus (HRSV) are major causes of respiratory disease in calves and children, respectively, and are priorities for vaccine development. We previously demonstrated that an experimental vaccine, BRSV-immunostimulating complex (ISCOM), is effective in calves with maternal antibodies. The present study focuses on the antigenic characterization of this vaccine for the design of new-generation subunit vaccines. The results of our study confirmed the presence of membrane glycoprotein (G), fusion glycoprotein (F), and nucleoprotein (N) proteins in the ISCOMs, and this knowledge was extended by the identification of matrix (M), M2-1, phosphoprotein (P), small hydrophobic protein (SH) and of cellular membrane proteins, such as the integrins αVβ1, αVβ3, and α3β1. The quantity of the major protein F was 4- to 5-fold greater than that of N (∼77 μg versus ∼17 μg/calf dose), whereas G, M, M2-1, P, and SH were likely present in smaller amounts. The polymerase (L), M2-2, nonstructural 1 (NS1), and NS2 proteins were not detected, suggesting that they are not essential for protection. Sera from the BRSV-ISCOM-immunized calves contained high titers of IgG antibody specific for F, G, N, and SH. Antibody responses against M and P were not detected; however, this does not exclude their role in protective T-cell responses. The absence of immunopathological effects of the cellular proteins, such as integrins, needs to be further confirmed, and their possible contribution to adjuvant functions requires elucidation. This work suggests that a combination of several surface and internal proteins should be included in subunit RSV vaccines and identifies absent proteins as potential candidates for differentiating infected from vaccinated animals. PMID:24828093

  17. Molecular mechanism of respiratory syncytial virus fusion inhibitors.

    PubMed

    Battles, Michael B; Langedijk, Johannes P; Furmanova-Hollenstein, Polina; Chaiwatpongsakorn, Supranee; Costello, Heather M; Kwanten, Leen; Vranckx, Luc; Vink, Paul; Jaensch, Steffen; Jonckers, Tim H M; Koul, Anil; Arnoult, Eric; Peeples, Mark E; Roymans, Dirk; McLellan, Jason S

    2016-02-01

    Respiratory syncytial virus (RSV) is a leading cause of pneumonia and bronchiolitis in young children and the elderly. Therapeutic small molecules have been developed that bind the RSV F glycoprotein and inhibit membrane fusion, yet their binding sites and molecular mechanisms of action remain largely unknown. Here we show that these inhibitors bind to a three-fold-symmetric pocket within the central cavity of the metastable prefusion conformation of RSV F. Inhibitor binding stabilizes this conformation by tethering two regions that must undergo a structural rearrangement to facilitate membrane fusion. Inhibitor-escape mutations occur in residues that directly contact the inhibitors or are involved in the conformational rearrangements required to accommodate inhibitor binding. Resistant viruses do not propagate as well as wild-type RSV in vitro, indicating a fitness cost for inhibitor escape. Collectively, these findings provide new insight into class I viral fusion proteins and should facilitate development of optimal RSV fusion inhibitors.

  18. Vaccines against respiratory syncytial virus: The time has finally come.

    PubMed

    Graham, Barney S

    2016-06-24

    Respiratory syncytial virus causes a significant public health burden, particularly in very young infants and the frail elderly. The legacy of enhanced RSV disease (ERD) from a whole formalin-inactivated RSV vaccine, and the complex biology of the virus and the neonate have delayed the development of effective vaccines. However, new insights into factors associated with ERD and breakthroughs in understanding the antigenic structure of the fusion (F) glycoprotein have increased optimism that vaccine development is possible. This has led to investment of time and resources by industry, regulatory authorities, governments, and nonprofit organizations to develop the infrastructure needed to make the advanced clinical development of RSV vaccine candidates a reality. PMID:27182820

  19. 4EBP1 Is Dephosphorylated by Respiratory Syncytial Virus Infection.

    PubMed

    Pérez-Gil, Gustavo; Landa-Cardeña, Adriana; Coutiño, Rocío; García-Román, Rebeca; Sampieri, Clara L; Mora, Silvia I; Montero, Hilda

    2015-01-01

    Respiratory syncytial virus (RSV) requires protein biosynthesis machinery to generate progeny. There is evidence that RSV might alter some translation components since stress granules are formed in their host cells. Consistent with these observations, we found that RSV induces dephosphorylation of 4EBP1 (eIF4E-binding protein), an important cellular translation factor. Our results show no correlation between the 4EBP1 dephosphorylation time and the decrease in the global rate of protein synthesis. Interestingly, treatment with rapamycin stimulates virus generation. The results suggest that RSV is a virus that still contains unknown mechanisms involved in the translation of their mRNAs through the alteration or modification of some translation factors, such as 4EBP1, possibly to favor its replicative cycle.

  20. Respiratory syncytial virus: its transmission in the hospital environment.

    PubMed

    Hall, C B

    1982-01-01

    Respiratory syncytial virus (RSV) over the past two decades has been recognized as the most important cause of lower respiratory tract disease in infants and young children. Recently, it has also been identified as a major nosocomial hazard on pediatric wards. The potential for RSV to spread on such wards is underlined by several singular characteristics of RSV. It arrives in yearly epidemics and is highly contagious in all age groups. Immunity is of short duration, allowing repeated infections to occur. Thus, during an epidemic 20--40 percent of infants admitted for other conditions may acquire nosocomial RSV infection, as well as 50 percent of the ward personnel. The usual infection control procedures for respiratory illnesses have had limited success in controlling the spread of RSV. This may be due in part to the modes of transmission of RSV. Inoculation occurs mainly through the eye and nose, rather than the mouth. This may be via large-particle aerosols or droplets, requiring close contact. The virus, however, does not seem capable of traversing distances by small-particle aerosols. Nevertheless, it is able to remain infectious on various environmental surfaces, suggesting fomites as a source of spread. Indeed, inoculation after touching such contaminated surfaces can occur, and may be a major second means of spread, in hospitals as well as in families.

  1. Respiratory Syncytial Virus Infections in Infants Affected by Primary Immunodeficiency

    PubMed Central

    Capretti, Maria Grazia; Lazzarotto, Tiziana; Faldella, Giacomo

    2014-01-01

    Primary immunodeficiencies are rare inherited disorders that may lead to frequent and often severe acute respiratory infections. Respiratory syncytial virus (RSV) is one of the most frequent pathogens during early infancy and the infection is more severe in immunocompromised infants than in healthy infants, as a result of impaired T- and B-cell immune response unable to efficaciously neutralize viral replication, with subsequent increased viral shedding and potentially lethal lower respiratory tract infection. Several authors have reported a severe clinical course after RSV infections in infants and children with primary and acquired immunodeficiencies. Environmental prophylaxis is essential in order to reduce the infection during the epidemic season in hospitalized immunocompromised infants. Prophylaxis with palivizumab, a humanized monoclonal antibody against the RSV F protein, is currently recommended in high-risk infants born prematurely, with chronic lung disease or congenital heart disease. Currently however the prophylaxis is not routinely recommended in infants with primary immunodeficiency, although some authors propose the extension of prophylaxis to this high risk population. PMID:25089282

  2. RSV604, a Novel Inhibitor of Respiratory Syncytial Virus Replication▿

    PubMed Central

    Chapman, Joanna; Abbott, Elizabeth; Alber, Dagmar G.; Baxter, Robert C.; Bithell, Sian K.; Henderson, Elisa A.; Carter, Malcolm C.; Chambers, Phil; Chubb, Ann; Cockerill, G. Stuart; Collins, Peter L.; Dowdell, Verity C. L.; Keegan, Sally J.; Kelsey, Richard D.; Lockyer, Michael J.; Luongo, Cindy; Najarro, Pilar; Pickles, Raymond J.; Simmonds, Mark; Taylor, Debbie; Tyms, Stan; Wilson, Lara J.; Powell, Kenneth L.

    2007-01-01

    Respiratory syncytial virus (RSV) is the most common cause of lower respiratory tract infections worldwide, yet no effective vaccine or antiviral treatment is available. Here we report the discovery and initial development of RSV604, a novel benzodiazepine with submicromolar anti-RSV activity. It proved to be equipotent against all clinical isolates tested of both the A and B subtypes of the virus. The compound has a low rate of in vitro resistance development. Sequencing revealed that the resistant virus had mutations within the nucleocapsid protein. This is a novel mechanism of action for anti-RSV compounds. In a three-dimensional human airway epithelial cell model, RSV604 was able to pass from the basolateral side of the epithelium effectively to inhibit virus replication after mucosal inoculation. RSV604, which is currently in phase II clinical trials, represents the first in a new class of RSV inhibitors and may have significant potential for the effective treatment of RSV disease. PMID:17576833

  3. Epidemiology, Molecular Epidemiology and Evolution of Bovine Respiratory Syncytial Virus

    PubMed Central

    Sarmiento-Silva, Rosa Elena; Nakamura-Lopez, Yuko; Vaughan, Gilberto

    2012-01-01

    The bovine respiratory syncytial virus (BRSV) is an enveloped, negative sense, single-stranded RNA virus belonging to the pneumovirus genus within the family Paramyxoviridae. BRSV has been recognized as a major cause of respiratory disease in young calves since the early 1970s. The analysis of BRSV infection was originally hampered by its characteristic lability and poor growth in vitro. However, the advent of numerous immunological and molecular methods has facilitated the study of BRSV enormously. The knowledge gained from these studies has also provided the opportunity to develop safe, stable, attenuated virus vaccine candidates. Nonetheless, many aspects of the epidemiology, molecular epidemiology and evolution of the virus are still not fully understood. The natural course of infection is rather complex and further complicates diagnosis, treatment and the implementation of preventive measures aimed to control the disease. Therefore, understanding the mechanisms by which BRSV is able to establish infection is needed to prevent viral and disease spread. This review discusses important information regarding the epidemiology and molecular epidemiology of BRSV worldwide, and it highlights the importance of viral evolution in virus transmission. PMID:23202546

  4. Respiratory syncytial virus infection increases chlorine-induced airway hyperresponsiveness.

    PubMed

    Song, Weifeng; Yu, Zhihong; Doran, Stephen F; Ambalavanan, Namasivayam; Steele, Chad; Garantziotis, Stavros; Matalon, Sadis

    2015-08-01

    Exposure to chlorine (Cl2) damages airway and alveolar epithelia resulting in acute lung injury and reactive airway hyperresponsiveness (AHR) to methacholine. However, little is known about the effect of preexisting respiratory disease on Cl2-induced lung injury. By using a murine respiratory syncytial virus (RSV) infection model, we found that preexisting RSV infection increases Cl2 (187 ppm for 30 min)-induced lung inflammation and airway AHR at 24 h after exposure (5 days after infection). RSV infection and Cl2 exposure synergistically induced oxygen desaturation and neutrophil infiltration and increased MCP-1, MIP-1β, IL-10, IFN-γ, and RANTES concentrations in the bronchoalveolar lavage fluid (BALF). In contrast, levels of type 2 cytokines (i.e., IL-4, IL-5, IL-9, and IL-13) were not significantly affected by either RSV infection or Cl2 exposure. Cl2 exposure, but not RSV infection, induced AHR to methacholine challenge as measured by flexiVent. Moreover, preexisting RSV infection amplified BALF levels of hyaluronan (HA) and AHR. The Cl2-induced AHR was mitigated by treatment with inter-α-trypsin inhibitor antibody, which inhibits HA signaling, suggesting a mechanism of HA-mediated AHR from exacerbated oxidative injury. Our results show for the first time that preexisting RSV infection predisposes the lung to Cl2-induced injury. These data emphasize the necessity for further research on the effects of Cl2 in vulnerable populations and the development of appropriate treatments.

  5. Epidemiology of Respiratory Syncytial Virus Infection in Preterm Infants

    PubMed Central

    Resch, Bernhard; Kurath, Stefan; Manzoni, Paolo

    2011-01-01

    This review focuses on the burden of respiratory syncytial virus (RSV) infection in preterm infants with and without chronic lung disease (bronchopulmonary dysplasia, BPD). The year-to-year and seasonal variations in RSV activity are key aspects of RSV epidemiology, and knowledge/monitoring of local RSV activity is mandatory for guidance of prophylaxis with the monoclonal antibodies palivizumab and in the near future motavizumab. Morbidity expressed in rates of hospitalizations attributable to RSV illness revealed a mean of 10 percent in preterm infants without and 19 percent (p=0.016) with BPD. Mortality rates diverged widely, and case fatality rates have been reported to range from 0 to 12 percent. The typical clinical picture of lower respiratory tract infection is not different in term and preterm infants, but rates of apnoeas are significantly increased in preterms, ranging from 4.9 to 37.5 percent with decreasing rates observed in more recent studies. Until a RSV vaccine is developed and will be available, prophylaxis with palivizumab is the only preventative strategy other than hand hygiene and contact measures that significantly reduces RSV hospitalization rates in preterm infants both with and without BPD. PMID:22262986

  6. Human and bovine respiratory syncytial virus vaccine research and development.

    PubMed

    Meyer, Gilles; Deplanche, Martine; Schelcher, François

    2008-03-01

    Human (HRSV) and bovine (BRSV) respiratory syncytial viruses (RSV) are two closely related viruses, which are the most important causative agents of respiratory tract infections of young children and calves, respectively. BRSV vaccines have been available for nearly 2 decades. They probably have reduced the prevalence of RSV infection but their efficacy needs improvement. In contrast, despite decades of research, there is no currently licensed vaccine for the prevention of HRSV disease. Development of a HRSV vaccine for infants has been hindered by the lack of a relevant animal model that develops disease, the need to immunize immunologically immature young infants, the difficulty for live vaccines to find the right balance between attenuation and immunogenicity, and the risk of vaccine-associated disease. During the past 15 years, intensive research into a HRSV vaccine has yielded vaccine candidates, which have been evaluated in animal models and, for some of them, in clinical trials in humans. Recent formulations have focused on subunit vaccines with specific CD4+ Th-1 immune response-activating adjuvants and on genetically engineered live attenuated vaccines. It is likely that different HRSV vaccines and/or combinations of vaccines used sequentially will be needed for the various populations at risk. This review discusses the recent advances in RSV vaccine development. PMID:17720245

  7. Noninvasive ventilation in acute respiratory failure from respiratory syncytial virus bronchiolitis

    PubMed Central

    Nizarali, Zahara; Cabral, Marta; Silvestre, Catarina; Abadesso, Clara; Nunes, Pedro; Loureiro, Helena; Almeida, Helena

    2012-01-01

    Objectives The present study focused on respiratory syncytial virus bronchiolitis with respiratory failure. The aim of the study was to determine whether noninvasive ventilation reduces the need for endotracheal intubation or slows the clinical progression of acute respiratory syncytial virus bronchiolitis by reducing the incidence of infectious complications. Methods The present study was a retrospective cohort study. Cohort A was comprised of children who were admitted to the pediatric intensive and special care unit from 2003-2005 before starting noninvasive ventilation; cohort B was comprised of children who were admitted to the pediatric intensive and special care unit from 2006-2008 after starting noninvasive ventilation. With the exception of noninvasive ventilation, the therapeutic support was the same for the two groups. All children who were diagnosed with respiratory syncytial virus bronchiolitis and respiratory failure between November 2003 and March 2008 were included in the cohort. Demographic, clinical and blood gas variables were analyzed. Results A total of 162 children were included; 75% of the subjects were less than 3 months old. Group A included 64 children, and group B included 98 children. In group B, 34 of the children required noninvasive ventilation. The distributions of the variables age, preterm birth, congenital heart disease, cerebral palsy and chronic lung disease were similar between the two groups. On admission, the data for blood gas analysis and the number of apneas were not significantly different between the groups. In group B, fewer children required invasive ventilation (group A: 12/64 versus group B: 7/98; p=0.02), and there was a reduction in the number of cases of bacterial pneumonia (group A: 19/64 versus group B: 12/98; p=0.008). There was no record of mortality in either of the groups. Conclusion By comparing children with the same disease both before and after noninvasive ventilation was used for ventilation support, we

  8. The pharmacologic mechanism by which inhaled epinephrine reduces airway obstruction in respiratory syncytial virus-associated bronchiolitis.

    PubMed

    Barr, F E; Patel, N R; Newth, C J

    2000-05-01

    Inhaled racemic epinephrine relieves airway obstruction in patients with respiratory syncytial virus bronchiolitis. The contribution of alpha- versus beta-adrenoreceptor stimulation toward this clinical effect is unknown. We describe an infant treated with propranolol for supraventricular tachycardia in whom respiratory syncytial virus bronchiolitis developed. Inhaled racemic epinephrine improved his respiratory symptoms, whereas nebulized albuterol did not.

  9. Risk Factors for Respiratory Failure Associated with Respiratory Syncytial Virus Infection in Adults

    PubMed Central

    Duncan, Coley B.; Walsh, Edward E.; Peterson, Derick R.; Lee, F.Eun-Hyung; Falsey, Ann R.

    2010-01-01

    Risk factors associated with respiratory failure during respiratory syncytial virus (RSV) infection have not been assessed in adults. We identified RSV by quantitative reverse transcription polymerase chain reaction in 58 adults during the 2007–2008 winter. Clinical variables and respiratory secretion viral loads were compared in 26 outpatients and 32 inpatients. Cardiopulmonary diseases were more common among inpatients than outpatients (91% vs 31%, P = .0001), whereas mean RSV load was similar. Nasal viral load was higher in ventilated vs nonventilated hospitalized patients (log10 3.7 ± 1.7 plaque-forming units (PFUs)/mL vs 2.4 ± 1.1 PFUs/mL, P = .02), and high viral load was independently associated with respiratory failure. PMID:19758094

  10. ISG15 Is Upregulated in Respiratory Syncytial Virus Infection and Reduces Virus Growth through Protein ISGylation

    PubMed Central

    González-Sanz, Rubén; Mata, Manuel; Bermejo-Martín, Jesús; Álvarez, Amparo; Cortijo, Julio; Melero, José A.

    2016-01-01

    ABSTRACT Human respiratory syncytial virus (RSV), for which neither a vaccine nor an effective therapeutic treatment is currently available, is the leading cause of severe lower respiratory tract infections in children. Interferon-stimulated gene 15 (ISG15) is a ubiquitin-like protein that is highly increased during viral infections and has been reported to have an antiviral or a proviral activity, depending on the virus. Previous studies from our laboratory demonstrated strong ISG15 upregulation during RSV infection in vitro. In this study, an in-depth analysis of the role of ISG15 in RSV infection is presented. ISG15 overexpression and small interfering RNA (siRNA)-silencing experiments, along with ISG15 knockout (ISG15−/−) cells, revealed an anti-RSV effect of the molecule. Conjugation inhibition assays demonstrated that ISG15 exerts its antiviral activity via protein ISGylation. This antiviral activity requires high levels of ISG15 to be present in the cells before RSV infection. Finally, ISG15 is also upregulated in human respiratory pseudostratified epithelia and in nasopharyngeal washes from infants infected with RSV, pointing to a possible antiviral role of the molecule in vivo. These results advance our understanding of the innate immune response elicited by RSV and open new possibilities to control infections by the virus. IMPORTANCE At present, no vaccine or effective treatment for human respiratory syncytial virus (RSV) is available. This study shows that interferon-stimulated gene 15 (ISG15) lowers RSV growth through protein ISGylation. In addition, ISG15 accumulation highly correlates with the RSV load in nasopharyngeal washes from children, indicating that ISG15 may also have an antiviral role in vivo. These results improve our understanding of the innate immune response to RSV and identify ISG15 as a potential target for virus control. PMID:26763998

  11. Effect of climatological factors on respiratory syncytial virus epidemics.

    PubMed

    Noyola, D E; Mandeville, P B

    2008-10-01

    Respiratory syncytial virus (RSV) presents as yearly epidemics in temperate climates. We analysed the association of atmospheric conditions to RSV epidemics in San Luis Potosí, S.L.P., Mexico. The weekly number of RSV detections from October 2002 and May 2006 were correlated to ambient temperature, barometric pressure, relative humidity, vapour tension, dew point, precipitation, and hours of light using time-series and regression analyses. Of the variation in RSV cases, 49.8% was explained by the study variables. Of the explained variation in RSV cases, 32.5% was explained by the study week and 17.3% was explained by meteorological variables (average daily temperature, maximum daily temperature, temperature at 08:00 hours, and relative humidity at 08:00 hours). We concluded that atmospheric conditions, particularly temperature, partly explain the year to year variability in RSV activity. Identification of additional factors that affect RSV seasonality may help develop a model to predict the onset of RSV epidemics. PMID:18177520

  12. Growth of respiratory syncytial virus in mink lung epithelial cells.

    PubMed

    Yeolekar, L R; Damle, R G; Basu, A; Rao, B L

    2002-12-01

    Mink lung epithelial cells (Mv-1-Lu) were tested for their ability to support the growth and serial passage of respiratory syncytial virus (RSV) in vitro. Indian isolates of RSV induced distinctive cytopathic effect with typical rounding of cells followed by detachment with more than 50 per cent cells showing bright fluorescence using anti-RSV monoclonal antibodies in immunofluorescence test. Serial passage of RSV was possible in Mv-1-Lu cells without loss of sensitivity of the cells for virus growth. Titration of cell associated virus and virus released in the supernatant indicated that 60 per cent of the virus was released in the supernatant, and 40 per cent remained cell associated. Transmission electron microscopic studies of negatively stained RSV particles and ultra-thin sections of RSV infected Mv-1-Lu cells showed roughly spherical particles with club shaped projections, budding from the cytoplasmic membrane. These results indicate that Mv-1-Lu cell line is suitable for the growth and propagation of RSV.

  13. Possible transmission by fomites of respiratory syncytial virus.

    PubMed

    Hall, C B; Douglas, R G; Geiman, J M

    1980-01-01

    To test whether nosocomial spread of respiratory syncytial virus (RSV) could occur through contact with environmental surfaces contaminated by RSV-infected nasal secretions, survival in the environment of RSV isolated from media, pooled adult secretions, and secretions from hospitalized infants was examined. RSV in freshly obtained infant secretions was recovered from countertops for up to 6 hr, from rubber gloves for up to 1 1/2 hr, from cloth gowns and paper tissue for 30--45 min, and from skin for up to 20 min. RSV in media and pooled secretions survived for slightly longer periods. Further experiments demonstrated that infectious virus could be transferred to hands touching these contaminated surfaces and could be recovered from these hands for up to 25 min. These studies suggest that survival of RSV in the environment of infected infant secretions is sufficient to allow transfer of infectious virus to the hands of hospital personnel. Thus, self-inoculation by contact with contaminated infant secretions may be a potential mode of nosocomial transmission of RSV.

  14. Brief History and Characterization of Enhanced Respiratory Syncytial Virus Disease.

    PubMed

    Acosta, Patricio L; Caballero, Mauricio T; Polack, Fernando P

    2016-03-01

    In 1967, infants and toddlers immunized with a formalin-inactivated vaccine against respiratory syncytial virus (RSV) experienced an enhanced form of RSV disease characterized by high fever, bronchopneumonia, and wheezing when they became infected with wild-type virus in the community. Hospitalizations were frequent, and two immunized toddlers died upon infection with wild-type RSV. The enhanced disease was initially characterized as a "peribronchiolar monocytic infiltration with some excess in eosinophils." Decades of research defined enhanced RSV disease (ERD) as the result of immunization with antigens not processed in the cytoplasm, resulting in a nonprotective antibody response and CD4(+) T helper priming in the absence of cytotoxic T lymphocytes. This response to vaccination led to a pathogenic Th2 memory response with eosinophil and immune complex deposition in the lungs after RSV infection. In recent years, the field of RSV experienced significant changes. Numerous vaccine candidates with novel designs and formulations are approaching clinical trials, defying our previous understanding of favorable parameters for ERD. This review provides a succinct analysis of these parameters and explores criteria for assessing the risk of ERD in new vaccine candidates. PMID:26677198

  15. Evaluation of the Adaptive Immune Response to Respiratory Syncytial Virus.

    PubMed

    Knudson, Cory J; Weiss, Kayla A; Stoley, Megan E; Varga, Steven M

    2016-01-01

    Evaluation of the adaptive immune response is critical to the advancement of our basic knowledge and understanding of respiratory syncytial virus (RSV). The cellular composition in the lung following RSV infection is often evaluated using flow cytometry. However, a limitation of this approach has been the inability to readily distinguish cells that are within the lung parenchyma from cells that remain in the pulmonary blood vessels. Herein, we detail a procedure to evaluate the adaptive immune response via flow cytometric analysis that incorporates an in vivo intravascular staining technique. This technique allows for discrimination of immune cells in the lung tissue from cells that remain in the pulmonary vasculature following perfusion. Therefore at any given time point following an RSV infection, the leukocytic populations in the lung parenchyma can be quantified and phenotypically assessed with high resolution. While we focus on the T lymphocyte response in the lung, this technique can be readily adapted to examine various leukocytic cell types in the lung following RSV infection. PMID:27464699

  16. Challenges and Opportunities in Developing Respiratory Syncytial Virus Therapeutics

    PubMed Central

    Simões, Eric A. F.; DeVincenzo, John P.; Boeckh, Michael; Bont, Louis; Crowe, James E.; Griffiths, Paul; Hayden, Frederick G.; Hodinka, Richard L.; Smyth, Rosalind L.; Spencer, Keith; Thirstrup, Steffen; Walsh, Edward E.; Whitley, Richard J.

    2015-01-01

    Two meetings, one sponsored by the Wellcome Trust in 2012 and the other by the Global Virology Foundation in 2013, assembled academic, public health and pharmaceutical industry experts to assess the challenges and opportunities for developing antivirals for the treatment of respiratory syncytial virus (RSV) infections. The practicalities of clinical trials and establishing reliable outcome measures in different target groups were discussed in the context of the regulatory pathways that could accelerate the translation of promising compounds into licensed agents. RSV drug development is hampered by the perceptions of a relatively small and fragmented market that may discourage major pharmaceutical company investment. Conversely, the public health need is far too large for RSV to be designated an orphan or neglected disease. Recent advances in understanding RSV epidemiology, improved point-of-care diagnostics, and identification of candidate antiviral drugs argue that the major obstacles to drug development can and will be overcome. Further progress will depend on studies of disease pathogenesis and knowledge provided from controlled clinical trials of these new therapeutic agents. The use of combinations of inhibitors that have different mechanisms of action may be necessary to increase antiviral potency and reduce the risk of resistance emergence. PMID:25713060

  17. Respiratory syncytial virus prophylaxis--the story so far.

    PubMed

    Simoes, E A F; Groothuis, J R

    2002-04-01

    Respiratory syncytial virus (RSV) is a common and highly contagious pathogen that infects nearly all children by the age of 2 years. It is responsible for significant morbidity and mortality worldwide among certain high-risk paediatric populations. Therapy is sub-optimal for RSV, thus treatment focuses on ameliorating symptoms. Since discovery of the virus in the 1950s, efforts have been ongoing to develop a safe and effective vaccine. These efforts have met with serious obstacles. Passive immunoprophylaxis presents a viable alternative to active immunization. In 1998, the genetically engineered humanized monoclonal antibody (palivizumab) was granted FDA (Food and Drug Administration) approval for prophylaxis of high-risk children in the United States; EMEA (European Agency for the Evaluation of Medicinal Products) approval followed in 1999 for Europe. It is now approved in over 45 countries worldwide. Palivizumab was shown to significantly reduce RSV-related hospitalizations in North America and Europe with few adverse effects. Clinical trial and outcomes data documenting experience with palivizumab to date continue to extend the initial safety and efficacy observations. PMID:11996400

  18. Effect of climatological factors on respiratory syncytial virus epidemics

    PubMed Central

    NOYOLA, D. E.; MANDEVILLE, P. B.

    2008-01-01

    SUMMARY Respiratory syncytial virus (RSV) presents as yearly epidemics in temperate climates. We analysed the association of atmospheric conditions to RSV epidemics in San Luis Potosí, S.L.P., Mexico. The weekly number of RSV detections between October 2002 and May 2006 were correlated to ambient temperature, barometric pressure, relative humidity, vapour tension, dew point, precipitation, and hours of light using time-series and regression analyses. Of the variation in RSV cases, 49·8% was explained by the study variables. Of the explained variation in RSV cases, 32·5% was explained by the study week and 17·3% was explained by meteorological variables (average daily temperature, maximum daily temperature, temperature at 08:00 hours, and relative humidity at 08:00 hours). We concluded that atmospheric conditions, particularly temperature, partly explain the year to year variability in RSV activity. Identification of additional factors that affect RSV seasonality may help develop a model to predict the onset of RSV epidemics. PMID:18177520

  19. Respiratory syncytial virus envelope glycoprotein (G) has a novel structure.

    PubMed Central

    Satake, M; Coligan, J E; Elango, N; Norrby, E; Venkatesan, S

    1985-01-01

    Amino acid sequence of human respiratory syncytial virus envelope glycoprotein (G) was deduced from the DNA sequence of a recombinant plasmid and confirmed by limited amino acid microsequencing of purified 90K G protein. The calculated molecular mass of the protein encoded by the only long open reading frame of 298 amino acids was 32,588 daltons and was somewhat smaller than the 36K polypeptide translated in vitro from mRNA selected by this plasmid. Inspection of the sequence revealed a single hydrophobic domain of 23 amino acids capable of membrane insertion at 41 residues from the N-terminus. There was no N-terminal signal sequence and the hydrophilic N-terminal 20 residues probably represent the cytoplasmic tail of the protein. The N-terminally oriented membrane insertion was somewhat analogous to paramyxovirus hemagglutinin-neuraminidase (HN) and influenza neuraminidase (NA). The protein was moderately hydrophilic and rich in hydroxy-amino acids. It was both N- and O-glycosylated with the latter contributing significantly to the net molecular mass 90K. Images PMID:4069997

  20. Simultaneous influenza and respiratory syncytial virus infection in human respiratory tract

    NASA Astrophysics Data System (ADS)

    Pinky, Lubna Jahan Rashid; Dobrovolny, Hana

    2015-03-01

    Studies have shown that simultaneous infection of the respiratory tract with at least two viruses is not uncommon in hospitalized patients, although it is not clear whether these infections are more or less severe than single infections. We use mathematical models to study the dynamics of simultaneous influenza (flu) and respiratory syncytial virus (RSV) infection, two of the more common respiratory viruses, in an effort to understand simultaneous infections. We examine the roles of initial viral inoculum, relative starting time, and cell regeneration on the severity of the infection. We also study the effect of antiviral treatment on the course of the infection. This study shows that, unless treated with antivirals, flu always takes over the infection no matter how small the initial dose and how delayed it starts with respect to RSV.

  1. In search of a small-molecule inhibitor for respiratory syncytial virus.

    PubMed

    Douglas, Janet L

    2004-08-01

    Respiratory syncytial virus has been an ongoing health problem for 50 years. Hospitalization rates due to virus-induced respiratory illness continue to be substantial for infants, small children, the elderly and the immunocompromised. The only currently available treatments are a broad-spectrum antiviral and two immunoprophylactic antibodies, all of which are reserved for high-risk patients. The combination of this limited therapeutic repertoire and the lack of a vaccine clearly demonstrates the need to continue the search for more efficacious and safe agents against respiratory syncytial virus. The following is a review on the current progress of that search.

  2. The Use of Humanized Monoclonal Antibodies for the Prevention of Respiratory Syncytial Virus Infection

    PubMed Central

    Arcuri, Santo; Galletti, Silvia; Faldella, Giacomo

    2013-01-01

    Monoclonal antibodies are widely used both in infants and in adults for several indications. Humanized monoclonal antibodies (palivizumab) have been used for many years for the prevention of respiratory syncytial virus infection in pediatric populations (preterm infants, infants with chronic lung disease or congenital heart disease) at high risk of severe and potentially lethal course of the infection. This drug was reported to be safe, well tolerated and effective to decrease the hospitalization rate and mortality in these groups of infants by several clinical trials. In the present paper we report the development and the current use of monoclonal antibodies for prophylaxis against respiratory syncytial virus. PMID:23840240

  3. Respiratory syncytial virus pneumonia in a lung transplant recipient: case report.

    PubMed

    Doud, J R; Hinkamp, T; Garrity, E R

    1992-01-01

    A 29-year-old man underwent bilateral lung transplantation and received maintenance immunosuppressive therapy. He was readmitted 11 months later with symptoms of cough, sneezing, and rhinorrhea. The physical examination was normal. Laboratory results were significant for a reduction of FEV1 and an interstitial infiltrate on chest films. The patient had recently undergone bronchoscopy for rejection surveillance, and 2 days before admission the bronchoalveolar lavage cultures returned positive for respiratory syncytial virus. The patient was treated with aerosolized ribavirin with complete resolution of symptoms. Respiratory syncytial virus must now be included in the list of pathogens causing pneumonia in the lung transplant recipient. PMID:1540615

  4. Induction and Antagonism of Antiviral Responses in Respiratory Syncytial Virus-Infected Pediatric Airway Epithelium

    PubMed Central

    Villenave, Rémi; Broadbent, Lindsay; Douglas, Isobel; Lyons, Jeremy D.; Coyle, Peter V.; Teng, Michael N.; Tripp, Ralph A.; Heaney, Liam G.; Shields, Michael D.

    2015-01-01

    ABSTRACT Airway epithelium is the primary target of many respiratory viruses. However, virus induction and antagonism of host responses by human airway epithelium remains poorly understood. To address this, we developed a model of respiratory syncytial virus (RSV) infection based on well-differentiated pediatric primary bronchial epithelial cell cultures (WD-PBECs) that mimics hallmarks of RSV disease in infants. RSV is the most important respiratory viral pathogen in young infants worldwide. We found that RSV induces a potent antiviral state in WD-PBECs that was mediated in part by secreted factors, including interferon lambda 1 (IFN-λ1)/interleukin-29 (IL-29). In contrast, type I IFNs were not detected following RSV infection of WD-PBECs. IFN responses in RSV-infected WD-PBECs reflected those in lower airway samples from RSV-hospitalized infants. In view of the prominence of IL-29, we determined whether recombinant IL-29 treatment of WD-PBECs before or after infection abrogated RSV replication. Interestingly, IL-29 demonstrated prophylactic, but not therapeutic, potential against RSV. The absence of therapeutic potential reflected effective RSV antagonism of IFN-mediated antiviral responses in infected cells. Our data are consistent with RSV nonstructural proteins 1 and/or 2 perturbing the Jak-STAT signaling pathway, with concomitant reduced expression of antiviral effector molecules, such as MxA/B. Antagonism of Jak-STAT signaling was restricted to RSV-infected cells in WD-PBEC cultures. Importantly, our study provides the rationale to further explore IL-29 as a novel RSV prophylactic. IMPORTANCE Most respiratory viruses target airway epithelium for infection and replication, which is central to causing disease. However, for most human viruses we have a poor understanding of their interactions with human airway epithelium. Respiratory syncytial virus (RSV) is the most important viral pathogen of young infants. To help understand RSV interactions with pediatric

  5. Inactivation of respiratory syncytial virus by zinc finger reactive compounds

    PubMed Central

    2010-01-01

    Background Infectivity of retroviruses such as HIV-1 and MuLV can be abrogated by compounds targeting zinc finger motif in viral nucleocapsid protein (NC), involved in controlling the processivity of reverse transcription and virus infectivity. Although a member of a different viral family (Pneumoviridae), respiratory syncytial virus (RSV) contains a zinc finger protein M2-1 also involved in control of viral polymerase processivity. Given the functional similarity between the two proteins, it was possible that zinc finger-reactive compounds inactivating retroviruses would have a similar effect against RSV by targeting RSV M2-1 protein. Moreover, inactivation of RSV through modification of an internal protein could yield a safer whole virus vaccine than that produced by RSV inactivation with formalin which modifies surface proteins. Results Three compounds were evaluated for their ability to reduce RSV infectivity: 2,2'-dithiodipyridine (AT-2), tetraethylthiuram disulfide and tetramethylthiuram disulfide. All three were capable of inactivating RSV, with AT-2 being the most potent. The mechanism of action of AT-2 was analyzed and it was found that AT-2 treatment indeed results in the modification of RSV M2-1. Altered intramolecular disulfide bond formation in M2-1 protein of AT-2-treated RSV virions might have been responsible for abrogation of RSV infectivity. AT-2-inactivated RSV was found to be moderately immunogenic in the cotton rats S.hispidus and did not cause a vaccine-enhancement seen in animals vaccinated with formalin-inactivated RSV. Increasing immunogenicity of AT-2-inactivated RSV by adjuvant (Ribi), however, led to vaccine-enhanced disease. Conclusions This work presents evidence that compounds that inactivate retroviruses by targeting the zinc finger motif in their nucleocapsid proteins are also effective against RSV. AT-2-inactivated RSV vaccine is not strongly immunogenic in the absence of adjuvants. In the adjuvanted form, however, vaccine induces

  6. Respiratory syncytial virus risk factors in late preterm infants.

    PubMed

    Lanari, Marcello; Silvestri, Michela; Rossi, Giovanni A

    2009-01-01

    The key role of respiratory syncytial virus (RSV) in causing infant morbidity and hospitalizations is worldwide well recognized. The late preterm infants (34-36 weeks of gestational age (WGA)) showed a higher risk of hospitalization for RSV-induced infection as compared with full-term infants and similar to that seen in very preterm infants. In addition to the prematurity, a number of risk factors have been identified in 33-35 WGA infants that are associated with RSV-hospitalization as demonstrated by the Canadian and the Spanish studies. However, prematurity per se is not the only factor, since RSV-hospitalization in the first year of life also occurs in numerous full-term and previously healthy infants. Indeed, a recent case-control study on mostly full-term children showed that chronological age at the beginning of RSV season, low birth weight, and birth order (>or=1 sibling) were associated with a higher likelihood to acquire RSV-induced LRTI, severe enough to lead to hospital admission. In order to maximize the cost effectiveness of immunoprophylaxis with palivizumab, the American Academy of Pediatrics have recently updated recommendations resulting in a restriction of its use to infants at highest risk of hospitalization during times when RSV is most likely to be circulating (i.e. in the first 3 months of life for late preterm infants). According to a variety of epidemiologic data in this gestational age group, the risk of exposure should include day-care attendance and having a sibling <5 years of age. However, in an Italian study, in addition to chronological age at the beginning of RSV season, birth weight and birth order were significant predictors for RSV infection with hospitalization. On the basis of the finding that among nations the difference for severe RSV may exist in environmental and demographic risk factors, an 'International' tool has been developed based on the data from the Spanish FLIP study to predict the likelihood of RSV

  7. A Druggable Pocket at the Nucleocapsid/Phosphoprotein Interaction Site of Human Respiratory Syncytial Virus

    PubMed Central

    Ouizougun-Oubari, Mohamed; Pereira, Nelson; Tarus, Bogdan; Galloux, Marie; Lassoued, Safa; Fix, Jenna; Tortorici, M. Alejandra; Hoos, Sylviane; Baron, Bruno; England, Patrick; Desmaële, Didier; Couvreur, Patrick; Bontems, François; Rey, Félix A.; Eléouët, Jean-François; Slama-Schwok, Anny

    2015-01-01

    ABSTRACT Presently, respiratory syncytial virus (RSV), the main cause of severe respiratory infections in infants, cannot be treated efficiently with antivirals. However, its RNA-dependent polymerase complex offers potential targets for RSV-specific drugs. This includes the recognition of its template, the ribonucleoprotein complex (RNP), consisting of genomic RNA encapsidated by the RSV nucleoprotein, N. This recognition proceeds via interaction between the phosphoprotein P, which is the main polymerase cofactor, and N. The determinant role of the C terminus of P, and more particularly of the last residue, F241, in RNP binding and viral RNA synthesis has been assessed previously. Here, we provide detailed structural insight into this crucial interaction for RSV polymerase activity. We solved the crystallographic structures of complexes between the N-terminal domain of N (N-NTD) and C-terminal peptides of P and characterized binding by biophysical approaches. Our results provide a rationale for the pivotal role of F241, which inserts into a well-defined N-NTD pocket. This primary binding site is completed by transient contacts with upstream P residues outside the pocket. Based on the structural information of the N-NTD:P complex, we identified inhibitors of this interaction, selected by in silico screening of small compounds, that efficiently bind to N and compete with P in vitro. One of the compounds displayed inhibitory activity on RSV replication, thereby strengthening the relevance of N-NTD for structure-based design of RSV-specific antivirals. IMPORTANCE Respiratory syncytial virus (RSV) is a widespread pathogen that is a leading cause of acute lower respiratory infections in infants worldwide. RSV cannot be treated efficiently with antivirals, and no vaccine is presently available, with the development of pediatric vaccines being particularly challenging. Therefore, there is a need for new therapeutic strategies that specifically target RSV. The interaction

  8. Role of bibersteinia trehalosi, respiratory syncytial virus, and parainfluenza-3 virus in bighorn sheep pneumonia

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pneumonic bighorn sheep (BHS) have been found to be culture- and/or sero-positive for Bibersteinia trehalosi, respiratory syncytial virus (RSV), and parainfluenza-3 virus (PI-3). The objective of this study was to determine whether these pathogens can cause fatal pneumonia in BHS. In the first study...

  9. A simple and rapid microassay for the titration of human respiratory syncytial virus.

    PubMed

    Trépanier, P; Payment, P; Trudel, M

    1980-01-01

    A microassay, using tissue culture microplates for the titration of human respiratory syncytial virus by syncytium formation, is described. Virus titers obtained agreed well with those obtained in a larger assay system; the microassay, however, is more rapid and economical. Large numbers of virus samples are easily and rapidly processed as the assay necessitates an incubation period of only three days.

  10. A randomized placebo controlled trial of ibuprofen for respiratory syncytial infection in a bovine model study

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: Respiratory syncytial virus (RSV) is the most common cause of bronchiolitis and hospital admission in infants. An analogous disease occurs in cattle and costs US agriculture a billion dollars a year. RSV causes much of its morbidity indirectly via adverse effects of the host response to ...

  11. Respiratory syncytial virus, adenoviruses, and mixed acute lower respiratory infections in children in a developing country.

    PubMed

    Rodríguez-Martínez, Carlos E; Rodríguez, Diego Andrés; Nino, Gustavo

    2015-05-01

    There is growing evidence suggesting greater severity and worse outcomes in children with mixed as compared to single respiratory virus infections. However, studies that assess the risk factors that may predispose a child to a mixture of respiratory syncytial virus (RSV) and adenoviral infections, are scarce. In a retrospective cohort study, the study investigated the epidemiology of RSV and adenovirus infections and predictors of mixed RSV-adenoviral infections in young children hospitalized with acute lower respiratory infection in Bogota, Colombia, South America, over a 2-year period 2009-2011. Of a total of 5,539 children admitted with a diagnosis of acute lower respiratory infection, 2,267 (40.9%) who were positive for RSV and/or adenovirus were selected. Out the total number of cases, 1,416 (62.5%) infections occurred during the 3-month period from March to May, the first rainy season of Bogota, Colombia. After controlling for gender, month when the nasopharyngeal sample was taken, and other pre-existing conditions, it was found that an age greater than 6 months (OR:1.74; CI 95%:1.05-2.89; P = 0.030) and malnutrition as a comorbidity (OR:9.92; CI 95%:1.01-100.9; P = 0.049) were independent predictors of mixed RSV-adenoviral infections in the sample of patients. In conclusion, RSV and adenovirus are significant causes of acute lower respiratory infection in infants and young children in Bogota, Colombia, especially during the first rainy season. The identified predictors of mixed RSV-adenoviral infections should be taken into account when planning intervention, in order to reduce the burden of acute lower respiratory infection in young children living in the country.

  12. Concurrent detection of other respiratory viruses in children shedding viable human respiratory syncytial virus.

    PubMed

    Gagliardi, T B; Paula, F E; Iwamoto, M A; Proença-Modena, J L; Santos, A E; Camara, A A; Cervi, M C; Cintra, O A L; Arruda, E

    2013-10-01

    Human respiratory syncytial virus (HRSV) is an important cause of respiratory disease. The majority of studies addressing the importance of virus co-infections to the HRSV-disease have been based on the detection of HRSV by RT-PCR, which may not distinguish current replication from prolonged shedding of remnant RNA from previous HRSV infections. To assess whether co-detections of other common respiratory viruses are associated with increased severity of HRSV illnesses from patients who were shedding viable-HRSV, nasopharyngeal aspirates from children younger than 5 years who sought medical care for respiratory infections in Ribeirão Preto (Brazil) were tested for HRSV by immunofluorescence, RT-PCR and virus isolation in cell culture. All samples with viable-HRSV were tested further by PCR for other respiratory viruses. HRSV-disease severity was assessed by a clinical score scale. A total of 266 samples from 247 children were collected and 111 (42%) were HRSV-positive. HRSV was isolated from 70 (63%), and 52 (74%) of them were positive for at least one additional virus. HRSV-positive diseases were more severe than HRSV-negative ones, but there was no difference in disease severity between patients with viable-HRSV and those HRSV-positives by RT-PCR. Co-detection of other viruses did not correlate with increased disease severity. HRSV isolation in cell culture does not seem to be superior to RT-PCR to distinguish infections associated with HRSV replication in studies of clinical impact of HRSV. A high rate of co-detection of other respiratory viruses was found in samples with viable-HRSV, but this was not associated with more severe HRSV infection.

  13. Risk factors for hypoxemia and respiratory failure in respiratory syncytial virus bronchiolitis.

    PubMed

    Chan, P W K; Lok, F Y L; Khatijah, S B

    2002-12-01

    Respiratory syncytial virus (RSV) bronchiolitis is a common infection in young children and may result in hospitalization. We examined the incidence of, and risk factors associated with, hypoxemia and respiratory failure in 216 children aged < 24 months admitted consecutively for proven RSV bronchiolitis. Hypoxemia was defined as SpO2 < 90% in room air and severe RSV bronchiolitis requiring intubation and ventilation was categorized as respiratory failure. Corrected age at admission was used for premature children (gestation < 37 weeks). Hypoxemia was suffered by 31 (14.3%) children. It was more likely to occur in children who were Malay (OR 2.56, 95%CI 1.05-6.23, p=0.03) or premature (OR 6.72, 95%CI 2.69-16.78, p<0.01). Hypoxemia was also more likely to develop in children with failure to thrive (OR 2.96, 95%CI 1.28-6.82, p<0.01). The seven (3.2%) children who were both premature (OR 11.94, 95%CI 2.50-56.99, p<0.01) and failure to thrive (OR 6.41, 95%CI 1.37-29.87, p=0.02) were more likely to develop respiratory failure. Prematurity was the only significant risk factor for hypoxemia and respiratory failure by logistic regression analysis (OR 1.17, 95%CI 1.06-1.55, p<0.01 and OR 1.14 95%CI 1.02-2.07, p=0.02 respectively). Prematurity was the single most important risk factor for both hypoxemia and respiratory failure in RSV bronchiolitis. PMID:12757230

  14. Respiratory Syncytial Virus Infection: Mechanisms of Redox Control and Novel Therapeutic Opportunities

    PubMed Central

    Garofalo, Roberto P.; Kolli, Deepthi

    2013-01-01

    Abstract Respiratory syncytial virus (RSV) is one of the most important causes of upper and lower respiratory tract infections in infants and young children, for which no effective treatment is currently available. Although the mechanisms of RSV-induced airway disease remain incompletely defined, the lung inflammatory response is thought to play a central pathogenetic role. In the past few years, we and others have provided increasing evidence of a role of reactive oxygen species (ROS) as important regulators of RSV-induced cellular signaling leading to the expression of key proinflammatory mediators, such as cytokines and chemokines. In addition, RSV-induced oxidative stress, which results from an imbalance between ROS production and airway antioxidant defenses, due to a widespread inhibition of antioxidant enzyme expression, is likely to play a fundamental role in the pathogenesis of RSV-associated lung inflammatory disease, as demonstrated by a significant increase in markers of oxidative injury, which correlate with the severity of clinical illness, in children with RSV infection. Modulation of ROS production and oxidative stress therefore represents a potential novel pharmacological approach to ameliorate RSV-induced lung inflammation and its long-term consequences. Antioxid. Redox Signal. 18, 186–217. PMID:22799599

  15. Immunological, Viral, Environmental, and Individual Factors Modulating Lung Immune Response to Respiratory Syncytial Virus

    PubMed Central

    Bottau, Paolo; Faldella, Giacomo

    2015-01-01

    Respiratory syncytial virus is a worldwide pathogen agent responsible for frequent respiratory tract infections that may become severe and potentially lethal in high risk infants and adults. Several studies have been performed to investigate the immune response that determines the clinical course of the infection. In the present paper, we review the literature on viral, environmental, and host factors influencing virus response; the mechanisms of the immune response; and the action of nonimmunological factors. These mechanisms have often been studied in animal models and in the present review we also summarize the main findings obtained from animal models as well as the limits of each of these models. Understanding the lung response involved in the pathogenesis of these respiratory infections could be useful in improving the preventive strategies against respiratory syncytial virus. PMID:26064963

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

    PubMed

    Klein, B S; Dollete, F R; Yolken, R H

    1982-07-01

    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.

  17. Association of bovine respiratory syncytial virus with atypical interstitial pneumonia in feedlot cattle.

    PubMed

    Collins, J K; Jensen, R; Smith, G H; Flack, D E; Kerschen, R; Bennett, B W; Jones, R L; Alexander, A F

    1988-07-01

    Thirty-three cattle with fatal respiratory tract disease were examined for gross and histologic lesions and for the presence of viral and bacterial agents in the lungs. Fifteen cattle had lesions characteristic of atypical interstitial pneumonia (AIP), and 18 had other respiratory tract diseases, including infectious bovine rhinotracheitis, shipping fever pneumonia, bronchopneumonia, pulmonary abscess, and edema of the trachea. Gross necropsy findings in the cattle with AIP were uncollapsed and emphysematous lungs; histopathologic findings included interstitial edema, thickening of alveolar walls, hyaline membrane formation, and hyperplasia of type-II pneumonocytes. The infective agents found in the lungs of the 33 cattle included bovine respiratory syncytial virus, bovine herpesvirus type 1, Pasteurella sp, mycoplasmas, and Corynebacterium pyogenes. Bovine respiratory syncytial virus was detected by use of immunofluorescence and immunoperoxidase on lung tissue sections; bovine herpesvirus type 1 was detected by these techniques and by isolation of the virus. Bovine respiratory syncytial virus was significantly (P = 0.01) associated with lesions of AIP (11 of 15), compared with those of other respiratory tract diseases (5 of 18).

  18. Experimental reproduction of respiratory tract disease with bovine respiratory syncytial virus.

    PubMed

    Ciszewski, D K; Baker, J C; Slocombe, R F; Reindel, J F; Haines, D M; Clark, E G

    1991-06-01

    An experiment was conducted to reproduce respiratory tract disease with bovine respiratory syncytial virus (BRSV) in one-month-old, colostrum-fed calves. The hypothesized role of viral hypersensitivity and persistent infection in the pathogenesis of BRSV pneumonia was also investigated. For BRSV inoculation a field isolate of BRSV, at the fifth passage level in cell culture, was administered by a combined respiratory tract route (intranasal and intratracheal) for four consecutive days. Four groups of calves were utilized as follows: Group I, 6 calves sham inoculated with uninfected tissue culture fluid and necropsied 21 days after the last inoculation; Group II, 6 calves inoculated with BRSV and necropsied at the time of maximal clinical response (4-6 days after the last inoculation); Group III, 6 calves inoculated with BRSV and necropsied at 21 days after the last inoculation; Group IV, 6 calves inoculated with BRSV, rechallenged with BRSV 10 days after initial exposure, and necropsied at 21 days after the initial inoculation. Clinical response was evaluated by daily monitoring of body temperature, heart rate, respiratory rate, arterial blood gas tensions, hematocrit, total protein, white blood cell count, and fibrinogen. Calves were necropsied and pulmonary surface lesions were quantitated by computer digitization. Viral pneumonia was reporduced in each principal group. Lesions were most extensive in Group II. Disease was not apparent in Group I (controls). Significant differences (p less than 0.05) in body temperature, heart rate, respiratory rate, arterial oxygen tension, and pneumonic surface area were demonstrated between control and infected calves. Results indicate that severe disease and lesions can be induced by BRSV in one-month-old calves that were colostrum-fed and seropositive to BRSV. BRSV rechallenge had minimal effect on disease progression. Based on clinical and pathological response, results did not support viral hypersensitivity or persistent

  19. Dimerization of Matrix Protein Is Required for Budding of Respiratory Syncytial Virus

    PubMed Central

    Förster, Andreas; Maertens, Goedele N.; Farrell, Paul J.

    2015-01-01

    ABSTRACT Respiratory syncytial virus (RSV) infects epithelial cells of the respiratory tract and is a major cause of bronchiolitis and pneumonia in children and the elderly. The virus assembles and buds through the plasma membrane, forming elongated membrane filaments, but details of how this happens remain obscure. Oligomerization of the matrix protein (M) is a key step in the process of assembly and infectious virus production. In addition, it was suggested to affect the conformation of the fusion protein, the major current target for RSV antivirals, in the mature virus. The structure and assembly of M are thus key parameters in the RSV antiviral development strategy. The structure of RSV M was previously published as a monomer. Other paramyxovirus M proteins have been shown to dimerize, and biochemical data suggest that RSV M also dimerizes. Here, using size exclusion chromatography-multiangle laser light scattering, we show that the protein is dimeric in solution. We also crystallized M in two crystal forms and show that it assembles into equivalent dimers in both lattices. Dimerization interface mutations destabilize the M dimer in vitro. To assess the biological relevance of dimerization, we used confocal imaging to show that dimerization interface mutants of M fail to assemble into viral filaments on the plasma membrane. Additionally, budding and release of virus-like particles are prevented in M mutants that fail to form filaments. Importantly, we show that M is biologically active as a dimer and that the switch from M dimers to higher-order oligomers triggers viral filament assembly and virus production. IMPORTANCE Human respiratory syncytial virus (RSV) is the most frequent cause of infantile bronchiolitis and pneumonia. The enormous burden of RSV makes it a major unmet target for a vaccine and antiviral drug therapy. Oligomerization of the matrix protein is a key step in the process of assembly and production of infectious virus, but the molecular

  20. The 22,000-kilodalton protein of respiratory syncytial virus is a major target for Kd-restricted cytotoxic T lymphocytes from mice primed by infection.

    PubMed Central

    Openshaw, P J; Anderson, K; Wertz, G W; Askonas, B A

    1990-01-01

    Recombinant vaccinia viruses containing the 22-kilodalton protein (matrixlike or 22K protein) or phosphoprotein gene from respiratory syncytial virus were constructed. These recombinant viruses expressed proteins which were immunoprecipitated by appropriate respiratory syncytial virus antibodies and comigrated with authentic proteins produced by respiratory syncytial virus infection. The new recombinant viruses (and others previously described containing the attachment glycoprotein, fusion, or nucleoprotein genes of respiratory syncytial virus) were used to infect target cells for cultured polyclonal cytotoxic T lymphocytes generated from the spleens of BALB/c or DBA/2 mice primed by intranasal infection with respiratory syncytial virus. Respiratory syncytial virus-specific cytotoxic T lymphocytes (CTL) showed strong Kd (but not Dd)-restricted recognition of the 22K protein. As previously reported, the fusion protein and nucleoprotein were both seen by CTL, but recognition of these proteins was comparatively weak. There was no detectable recognition of other respiratory syncytial virus proteins tested (including phosphoprotein). 22K protein-specific splenic memory CTL persisted for at least 11 months after infection of BALB/c mice. Priming BALB/c mice with recombinant vaccinia virus containing the 22K protein gene induced respiratory syncytial virus-specific memory CTL at lower levels than that previously reported following infection with a similar recombinant containing the fusion protein gene. These data identify the 22K protein as a major target antigen for respiratory syncytial virus-specific CTL from H-2d mice primed by respiratory syncytial virus infection. Images PMID:2319650

  1. CT findings in viral lower respiratory tract infections caused by parainfluenza virus, influenza virus and respiratory syncytial virus

    PubMed Central

    Kim, Min-Chul; Kim, Mi Young; Lee, Hyun Joo; Lee, Sang-Oh; Choi, Sang-Ho; Kim, Yang Soo; Woo, Jun Hee; Kim, Sung-Han

    2016-01-01

    Abstract Viral lower respiratory tract infections (LRTIs) can present with a variety of computed tomography (CT) findings. However, identifying the contribution of a particular virus to CT findings is challenging due to concomitant infections and the limited data on the CT findings in viral LRTIs. We therefore investigate the CT findings in different pure viral LRTIs. All patients who underwent bronchoalveolar lavage (BAL) and were diagnosed with LRTIs caused by parainfluenza virus (PIV), influenza virus, or respiratory syncytial virus (RSV) between 1998 and 2014 were enrolled in a tertiary hospital in Seoul, South Korea. A pure viral LRTI was defined as a positive viral culture from BAL without any positive evidence from respiratory or blood cultures, or from polymerase chain reaction (PCR), or from serologic tests for bacteria, fungi, mycobacteria, or other viruses. CT images of 40 patients with viral LRTIs were analyzed: 14 with PIV, 14 with influenza virus, and 12 with RSV. Patch consolidation (≥1 cm or more than 1 segmental level) was found only in PIV (29%) (P = 0.03), by which CT findings caused by PIV could resemble those seen in bacterial LRTIs. Ground-glass opacities were seen in all cases of influenza virus and were more frequent than in PIV (71%) and RSV (67%) (P = 0.05). Bronchial wall thickening was more common in influenza virus (71%) and RSV (67%) LRTIs than PIV LRTIs (21%) (P = 0.02). With respect to anatomical distribution, PIV infections generally affected the lower lobes (69%), while influenza virus mostly caused diffuse changes throughout the lungs (57%), and RSV frequently formed localized patterns in the upper and mid lobes (44%). The CT findings in LRTIs of PIV, influenza virus, and RSV can be distinguished by certain characteristics. These differences could be useful for early differentiation of these viral LRTIs, and empirical use of appropriate antiviral agents. PMID:27368011

  2. Investigation of Respiratory Syncytial Virus-Associated Deaths Among US Children Aged <2 Years, 2004-2007.

    PubMed

    Prill, Mila M; Iwane, Marika K; Little, Delmar; Gerber, Susan I

    2016-09-01

    We validated the respiratory syncytial virus-coded deaths of children aged <2 years in 2004-2007 using national/state death data and medical records. There were 48 deaths in 4 states, and hospital records for 32 of them were available; 26 of those 32 (81%) had a laboratory finding of respiratory syncytial virus, and 21 of those 26 (81%) had a potential high-risk condition, most commonly preterm birth (35%). PMID:27534673

  3. Progress in the development of respiratory syncytial virus and parainfluenza virus vaccines.

    PubMed

    Durbin, Anna P; Karron, Ruth A

    2003-12-15

    Respiratory syncytial virus (RSV) and human parainfluenza viruses (hPIVs) are leading causes of viral lower respiratory tract illness in children and in high-risk adult populations. Despite decades of research, licensed vaccines for RSV and hPIVs do not exist. Recently, however, genetically engineered live attenuated RSV and hPIV candidate vaccines have been generated, several of which are already being evaluated in clinical trials. Recombinant technology allows candidate vaccines to be "fine-tuned" in response to clinical data, which should hasten the development of vaccines against these important respiratory pathogens. PMID:14689350

  4. Application of Traditional Chinese Medical Herbs in Prevention and Treatment of Respiratory Syncytial Virus

    PubMed Central

    Lin, Li Li; Xie, Tong; Xu, Jian Ya; Shen, Cun Si; Di, Liu Qing; Chen, Jia Bin

    2016-01-01

    Respiratory syncytial virus (RSV) is a common viral pathogen of the lower respiratory tract, which, in the absence of effective management, causes millions of cases of severe illness per year. Many of these infections develop into fatal pneumonia. In a review of English and Chinese medical literature, recent traditional Chinese medical herb- (TCMH-) based progress in the area of prevention and treatment was identified, and the potential anti-RSV compounds, herbs, and formulas were explored. Traditional Chinese medical herbs have a positive effect on inhibiting viral attachment, inhibiting viral internalization, syncytial formation, alleviation of airway inflammation, and stimulation of interferon secretion and immune system; however, the anti-RSV mechanisms of TCMHs are complicated, which should be further investigated. PMID:27688789

  5. Application of Traditional Chinese Medical Herbs in Prevention and Treatment of Respiratory Syncytial Virus

    PubMed Central

    Lin, Li Li; Xie, Tong; Xu, Jian Ya; Shen, Cun Si; Di, Liu Qing; Chen, Jia Bin

    2016-01-01

    Respiratory syncytial virus (RSV) is a common viral pathogen of the lower respiratory tract, which, in the absence of effective management, causes millions of cases of severe illness per year. Many of these infections develop into fatal pneumonia. In a review of English and Chinese medical literature, recent traditional Chinese medical herb- (TCMH-) based progress in the area of prevention and treatment was identified, and the potential anti-RSV compounds, herbs, and formulas were explored. Traditional Chinese medical herbs have a positive effect on inhibiting viral attachment, inhibiting viral internalization, syncytial formation, alleviation of airway inflammation, and stimulation of interferon secretion and immune system; however, the anti-RSV mechanisms of TCMHs are complicated, which should be further investigated.

  6. Application of Traditional Chinese Medical Herbs in Prevention and Treatment of Respiratory Syncytial Virus.

    PubMed

    Lin, Li Li; Shan, Jin Jun; Xie, Tong; Xu, Jian Ya; Shen, Cun Si; Di, Liu Qing; Chen, Jia Bin; Wang, Shou Chuan

    2016-01-01

    Respiratory syncytial virus (RSV) is a common viral pathogen of the lower respiratory tract, which, in the absence of effective management, causes millions of cases of severe illness per year. Many of these infections develop into fatal pneumonia. In a review of English and Chinese medical literature, recent traditional Chinese medical herb- (TCMH-) based progress in the area of prevention and treatment was identified, and the potential anti-RSV compounds, herbs, and formulas were explored. Traditional Chinese medical herbs have a positive effect on inhibiting viral attachment, inhibiting viral internalization, syncytial formation, alleviation of airway inflammation, and stimulation of interferon secretion and immune system; however, the anti-RSV mechanisms of TCMHs are complicated, which should be further investigated. PMID:27688789

  7. Dual Proinflammatory and Antiviral Properties of Pulmonary Eosinophils in Respiratory Syncytial Virus Vaccine-Enhanced Disease

    PubMed Central

    Su, Yung-Chang; Townsend, Dijana; Herrero, Lara J.; Zaid, Ali; Rolph, Michael S.; Gahan, Michelle E.; Nelson, Michelle A.; Rudd, Penny A.; Matthaei, Klaus I.; Foster, Paul S.; Dent, Lindsay; Tripp, Ralph A.; Lee, James; Simson, Ljubov

    2014-01-01

    ABSTRACT Human respiratory syncytial virus (RSV) is a major cause of morbidity and severe lower respiratory tract disease in the elderly and very young, with some infants developing bronchiolitis, recurrent wheezing, and asthma following infection. Previous studies in humans and animal models have shown that vaccination with formalin-inactivated RSV (FI-RSV) leads to prominent airway eosinophilic inflammation following RSV challenge; however, the roles of pulmonary eosinophilia in the antiviral response and in disease pathogenesis are inadequately understood. In vivo studies in mice with eotaxin and/or interleukin 5 (IL-5) deficiency showed that FI-RSV vaccination did not lead to enhanced pulmonary disease, where following challenge there were reduced pulmonary eosinophilia, inflammation, Th2-type cytokine responses, and altered chemokine (TARC and CCL17) responses. In contrast to wild-type mice, RSV was recovered at high titers from the lungs of eotaxin- and/or IL-5-deficient mice. Adoptive transfer of eosinophils to FI-RSV-immunized eotaxin- and IL-5-deficient (double-deficient) mice challenged with RSV was associated with potent viral clearance that was mediated at least partly through nitric oxide. These studies show that pulmonary eosinophilia has dual outcomes: one linked to RSV-induced airway inflammation and pulmonary pathology and one with innate features that contribute to a reduction in the viral load. IMPORTANCE This study is critical to understanding the mechanisms attributable to RSV vaccine-enhanced disease. This study addresses the hypothesis that IL-5 and eotaxin are critical in pulmonary eosinophil response related to FI-RSV vaccine-enhanced disease. The findings suggest that in addition to mediating tissue pathology, eosinophils within a Th2 environment also have antiviral activity. PMID:25410867

  8. Revised indications for the use of palivizumab and respiratory syncytial virus immune globulin intravenous for the prevention of respiratory syncytial virus infections.

    PubMed

    Meissner, H Cody; Long, Sarah S

    2003-12-01

    Palivizumab and Respiratory Syncytial Virus Immune Globulin Intravenous (RSV-IGIV) are licensed by the Food and Drug Administration for use in preventing severe respiratory syncytial virus (RSV) infections in high-risk infants, children younger than 24 months with chronic lung disease (formerly called bronchopulmonary dysplasia), and certain preterm infants. This report summarizes the clinical trial information on which the guidance in the accompanying policy statement for administering RSV prophylaxis to certain children with a history of preterm birth, chronic lung disease, or congenital heart disease is based. On the basis of results of a recently completed clinical trial, palivizumab is appropriate for infants and young children with hemodynamically significant congenital heart disease. RSV-IGIV should not be used in children with hemodynamically significant heart disease. Palivizumab is preferred for most high-risk infants and children because of ease of intramuscular administration. Monthly administration of palivizumab during the RSV season results in a 45% to 55% decrease in the rate of hospitalization attributable to RSV. Because of the large number of infants born after 32 to 35 weeks' gestation and because of the high cost, immunoprophylaxis should be considered for this category of preterm infants only if 2 or more risk factors are present. PMID:14654628

  9. Secretory Expression and Purification of Respiratory Syncytial Virus G and F Proteins in Human Cells.

    PubMed

    Jadhao, Samadhan J; Anderson, Larry J

    2016-01-01

    Respiratory syncytial virus (RSV) is one of the leading causes of range of symptoms from mild upper to serious lower respiratory virus infections in infants, immunocompromised individuals, and the elderly. Despite many decades of research and development, a licensed RSV vaccine is not available for use in human. Since the RSV F and G proteins induce neutralizing antibodies and confer protection from infection, they are important for understanding disease and for developing vaccines and access to purified, expressed proteins is important to RSV research and diagnostics. We describe methods to produce recombinant RSV F and G proteins in human cells and purify these proteins using Ni Sepharose affinity chromatography. PMID:27464687

  10. Novel insights into human respiratory syncytial virus-host factor interactions through integrated proteomics and transcriptomics analysis

    PubMed Central

    Dapat, Clyde; Oshitani, Hitoshi

    2016-01-01

    ABSTRACT The lack of vaccine and limited antiviral options against respiratory syncytial virus (RSV) highlights the need for novel therapeutic strategies. One alternative is to develop drugs that target host factors required for viral replication. Several microarray and proteomics studies had been published to identify possible host factors that are affected during RSV replication. In order to obtain a comprehensive understanding of RSV-host interaction, we integrated available proteome and transcriptome datasets and used it to construct a virus-host interaction network. Then, we interrogated the network to identify host factors that are targeted by the virus and we searched for drugs from the DrugBank database that interact with these host factors, which may have potential applications in repositioning for future treatment options of RSV infection. PMID:26760927

  11. Global gene expression profiling in infants with acute respiratory syncytial virus broncholitis demonstrates systemic activation of interferon signaling networks

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Respiratory syncytial virus (RSV) is a leading cause of pediatric lower respiratory tract infections and has a high impact on pediatric emergency department utilization. Variation in host response may influence the pathogenesis and disease severity. We evaluated global gene expression profiles to be...

  12. Systematic review of the biology and medical management of respiratory syncytial virus infection.

    PubMed

    Black, Craig Patrick

    2003-03-01

    Respiratory syncytial virus, the leading cause of serious upper and lower respiratory tract infection in infants and children, accounts for 125,000 hospitalizations and 450 deaths annually in the United States. It also may predispose to development of asthma later in life. Annual epidemics occur from November to April, and virtually all infants are infected by age 2. Immunity is not durable; hence, reinfection occurs throughout life, although subsequent infections are nearly always mild. Certain populations (eg, premature infants, infants with chronic lung disease, and immunocompromised individuals) are at risk for severe morbidity and have higher risk of mortality. Infection is spread to the nose and eyes by large droplets and direct contact with secretions, and fomites may remain infectious for up to 12 hours. Nosocomial infection is common. The virus infects airway ciliated epithelial cells, spreading by the formation of syncytia. Cellular debris and inflammation cause airway obstruction, hyperinflation, localized atelectasis, wheezing, and impaired gas exchange. Both humoral and cellular immune response are critical to ending the acute infection, but wheezing and reactive airways may persist for as long as 5-10 years after acute infection. No cure exists for respiratory syncytial virus infection, but commonly employed palliative treatments include oxygen, inhaled beta(2) agonists, racemic epinephrine, dornase alfa, systemic and inhaled corticosteroids, inhaled ribavirin, and nasopharyngeal suctioning. Infants suffering severe lower airways disease may require mechanical ventilation. Prophylactic measures include rigorous infection control and administration of polyclonal (RSV-IGIV [respiratory syncytial virus - immunoglobulin intravenous]) and monoclonal (palivizumab) antibodies. The cost of the prophylactic antibody treatment is high; it is cost-effective for only the highest risk patients. Development of a vaccine remains far in the future. Application of

  13. Bacteremia in Children Hospitalized with Respiratory Syncytial Virus Infection

    PubMed Central

    Pardo-Seco, Jacobo; Gómez-Carballa, Alberto; Martinón-Torres, Nazareth; Martinón-Sánchez, José María; Justicia-Grande, Antonio; Rivero-Calle, Irene; Pinnock, Elli; Salas, Antonio; Fink, Colin

    2016-01-01

    Background The risk of bacteremia is considered low in children with acute bronchiolitis. However the rate of occult bacteremia in infants with RSV infection is not well established. The aim was to determine the actual rate and predictive factors of bacteremia in children admitted to hospital due to confirmed RSV acute respiratory illness (ARI), using both conventional culture and molecular techniques. Methods A prospective multicenter study (GENDRES-network) was conducted between 2011–2013 in children under the age of two admitted to hospital because of an ARI. Among those RSV-positive, bacterial presence in blood was assessed using PCR for Meningococcus, Streptococcus pneumoniae, Haemophilus influenzae, Streptococcus pyogenes, Klebsiella pneumoniae, Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus, in addition to conventional cultures. Results 66 children with positive RSV respiratory illness were included. In 10.6% patients, bacterial presence was detected: H. influenzae (n = 4) and S. pneumoniae (n = 2). In those patients with bacteremia, there was a previous suspicion of bacterial superinfection and had received empirical antibiotic treatment 6 out of 7 (85.7%) patients. There were significant differences in terms of severity between children with positive bacterial PCR and those with negative results: PICU admission (100% vs. 50%, P-value = 0.015); respiratory support necessity (100% vs. 18.6%, P-value < 0.001); Wood-Downes score (mean = 8.7 vs. 4.8 points, P-value < 0.001); GENVIP scale (mean = 17 vs. 10.1, P-value < 0.001); and length of hospitalization (mean = 12.1 vs. 7.5 days, P-value = 0.007). Conclusion Bacteremia is not frequent in infants hospitalized with RSV respiratory infection, however, it should be considered in the most severe cases. PMID:26872131

  14. Bovine Respiratory Syncytial Virus (BRSV) Pneumonia in Beef Calf Herds Despite Vaccination

    PubMed Central

    Larsen, LE; Tegtmeier, C; Pedersen, E

    2001-01-01

    The present report describes the clinical, pathological, serological and virological findings in calves from 2 larger Danish beef herds experiencing outbreaks of pneumonia. The calves had been vaccinated with an inactivated bovine respiratory syncytial virus (BRSV) vaccine 2 months prior to the outbreak. The clinical signs comprised nasal discharge, pyrexia, cough and increased respiratory rates. A total of 28 calves died in the 2 herds. The laboratory investigations revealed that BRSV was involved and probably initiated both outbreaks. Furthermore, the serological results suggested that the vaccine induced only sparse levels of antibodies probably due to the presence of maternally derived antibodies at the time of vaccination. Necropsy findings in 5 calves revealed changes typical for infectious pneumonia with involvment of BRSV. In conclusion, vaccination of calves against BRSV in 2 Danish beef herds failed to protect the calves against severe or even fatal BRSV mediated respiratory disease 2 months later. PMID:11455891

  15. Outbreak of respiratory syncytial virus (RSV) infection in immunocompromised adults on a hematology ward.

    PubMed

    Jensen, Tomas Ostergaard; Stelzer-Braid, Sacha; Willenborg, Christiana; Cheung, Carol; Andresen, David; Rawlinson, William; Clezy, Kate

    2016-10-01

    We describe an outbreak of respiratory syncytial virus (RSV) infection on a hematology ward without allogeneic stem cell transplant patients. Twelve patients and one staff member infected with RSV were identified from the laboratory database. Five patients had lower respiratory tract infection, seven had upper respiratory tract infection, one was asymptomatic, and there were two (15.4%) deaths. Most patients had overlapping periods of potential infectiousness on the ward. Sequencing was possible on eight specimens and five of these had identical sequences. Results were consistent with transmission occurring both on the ward and by introduction of RSV from the community. J. Med. Virol. 88:1827-1831, 2016. © 2016 Wiley Periodicals, Inc.

  16. Respiratory syncytial virus fusion inhibitors. Part 4: optimization for oral bioavailability.

    PubMed

    Yu, Kuo-Long; Sin, Ny; Civiello, Rita L; Wang, X Alan; Combrink, Keith D; Gulgeze, H Belgin; Venables, Brian L; Wright, J J Kim; Dalterio, Richard A; Zadjura, Lisa; Marino, Anthony; Dando, Sandra; D'Arienzo, Celia; Kadow, Kathleen F; Cianci, Christopher W; Li, Zhufang; Clarke, Junius; Genovesi, Eugene V; Medina, Ivette; Lamb, Lucinda; Colonno, Richard J; Yang, Zheng; Krystal, Mark; Meanwell, Nicholas A

    2007-02-15

    A series of benzimidazole-based inhibitors of respiratory syncytial virus (RSV) fusion were optimized for antiviral potency, membrane permeability and metabolic stability in human liver microsomes. 1-Cyclopropyl-1,3-dihydro-3-[[1-(4-hydroxybutyl)-1H-benzimidazol-2-yl]methyl]-2H-imidazo[4,5-c]pyridin-2-one (6m, BMS-433771) was identified as a potent RSV inhibitor demonstrating good bioavailability in the mouse, rat, dog and cynomolgus monkey that demonstrated antiviral activity in the BALB/c and cotton rat models of infection following oral administration.

  17. [Bovine serum albumin inhibits the adsorption of respiratory syncytial virus on MDBK cells].

    PubMed

    Fassi Fihri, O; Mohanty, J; Elazhary, Y

    1993-01-01

    In order to obtain optimal bovine respiratory syncytial virus adsorption to host cells, the effect of several products (fetal bovine serum, bovine serum albumin, ovalbumin and cytochrome c) was studied. The adsorption of the virus to MDBK cells was higher in the presence of 2% than in the presence of 5% fetal bovine serum. Adsorption was inhibited in the presence of bovine serum albumin at concentrations > 0.2% when added before or at the same time as the virus. Ovalbumin and cytochrome c did not inhibit adsorption. These results will allow the study of virus adsorption on cell receptors.

  18. Active and passive immunisation against respiratory syncytial virus.

    PubMed

    Zambon, M

    1999-01-01

    RSV is a major cause of respiratory illness in infants under 2 years of age. Evidence is accumulating that it is also underestimated as a cause of respiratory infection in adults, the elderly and immunocompromised individuals. Active interventions to control the impact of RSV infection have been hampered by a lack of understanding of the immune response to RSV in different age groups. A number of different strategies for developing RSV vaccines have been pursued, including live attenuated vaccines, genetically engineered live and subunit vaccines and peptide vaccines with varying degrees of success. The target populations for RSV vaccines include infants, the elderly and women of childbearing age, but the efficacy of different vaccines may differ according to age. Desirable immune responses and immune correlates of protection to RSV in humans remain uncertain and determining these is critical for introduction of any vaccines. Prophylaxis and treatment of RSV in infants using human immunoglobulin containing high titres of RSV specific neutralising antibody (RSV-Ig) has shown limited success in different infant populations. Prophylaxis of premature infants with RSV-Ig, particularly those with bronchopulmonary dysplasia, has demonstrated limited clinical efficacy against RSV. In contrast, there are significant safety concerns for use of this preparation for prophylaxis in infants with congenital heart disease and no demonstrable efficacy in treatment of RSV disease in healthy infants. The cost of the preparation will limit use to highly selected infant groups. Production of humanized monoclonal antibodies to RSV offers another potential passive immunotherapy intervention for RSV, with increased specific activity and reduced side effects, although its use remains experimental. PMID:10578118

  19. Neural-endocrine mechanisms of respiratory syncytial virus-associated asthma in a rat model.

    PubMed

    Li, Q G; Wu, X R; Li, X Z; Yu, J; Xia, Y; Wang, A P; Wang, J

    2012-08-24

    We examined the underlying neural-endocrine mechanisms of asthma associated with respiratory syncytial virus infection. Thirty Sprague-Dawley rats were randomly divided into control group, respiratory syncytial virus (RSV) group, and anti-nerve growth factor (NGF) IgG group. An RSV infection model was established by nasal drip once a week. In the anti-NGF antibody intervention group, each rat was given an intraperitoneal injection of anti-NGF IgG 3 h before RSV infection. Optical microscopy and transmission electron microscopy were used to observe the structural changes in adrenal medulla cells. Changes in adrenaline and norepinephrine in serum were detected by ELISA. NGF expression was assayed by immunohistochemistry. Expression differences in synaptophysin mRNA were detected by RT-PCR. Transmission electron microscopy displayed widened adrenal medulla intercellular spaces, reduced chromaffin particle concentration, and increased mitochondria in the RSV infection group. At the same time, NGF expression was increased in the RSV infection group significantly. In addition, the adrenaline concentration was significantly decreased compared with the control and anti-NGF antibody groups. Synaptophysin mRNA expression was significantly increased in the RSV infection and anti-NGF antibody groups. However, compared with the RSV infection group, synaptophysin mRNA expression was significantly decreased in the anti-NGF antibody group. We conclude that RSV infection could induce adrenal medulla cell differentiation to nerve cells by over-expression of NGF, resulting in the decreased endocrine function found in asthma progression.

  20. Respiratory syncytial virus infection in infants and correlation with meteorological factors and air pollutants

    PubMed Central

    2013-01-01

    Background Respiratory Syncytial Virus (RSV) is the most important cause of severe respiratory infections in infants with seasonal epidemics. Environmental factors (temperature, humidity, air pollution) could influence RSV epidemics through their effects on virus activity and diffusion. Methods We conducted a retrospective study on a paediatric population who referred to our Paediatric Emergency Unit in order to analyze the correlation between weekly incidence of RSV positive cases during winter season in Bologna and meteorological factors and air pollutants concentration. Results We observed a significant correlation between the incidence of RSV infections and the mean minimum temperature registered during the same week and the previous weeks. The weekly number of RSV positive cases was also correlated to the mean PM10 concentration of the week before. Conclusions RSV epidemic trend in Bologna (Italy) is related to the mean minimum temperature, and the mean PM10 concentration. PMID:23311474

  1. Rapid antigen-based testing for respiratory syncytial virus: moving diagnostics from bench to bedside?

    PubMed

    Prendergast, Caitlin; Papenburg, Jesse

    2013-04-01

    Respiratory syncytial virus (RSV) is the most important cause of infantile bronchiolitis and pneumonia. It is ubiquitous, with most children acquiring their primary infection within the first year of life and with subsequent reinfection occurring in all age groups. Clinically, RSV is virtually indistinguishable from other viral respiratory infections. Traditionally, the microbiologic diagnosis of RSV has been based on moderate to complex techniques performed in a laboratory (cell culture, nucleic acid amplification and immunofluorescence assays); however, rapid antigen-detection tests offer potential advantages associated with point-of-care testing. This review seeks to familiarize the readers with RSV rapid antigen-detection tests, describe their performance characteristics and comment on their strengths and weaknesses. The authors will discuss the impact of rapid RSV testing on clinical practice, with a look to the future of what the field ultimately requires of a point-of-care diagnostic technique.

  2. Performance of the Kallestad Pathfinder enzyme immunoassay in the diagnosis of respiratory syncytial virus infections.

    PubMed

    Olsen, M A; Shuck, K M; Sambol, A R; Bohnert, V A; Henery, M L

    1993-01-01

    The Kallestad Pathfinder enzyme immunoassay (EIA) for the rapid detection of respiratory syncytial virus (RSV) antigen was compared with virus culture and direct fluorescent antibody (DFA) to determine the reliability of the EIA. During two consecutive winter respiratory seasons, 270 nasopharyngeal wash specimens were tested. RSV was detected in culture by the presence of cytopathic effect and/or an indirect immunofluorescence assay. The sensitivity of the Pathfinder EIA in comparison with isolation in tube culture was 72% (73 of 101) and the specificity was 99% (167 of 169). During the second year of the evaluation period, DFA was performed on all specimens. The sensitivity of the DFA compared with isolation in tube culture was 94%. This study indicates that the Pathfinder EIA is a very specific test for diagnosis of RSV infections, but lacks sensitivity in comparison with tube culture or direct immunofluorescence.

  3. In Hot Pursuit of the First Vaccine Against Respiratory Syncytial Virus.

    PubMed

    Kim, Joo Young; Chang, Jun

    2016-07-01

    Human respiratory syncytial virus (RSV) is the leading cause of severe lower respiratory tract infection, such as bronchiolitis, bronchitis, or pneumonia, in both infants and the elderly. Despite the global burden of diseases attributable to RSV infection, no clinically approved vaccine is available, and a humanized monoclonal antibody for prophylaxis is not readily affordable in developing countries. There are several hurdles to the successful development of RSV vaccines: immune-vulnerable target populations such as premature infants, pregnant women, and immunocompromised people; safety concerns associated with vaccine-enhanced diseases; repeated infection; and waning memory. To develop successful strategies for the prevention of RSV infection, it is necessary to understand the protective and pathologic roles of host immune responses to RSV infection. In this review, we will summarize the positive and negative relationship between RSV infection and host immunity and discuss strategies for the development of the first successful RSV vaccine. PMID:27189271

  4. IgG-subclass-specific antibody reactivity to respiratory syncytial virus polypeptides investigated by western blot.

    PubMed

    Jankowski, M; Hornsleth, A; Olsen, P G

    1990-01-01

    IgG1 and IgG3 subclass-specific antibody reactivity (ABR) in serum samples obtained from infants and children in relation to acute lower respiratory disease caused by respiratory syncytial virus (RSV) infection was investigated by Western blot. IgG1 ABR was directed against the nucleocapsid polypeptides VPN and VPP as well as against the glycoproteins GP48 (F1) and GP90. IgG3 ABR was directed only against VPN and VPP. In infants, a low IgG1 reactivity against glycoproteins was observed. When serum samples obtained in the early convalescent phase were tested, ABR against GP48 and GP90 as well as against VPP differed with respect to RSV subtypes A and B. IgG1 ABR increased in the late convalescent phase, while IgG3 ABR decreased during this phase when serum samples from primary infections were tested.

  5. In Hot Pursuit of the First Vaccine Against Respiratory Syncytial Virus

    PubMed Central

    Kim, Joo Young

    2016-01-01

    Human respiratory syncytial virus (RSV) is the leading cause of severe lower respiratory tract infection, such as bronchiolitis, bronchitis, or pneumonia, in both infants and the elderly. Despite the global burden of diseases attributable to RSV infection, no clinically approved vaccine is available, and a humanized monoclonal antibody for prophylaxis is not readily affordable in developing countries. There are several hurdles to the successful development of RSV vaccines: immune-vulnerable target populations such as premature infants, pregnant women, and immunocompromised people; safety concerns associated with vaccine-enhanced diseases; repeated infection; and waning memory. To develop successful strategies for the prevention of RSV infection, it is necessary to understand the protective and pathologic roles of host immune responses to RSV infection. In this review, we will summarize the positive and negative relationship between RSV infection and host immunity and discuss strategies for the development of the first successful RSV vaccine. PMID:27189271

  6. Role of respiratory syncytial virus in acute otitis media: implications for vaccine development.

    PubMed

    Patel, Janak A; Nguyen, Dang T; Revai, Krystal; Chonmaitree, Tasnee

    2007-02-19

    We summarize herein the results of various virologic studies of acute otitis media (AOM) conducted at our site over a 10-year period. Among 566 children with AOM, respiratory syncytial virus (RSV) was the most common virus identified in either middle ear fluid or nasal wash; it was found in 16% of all children and 38% of virus-positive children. Seventy-one percent of the children with RSV were 1 year of age or older, which was significantly older than all other viruses combined (P=0.045). RSV infection was associated with the common bacterial pathogens causing AOM. Past efforts to develop vaccines for RSV have emphasized prevention of lower respiratory tract infection in infants, which is a more serious problem but less common than AOM. Our results suggest that RSV vaccines that work only against infection in older children may have value in preventing AOM, the most common pediatric disease.

  7. Recent Advances in Diagnosis, Prevention, and Treatment of Human Respiratory Syncytial Virus

    PubMed Central

    Bawage, Swapnil Subhash; Tiwari, Pooja Munnilal; Singh, Shree Ram

    2013-01-01

    Human respiratory syncytial virus (RSV) is a common cause of respiratory infection in infants and the elderly, leading to significant morbidity and mortality. The interdisciplinary fields, especially biotechnology and nanotechnology, have facilitated the development of modern detection systems for RSV. Many anti-RSV compounds like fusion inhibitors and RNAi molecules have been successful in laboratory and clinical trials. But, currently, there are no effective drugs for RSV infection even after decades of research. Effective diagnosis can result in effective treatment, but the progress in both of these facets must be concurrent. The development in prevention and treatment measures for RSV is at appreciable pace, but the implementation into clinical practice still seems a challenge. This review attempts to present the promising diverse research approaches and advancements in the area of diagnosis, prevention, and treatment that contribute to RSV management. PMID:24382964

  8. Protection against respiratory disease in calves induced by vaccines containing respiratory syncytial virus, parainfluenza type 3 virus, Mycoplasma bovis and M dispar.

    PubMed

    Howard, C J; Stott, E J; Thomas, L H; Gourlay, R N; Taylor, G

    1987-10-17

    A field trial to assess the ability of two vaccines to protect calves against respiratory disease was carried out on a large beef rearing unit in southern England over the two winters of 1983 to 1984 and 1984 to 1985. A quadrivalent vaccine containing the killed antigens of respiratory syncytial virus, parainfluenza virus type 3, Mycoplasma bovis and M dispar or a vaccine containing only the respiratory syncytial virus component were inoculated into 246 and 245 calves, respectively; 245 calves remained as unvaccinated controls. The calves were reared in seven batches and outbreaks of disease occurred in five; significant protection was achieved in the four batches in which disease was associated with respiratory syncytial virus and M bovis infection, together or independently. The death rate from pneumonia was 9 per cent in the control group, 2 per cent in the calves inoculated with the quadrivalent vaccine (P less than 0.001), a protection rate of 77 per cent, and 3 per cent in the calves inoculated with the respiratory syncytial virus vaccine (P less than 0.01), a protection rate of 68 per cent. The proportion of calves receiving treatment for respiratory disease was 38 per cent in the control group, 25 per cent in the calves inoculated with the quadrivalent vaccine (P less than 0.001) and 27 per cent in the calves inoculated with the respiratory syncytial virus vaccine (P less than 0.01). The results show that protection against respiratory disease can be achieved by parenteral vaccination of calves with the appropriate inactivated microorganisms.

  9. Brief report: respiratory syncytial virus activity--United States, 2005-2006.

    PubMed

    2006-12-01

    Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infections (LRTIs) (e.g., bronchiolitis and pneumonia) among young children in the United States. RSV also causes severe respiratory disease and a substantial number of deaths among older adults and persons with compromised respiratory, cardiac, or immune systems. RSV is transmitted person to person through close contact or inhalation of large droplets from a sneeze or cough; infection also can occur through contact with fomites (i.e., contaminated surfaces or objects). In temperate climates, peak RSV activity typically occurs during the winter. This report presents preliminary data on RSV activity reported to the National Respiratory and Enteric Virus Surveillance System (NREVSS) for the weeks ending July 8-November 18, 2006, indicating the onset of the 2006-2007 RSV season, and summarizes RSV trends during July 2005-June 2006. Health-care providers should consider RSV in the differential diagnosis for persons of all ages with LRTIs and implement appropriate isolation precautions to prevent nosocomial transmission from RSV-infected patients. Immune prophylaxis should be considered for certain infants and young children at high risk for complications from RSV infection (e.g., certain premature infants or infants and children with chronic lung and heart disease). PMID:17136023

  10. Respiratory syncytial virus infection enhances Pseudomonas aeruginosa biofilm growth through dysregulation of nutritional immunity.

    PubMed

    Hendricks, Matthew R; Lashua, Lauren P; Fischer, Douglas K; Flitter, Becca A; Eichinger, Katherine M; Durbin, Joan E; Sarkar, Saumendra N; Coyne, Carolyn B; Empey, Kerry M; Bomberger, Jennifer M

    2016-02-01

    Clinical observations link respiratory virus infection and Pseudomonas aeruginosa colonization in chronic lung disease, including cystic fibrosis (CF) and chronic obstructive pulmonary disease. The development of P. aeruginosa into highly antibiotic-resistant biofilm communities promotes airway colonization and accounts for disease progression in patients. Although clinical studies show a strong correlation between CF patients' acquisition of chronic P. aeruginosa infections and respiratory virus infection, little is known about the mechanism by which chronic P. aeruginosa infections are initiated in the host. Using a coculture model to study the formation of bacterial biofilm formation associated with the airway epithelium, we show that respiratory viral infections and the induction of antiviral interferons promote robust secondary P. aeruginosa biofilm formation. We report that the induction of antiviral IFN signaling in response to respiratory syncytial virus (RSV) infection induces bacterial biofilm formation through a mechanism of dysregulated iron homeostasis of the airway epithelium. Moreover, increased apical release of the host iron-binding protein transferrin during RSV infection promotes P. aeruginosa biofilm development in vitro and in vivo. Thus, nutritional immunity pathways that are disrupted during respiratory viral infection create an environment that favors secondary bacterial infection and may provide previously unidentified targets to combat bacterial biofilm formation.

  11. Brief report: respiratory syncytial virus activity--United States, 2005-2006.

    PubMed

    2006-12-01

    Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infections (LRTIs) (e.g., bronchiolitis and pneumonia) among young children in the United States. RSV also causes severe respiratory disease and a substantial number of deaths among older adults and persons with compromised respiratory, cardiac, or immune systems. RSV is transmitted person to person through close contact or inhalation of large droplets from a sneeze or cough; infection also can occur through contact with fomites (i.e., contaminated surfaces or objects). In temperate climates, peak RSV activity typically occurs during the winter. This report presents preliminary data on RSV activity reported to the National Respiratory and Enteric Virus Surveillance System (NREVSS) for the weeks ending July 8-November 18, 2006, indicating the onset of the 2006-2007 RSV season, and summarizes RSV trends during July 2005-June 2006. Health-care providers should consider RSV in the differential diagnosis for persons of all ages with LRTIs and implement appropriate isolation precautions to prevent nosocomial transmission from RSV-infected patients. Immune prophylaxis should be considered for certain infants and young children at high risk for complications from RSV infection (e.g., certain premature infants or infants and children with chronic lung and heart disease).

  12. Respiratory Syncytial Virus (RSV) RNA loads in peripheral blood correlates with disease severity in mice

    PubMed Central

    2010-01-01

    Background Respiratory Syncytial Virus (RSV) infection is usually restricted to the respiratory epithelium. Few studies have documented the presence of RSV in the systemic circulation, however there is no consistent information whether virus detection in the blood correlates with disease severity. Methods Balb/c mice were inoculated with live RSV, heat-inactivated RSV or medium. A subset of RSV-infected mice was treated with anti-RSV antibody 72 h post-inoculation. RSV RNA loads were measured by PCR in peripheral blood from day 1-21 post-inoculation and were correlated with upper and lower respiratory tract viral loads, the systemic cytokine response, lung inflammation and pulmonary function. Immunohistochemical staining was used to define the localization of RSV antigens in the respiratory tract and peripheral blood. Results RSV RNA loads were detected in peripheral blood from day 1 to 14 post-inoculation, peaked on day 5 and significantly correlated with nasal and lung RSV loads, airway obstruction, and blood CCL2 and CXCL1 expression. Treatment with anti-RSV antibody reduced blood RSV RNA loads and improved airway obstruction. Immunostaining identified RSV antigens in alveolar macrophages and peripheral blood monocytes. Conclusions RSV RNA was detected in peripheral blood upon infection with live RSV, followed a time-course parallel to viral loads assessed in the respiratory tract and was significantly correlated with RSV-induced airway disease. PMID:20843364

  13. Respiratory syncytial virus infection enhances Pseudomonas aeruginosa biofilm growth through dysregulation of nutritional immunity

    PubMed Central

    Hendricks, Matthew R.; Lashua, Lauren P.; Fischer, Douglas K.; Flitter, Becca A.; Eichinger, Katherine M.; Durbin, Joan E.; Sarkar, Saumendra N.; Coyne, Carolyn B.; Empey, Kerry M.; Bomberger, Jennifer M.

    2016-01-01

    Clinical observations link respiratory virus infection and Pseudomonas aeruginosa colonization in chronic lung disease, including cystic fibrosis (CF) and chronic obstructive pulmonary disease. The development of P. aeruginosa into highly antibiotic-resistant biofilm communities promotes airway colonization and accounts for disease progression in patients. Although clinical studies show a strong correlation between CF patients’ acquisition of chronic P. aeruginosa infections and respiratory virus infection, little is known about the mechanism by which chronic P. aeruginosa infections are initiated in the host. Using a coculture model to study the formation of bacterial biofilm formation associated with the airway epithelium, we show that respiratory viral infections and the induction of antiviral interferons promote robust secondary P. aeruginosa biofilm formation. We report that the induction of antiviral IFN signaling in response to respiratory syncytial virus (RSV) infection induces bacterial biofilm formation through a mechanism of dysregulated iron homeostasis of the airway epithelium. Moreover, increased apical release of the host iron-binding protein transferrin during RSV infection promotes P. aeruginosa biofilm development in vitro and in vivo. Thus, nutritional immunity pathways that are disrupted during respiratory viral infection create an environment that favors secondary bacterial infection and may provide previously unidentified targets to combat bacterial biofilm formation. PMID:26729873

  14. Prevalence of Rhinovirus and Respiratory Syncytial Virus Among Patients With Chronic Rhinosinusitis

    PubMed Central

    Abshirini, Hassan; Makvandi, Manochher; Seyyed Ashrafi, Meghdad; Hamidifard, Mojtaba; Saki, Nader

    2015-01-01

    Background: Chronic rhinosinusitis (CRS) is one of the most common chronic illnesses, but the etiology and pathogenesis of CRS are not well understood. Few studies have been carried out on the role of viruses in patients with chronic sinusitis so far. Objectives: Regarding the high number of patients, we intended to evaluate the prevalence of rhinovirus and respiratory syncytial virus in patients with CRS. Doing so, we may pave the way for definitely achieving the causes and factors of the disease and consequently definite treatment of this debilitating disease in future studies. Patients and Methods: This cross-sectional study was carried out on 76 patients. Sample of the study consisted of patients with CRS who were candidates for functional endoscopic sinus surgery (FESS). The specimens were collected during FESS between February 2013 and December 2013. For this purpose, after entering into sinuses, the specimens were collected from their mucus. They were then placed in Dulbecco's modified Eagle's viral transport medium (DMEM). They were transferred to the virology lab of the university in a cold chain. The specimens were maintained in -70°C before examination. Reverse transcriptase-polymerase chain reaction (RT-PCR) was applied to explore the presence of rhinovirus and respiratory syncytial virus. Results: In this study, 76 patients with rhinosinusitis underwent FESS as qualified candidates. The sample of the study consisted of 48 males (63.2%) and 28 females (36.8%) with the mean age of 44.3 years and an age range of 19-76 years. Among the 76 patients, 53 were with polyps and 23 without polyps. Among the patients, 66 were candidates of FESS for the first time. Other 11 patients had previously undergone the surgery. The results from PCR indicated that 22 (28.94%) patients had rhinovirus and 9 (11.84%) had respiratory syncytial virus (RSV). A total of 25 patients (32.89%) had one of the two viruses. In 6 (7.89%) specimens, both viruses were reported

  15. Mucosal IgG Levels Correlate Better with Respiratory Syncytial Virus Load and Inflammation than Plasma IgG Levels

    PubMed Central

    Vissers, Marloes; Ahout, Inge M. L.; de Jonge, Marien I.

    2015-01-01

    Maternal vaccination is currently considered a strategy against respiratory syncytial virus (RSV) infections. In RSV-infected infants, high mucosal IgG levels correlated better with reduced RSV load and lower mucosal CXCL10 levels than plasma IgG levels. For future vaccination strategies against RSV, more focus should be on the mucosal humoral immune response. PMID:26656116

  16. Diagnostic virology practices for respiratory syncytial virus and influenza virus among children in the hospital setting: a national survey.

    PubMed

    Jafri, Hasan S; Ramilo, Octavio; Makari, Doris; Charsha-May, Deborah; Romero, José R

    2007-10-01

    A survey was sent to the emergency room and laboratory directors of 400 randomly selected US hospitals to assess the diagnostic testing practices for respiratory syncytial virus and influenza virus in children. The results demonstrate that the majority of hospitals routinely perform viral testing for both viruses and use virology testing practices appropriate for the reasons reported for testing.

  17. Evaluation of a multiplex real-time polymerase chain reaction assay for the detection of influenza and respiratory syncytial viruses.

    PubMed

    Esposito, Susanna; Scala, Alessia; Tagliabue, Claudia; Zampiero, Alberto; Bianchini, Sonia; Principi, Nicola

    2016-01-01

    Nasopharyngeal swabs from 424 children were used to compare the performances of the new multiplex real-time polymerase chain reaction (RT-PCR) RIDA®GENE Flu & RSV kit and monospecific RT-PCR assays in detecting respiratory syncytial and influenza viruses. The easy-to-use kit was highly sensitive and specific and is recommended for routine practice.

  18. INHIBITION OF RESPIRATORY SYNCYTIAL VIRUS (RSV)-INDUCED INFLAMMATION BY 3-NITROTYROSINE IN HUMAN BRONCHIAL EPITHELIAL CELLS

    EPA Science Inventory

    Inhibition of Respiratory Syncytial Virus (RSV)-Induced Inflammation by 3-Nitrotyrosine in Human Bronchial Epithelial Cells. J. M. Soukup, MPH 1, ZW. Li, MD 2 and YC. T. Huang, MD 1. 1 NHEERL, US Environmental Protection Agency, RTP, NC and 2 CEMALB, University of North Carolina,...

  19. Molecular Evolutionary Dynamics of Respiratory Syncytial Virus Group A in Recurrent Epidemics in Coastal Kenya

    PubMed Central

    Agoti, Charles N.; Gitahi, Caroline W.; Bett, Ann; Ngama, Mwanajuma; Medley, Graham F.; Cane, Patricia A.; Nokes, D. James

    2016-01-01

    ABSTRACT The characteristic recurrent epidemics of human respiratory syncytial virus (RSV) within communities may result from the genetic variability of the virus and associated evolutionary adaptation, reducing the efficiency of preexisting immune responses. We analyzed the molecular evolutionary changes in the attachment (G) glycoprotein of RSV-A viruses collected over 13 epidemic seasons (2000 to 2012) in Kilifi (n = 649), Kenya, and contemporaneous sequences (n = 1,131) collected elsewhere within Kenya and 28 other countries. Genetic diversity in the G gene in Kilifi was dynamic both within and between epidemics, characterized by frequent new variant introductions and limited variant persistence between consecutive epidemics. Four RSV-A genotypes were detected in Kilifi: ON1 (11.9%), GA2 (75.5%), GA5 (12.3%), and GA3 (0.3%), with predominant genotype replacement of GA5 by GA2 and then GA2 by ON1. Within these genotypes, there was considerable variation in potential N-glycosylation sites, with GA2 and ON1 viruses showing up to 15 different patterns involving eight possible sites. Further, we identified 15 positively selected and 34 genotype-distinguishing codon sites, with six of these sites exhibiting both characteristics. The mean substitution rate of the G ectodomain for the Kilifi data set was estimated at 3.58 × 10−3 (95% highest posterior density interval = 3.04 to 4.16) nucleotide substitutions/site/year. Kilifi viruses were interspersed in the global phylogenetic tree, clustering mostly with Kenyan and European sequences. Our findings highlight ongoing genetic evolution and high diversity of circulating RSV-A strains, locally and globally, with potential antigenic differences. Taken together, these provide a possible explanation on the nature of recurrent local RSV epidemics. IMPORTANCE The mechanisms underlying recurrent epidemics of RSV are poorly understood. We observe high genetic diversity in circulating strains within and between epidemics in

  20. Neonatal Immunization with Respiratory Syncytial Virus Glycoprotein Fragment Induces Protective Immunity in the Presence of Maternal Antibodies in Mice

    PubMed Central

    Noh, Youran; Shim, Byoung-Shik; Cheon, In Su; Rho, Semi; Kim, Hee Joo; Choi, Youngjoo; Kang, Chang-Yuil; Chang, Jun

    2013-01-01

    Abstract Respiratory syncytial virus (RSV) is a major cause of severe lower respiratory tract infections in infants and the elderly worldwide. The significant morbidity and mortality associated with this infection underscores the urgent need for development of RSV vaccine. In this study, we first show that intranasal administration of RSV glycoprotein core fragment (Gcf) to neonatal mice can induce systemic humoral immune responses and protective immunity against RSV without causing lung eosinophilia, although antibody response was shifted to a Th2 response. Next, we examined whether the presence of maternal anti-RSV antibodies would affect the responsiveness and protection efficacy of Gcf in newborn mice, since infants can possess RSV-specific maternal antibodies due to frequent RSV re-infections to adults. Intranasal administration of Gcf induced antibody response and increased IFNγ secretion and protected mice against RSV challenge without severe lung eosinophilia, even in the presence of high levels of RSV-specific maternal antibodies. Thus, our findings suggest that Gcf may be an effective and safe RSV vaccine during the neonatal period. PMID:23869549

  1. A genetic model of differential susceptibility to human respiratory syncytial virus (RSV) infection

    PubMed Central

    Ciencewicki, Jonathan M.; Wang, Xuting; Marzec, Jacqui; Serra, M. Elina; Bell, Douglas A.; Polack, Fernando P.; Kleeberger, Steven R.

    2014-01-01

    Respiratory syncytial virus (RSV) is the primary cause of lower respiratory tract infection during childhood and causes severe symptoms in some patients, which may cause hospitalization and death. Mechanisms for differential responses to RSV are unknown. Our objective was to develop an in vitro model of RSV infection to evaluate interindividual variation in response to RSV and identify susceptibility genes. Populations of human-derived HapMap lymphoblastoid cell lines (LCLs) were infected with RSV. Compared with controls, RSV-G mRNA expression varied from ∼1- to 400-fold between LCLs. Basal expression of a number of gene transcripts, including myxovirus (influenza virus) resistance 1 (MX1), significantly correlated with RSV-G expression in HapMap LCLs. Individuals in a case-control population of RSV-infected children who were homozygous (n=94) or heterozygous (n=172) for the predicted deleterious A allele in a missense G/A SNP in MX1 had significantly greater risk for developing severe RSV disease relative to those with the major allele (n=108) (χ2=5.305, P=0.021; OR: 1.750, 95% CI: 1.110, 2.758, P=0.021). We conclude that genetically diverse human LCLs enable identification of susceptibility genes (e.g., MX1) for RSV disease severity in children, providing insight for disease risk.—Ciencewicki, J. M., Wang, X., Marzec, J., Serra, M. E., Bell, D. A., Polack, F. P., Kleeberger, S. R. A genetic model of differential susceptibility to human respiratory syncytial virus (RSV) infection. PMID:24421397

  2. [THE COMPARATIVE ANALYSIS OF EFFECTIVENESS OF QUICK TESTS IN DIAGNOSTIC OF INFLUENZA AND RESPIRATORY SYNCYTIAL VIRAL INFECTION IN CHILDREN].

    PubMed

    Petrova, E R; Sukhovetskaia, V P; Pisareva, M M; Maiorova, V G; Sverlova, M V; Danilenko, D M; Petrova, P A; Krivitskaia, V Z; Sominina, A A

    2015-11-01

    The analysis was implemented concerning diagnostic parameters of commercial quick tests (immune chromatographic tests BinaxNOW Influenza A&B and BinaxNow RSV Alere, Scarborough Inc., USA) under detection of antigens of influenza virus A and respiratory syncytial virus in clinical materials. The polymerase chain reaction in real-time and isolation ofviruses in cell cultures. The analysis of naso-pharyngeal smears from 116 children demonstrated that sensitivity and specifcity of detection of influenza virus A using device mariPOC in comparison with polymerase chain reaction made up to 93.8% and 99.0% correspondingly at total concordance of results of both techniques as 98.3%. At diagnosing of respiratory syncytial virus using device mariPOC parameters made up to 77.3%, 98.9% and 862% as compared with polymerase chain reaction. The sensitivity, specificity and total concordance of results of immune chromatographic tests BinaxNOW in comparison ofpolymerase chain reaction made up to 86.7%, 100% and 96.2% correspondingly at detection of influenza virus A and 80.9%, 97.4% and 91.6% correspondingly at detection of respiratory syncytial virus. In comparison with isolation technique in cell cultures sensitivity of system mariPOC and immune chromatographic tests proved to be in 1.3-1.4 times higher at detection of influenza virus A and in 1.7-2 times higher in case of isolation of respiratory syncytial virus. There is no statistically significant differences between diagnostic parameters received for mariPOC and immune chromatographic tests at diagnosing influenza virus A and respiratory syncytial viral infection. PMID:26999865

  3. Genetic Vaccine for Respiratory Syncytial Virus Provides Protection Without Disease Potentiation

    PubMed Central

    Johnson, Teresa R; Rangel, David; Graham, Barney S; Brough, Douglas E; Gall, Jason G

    2014-01-01

    Respiratory syncytial virus (RSV) is a major cause of infectious lower respiratory disease in infants and the elderly. As there is no vaccine for RSV, we developed a genetic vaccine approach that induced protection of the entire respiratory tract from a single parenteral administration. The approach was based on adenovirus vectors derived from newly isolated nonhuman primate viruses with low seroprevalence. We show for the first time that a single intramuscular (IM) injection of the replication-deficient adenovirus vectors expressing the RSV fusion (F0) glycoprotein induced immune responses that protected both the lungs and noses of cotton rats and mice even at low doses and for several months postimmunization. The immune response included high titers of neutralizing antibody that were maintained ≥24 weeks and RSV-specific CD8+ and CD4+ T cells. The vectors were as potently immunogenic as a human adenovirus 5 vector in these two key respiratory pathogen animal models. Importantly, there was minimal alveolitis and granulocytic infiltrates in the lung, and type 2 cytokines were not produced after RSV challenge even under conditions of partial protection. Overall, this genetic vaccine is highly effective without potentiating immunopathology, and the results support development of the vaccine candidate for human testing. PMID:23752342

  4. Reliable Detection of Respiratory Syncytial Virus Infection in Children for Adequate Hospital Infection Control Management

    PubMed Central

    Abels, Susanne; Nadal, David; Stroehle, Angelika; Bossart, Walter

    2001-01-01

    By using a rapid test for respiratory syncytial virus (RSV) detection (Abbott TestPack RSV), a number of patients were observed, showing repeatedly positive results over a period of up to 10 weeks. A prospective study was initiated to compare the rapid test with an antigen capture enzyme immunoassay (EIA) and a nested reverse transcriptase PCR (RT-PCR) protocol for detection of RSV serotypes A and B. Only respiratory samples from children exhibiting the prolonged presence of RSV (≥5 days) as determined by the rapid test were considered. A total of 134 specimens from 24 children was investigated by antigen capture EIA and nested RT-PCR. Using RT-PCR as the reference method, we determined the RSV rapid test to have a specificity of 63% and a sensitivity of 66% and the antigen capture EIA to have a specificity of 96% and a sensitivity of 69% for acute-phase samples and the homologous virus serotype A. In 7 (29%) of 24 patients, the positive results of the RSV rapid test could not be confirmed by either nested RT-PCR or antigen capture EIA. In these seven patients a variety of other respiratory viruses were detected. For general screening the RSV rapid test was found to be a reasonable tool to get quick results. However, its lack of specificity in some patients requires confirmation by additional tests to rule out false-positive results and/or detection of other respiratory viruses. PMID:11526141

  5. Identification and Characterization of the Binding Site of the Respiratory Syncytial Virus Phosphoprotein to RNA-Free Nucleoprotein

    PubMed Central

    Galloux, Marie; Gabiane, Gaëlle; Sourimant, Julien; Richard, Charles-Adrien; England, Patrick; Moudjou, Mohammed; Aumont-Nicaise, Magali; Fix, Jenna; Rameix-Welti, Marie-Anne

    2015-01-01

    ABSTRACT The RNA genome of respiratory syncytial virus (RSV) is constitutively encapsidated by the viral nucleoprotein N, thus forming a helical nucleocapsid. Polymerization of N along the genomic and antigenomic RNAs is concomitant to replication and requires the preservation of an unassembled monomeric nucleoprotein pool. To this end, and by analogy with Paramyxoviridae and Rhabdoviridae, it is expected that the viral phosphoprotein P acts as a chaperone protein, forming a soluble complex with the RNA-free form of N (N0-P complex). Here, we have engineered a mutant form of N that is monomeric, is unable to bind RNA, still interacts with P, and could thus mimic the N0 monomer. We used this N mutant, designated Nmono, as a substitute for N0 in order to characterize the P regions involved in the N0-P complex formation. Using a series of P fragments, we determined by glutathione S-transferase (GST) pulldown assays that the N and C termini of P are able to interact with Nmono. We analyzed the functional role of amino-terminal residues of P by site-directed mutagenesis, using an RSV polymerase activity assay based on a human RSV minireplicon, and found that several residues were critical for viral RNA synthesis. Using GST pulldown and surface plasmon resonance assays, we showed that these critical residues are involved in the interaction between P[1-40] peptide and Nmono in vitro. Finally, we showed that overexpression of the peptide P[1-29] can inhibit the polymerase activity in the context of the RSV minireplicon, thus demonstrating that targeting the N0-P interaction could constitute a potential antiviral strategy. IMPORTANCE Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract illness in infants. Since no vaccine or efficient antiviral treatment is available against RSV, it is essential to better understand how the viral machinery functions in order to develop new antiviral strategies. RSV phosphoprotein P, the main RNA polymerase

  6. Respiratory Syncytial Virus (RSV) Pulmonary Infection in Humanized Mice Induces Human Anti-RSV Immune Responses and Pathology

    PubMed Central

    Sharma, Anurag; Wu, Wenzhu; Sung, Biin; Huang, Jing; Tsao, Tiffany; Li, Xiangming; Gomi, Rika; Tsuji, Moriya

    2016-01-01

    ABSTRACT Respiratory syncytial virus (RSV) is a leading cause of lower respiratory tract disease, which causes high rates of morbidity and mortality in infants and the elderly. Models of human RSV pulmonary disease are needed to better understand RSV pathogenesis and to assess the efficacy of RSV vaccines. We assessed the RSV-specific human innate, humoral, and cellular immune responses in humanized mice (mice with a human immune system [HIS mice]) with functional human CD4+ T and B cells. These mice were generated by introduction of HLA class II genes, various human cytokines, and human B cell activation factor into immunodeficient NOD scid gamma (NSG) mice by the use of an adeno-associated virus vector, followed by engraftment of human hematopoietic stem cells. During the first 3 days of infection, HIS mice lost more weight and cleared RSV faster than NSG mice. Human chemokine (C-C motif) ligand 3 (CCL3) and human interleukin-1β (IL-1β) expression was detected in the RSV-infected HIS mice. The pathological features induced by RSV infection in HIS mice included peribronchiolar inflammation, neutrophil predominance in the bronchioalveolar lavage fluid, and enhanced airway mucus production. Human anti-RSV IgG and RSV-neutralizing antibodies were detected in serum and human anti-RSV mucosal IgA was detected in bronchioalveolar lavage fluid for up to 6 weeks. RSV infection induced an RSV-specific human gamma interferon response in HIS mouse splenocytes. These results indicate that human immune cells can induce features of RSV lung disease, including mucus hyperplasia, in murine lungs and that HIS mice can be used to elicit human anti-RSV humoral and cellular immunity. IMPORTANCE Infections with respiratory syncytial virus (RSV) are common and can cause severe lung disease in infants and the elderly. The lack of a suitable animal model with disease features similar to those in humans has hampered efforts to predict the efficacy of novel anti-RSV therapies and

  7. Discovery of Piperazinylquinoline Derivatives as Novel Respiratory Syncytial Virus Fusion Inhibitors.

    PubMed

    Zheng, Xiufang; Wang, Lisha; Wang, Baoxia; Miao, Kun; Xiang, Kunlun; Feng, Song; Gao, Lu; Shen, Hong C; Yun, Hongying

    2016-06-01

    A novel series of piperazinylquinoline derivatives were discovered as respiratory syncytial virus (RSV) fusion inhibitors by the ligand-based screening approach. Among 3,000 hits, 1-amino-3-[[2-(4-phenyl-1-piperidyl)-4-quinolyl]amino]propan-2-ol (7) was proven to be active against the RSV long (A) strain. The anti-RSV activity was improved by converting piperidine to benzylcarbonyl substituted piperazine. The basic side chain was also found to be crucial for anti-RSV activity. The selected analogues, 45 and 50, demonstrated anti-RSV activities up to EC50 = 0.028 μM and 0.033 μM, respectively. A direct anti-RSV effect was confirmed by a plaque reduction assay and a fusion inhibition assay. Both 45 and 50 showed promising DMPK properties with good oral bioavailability, and could potentially lead to novel therapeutic agents targeting the RSV fusion process. PMID:27326326

  8. Cell-mediated immune responses of lambs to challenge with bovine respiratory syncytial virus.

    PubMed Central

    Sharma, A K; Woldehiwet, Z

    1995-01-01

    The lamb is a good model to study the pathogenesis and immune responses to infections with respiratory syncytial virus (RSV) as lambs experimentally infected with bovine or human RSV may develop overt clinical disease. In the present study the development of cellular cytotoxic responses was studied in splenic, pulmonary and peripheral blood mononuclear cells obtained from lambs after primary and secondary infection with bovine RSV. Infection with bovine RSV was followed by the appearance of cytotoxic cells in the peripheral blood, the spleen and lung lavage fluids. These effector cells lysed virus-infected targets in a self-restricted manner. Depletion techniques revealed that cytotoxic activity was largely due to OvCD8+ cells. When effector cells obtained from primed lambs were stimulated with inactivated bovine RSV or with virus-infected cells in vitro, virus-specific cytotoxicity was significantly increased. PMID:7544251

  9. The host response and molecular pathogenesis associated with respiratory syncytial virus infection

    PubMed Central

    Oshansky, Christine M; Zhang, Wenliang; Moore, Elizabeth; Tripp, Ralph A

    2009-01-01

    Since the isolation of respiratory syncytial virus (RSV) in 1956, its significance as an important human pathogen in infants, the elderly and the immunocompromised has been established. Many important mechanisms contributing to RSV infection, replication and disease pathogenesis have been uncovered; however, there is still insufficient knowledge in these and related areas, which must be addressed to facilitate the development of safe and effective vaccines and therapeutic treatments. A better understanding of the molecular pathogenesis of RSV infection, particularly the host-cell response and transcription profiles to RSV infection, is required to advance disease intervention strategies. Substantial information is accumulating regarding how RSV proteins modulate molecular signaling and regulation of cytokine and chemokine responses to infection, molecular signals regulating programmed cell death, and innate and adaptive immune responses to infection. This review discusses RSV manipulation of the host response to infection and related disease pathogenesis. PMID:19327115

  10. Structure-Based Design of a Fusion Glycoprotein Vaccine for Respiratory Syncytial Virus

    PubMed Central

    McLellan, Jason S.; Chen, Man; Joyce, M. Gordon; Sastry, Mallika; Stewart-Jones, Guillaume B. E.; Yang, Yongping; Zhang, Baoshan; Chen, Lei; Srivatsan, Sanjay; Zheng, Anqi; Zhou, Tongqing; Graepel, Kevin W.; Kumar, Azad; Moin, Syed; Boyington, Jeffrey C.; Chuang, Gwo-Yu; Soto, Cinque; Baxa, Ulrich; Bakker, Arjen Q.; Spits, Hergen; Beaumont, Tim; Zheng, Zizheng; Xia, Ningshao; Ko, Sung-Youl; Todd, John-Paul; Rao, Srinivas; Graham, Barney S.; Kwong, Peter D.

    2015-01-01

    Respiratory syncytial virus (RSV) is the leading cause of hospitalization for children under five years of age. We sought to engineer a viral antigen that provides greater protection than currently available vaccines and focused on antigenic site Ø, a metastable site specific to the prefusion state of the RSV fusion (F) glycoprotein, as this site is targeted by extremely potent RSV-neutralizing antibodies. Structure-based design yielded stabilized versions of RSV F that maintained antigenic site Ø when exposed to extremes of pH, osmolality, and temperature. Six RSV F-crystal structures provided atomic-level data on how introduced cysteine residues and filled hydrophobic cavities improved stability. Immunization with site Ø-stabilized variants of RSV F in mice and macaques elicited levels of RSV-specific neutralizing activity many times the protective threshold. PMID:24179220

  11. Enzyme-linked immunosorbent assay for detection of respiratory syncytial virus infection: development and description.

    PubMed Central

    Hendry, R M; McIntosh, K

    1982-01-01

    An indirect enzyme-linked immunosorbent assay for detection of respiratory syncytial virus (RSV) antigens was developed, using commercially available antisera. Horse anti-RSV and calf antiserum to bovine RSV were used as capture and detector antibodies, respectively. The assay could detect as few as 50 PFU of unpurified RSV per ml in infected cell culture supernatant fluids and as little as 10 ng of affinity-purified RSV antigen per ml. No cross-reactions were observed with heterologous virus types. Freeze-thaw treatment had no effect on RSV enzyme-linked immunosorbent assay titers, but viral transport medium inhibited RSV enzyme-linked immunosorbent assay titers from 10- to 100-fold. The assay can be easily performed in 24 h and is a sensitive and specific method for the detection of RSV antigens. PMID:6749894

  12. Streptococcus pneumoniae Enhances Human Respiratory Syncytial Virus Infection In Vitro and In Vivo

    PubMed Central

    Nguyen, D. Tien; Louwen, Rogier; Elberse, Karin; van Amerongen, Geert; Yüksel, Selma; Luijendijk, Ad; Osterhaus, Albert D. M. E.; Duprex, W. Paul; de Swart, Rik L.

    2015-01-01

    Human respiratory syncytial virus (HRSV) and Streptococcus pneumoniae are important causative agents of respiratory tract infections. Both pathogens are associated with seasonal disease outbreaks in the pediatric population, and can often be detected simultaneously in infants hospitalized with bronchiolitis or pneumonia. It has been described that respiratory virus infections may predispose for bacterial superinfections, resulting in severe disease. However, studies on the influence of bacterial colonization of the upper respiratory tract on the pathogenesis of subsequent respiratory virus infections are scarce. Here, we have investigated whether pneumococcal colonization enhances subsequent HRSV infection. We used a newly generated recombinant subgroup B HRSV strain that expresses enhanced green fluorescent protein and pneumococcal isolates obtained from healthy children in disease-relevant in vitro and in vivo model systems. Three pneumococcal strains specifically enhanced in vitro HRSV infection of primary well-differentiated normal human bronchial epithelial cells grown at air-liquid interface, whereas two other strains did not. Since previous studies reported that bacterial neuraminidase enhanced HRSV infection in vitro, we measured pneumococcal neuraminidase activity in these cultures but found no correlation with the observed infection enhancement in our model. Subsequently, a selection of pneumococcal strains was used to induce nasal colonization of cotton rats, the best available small animal model for HRSV. Intranasal HRSV infection three days later resulted in strain-specific enhancement of HRSV replication in vivo. One S. pneumoniae strain enhanced HRSV both in vitro and in vivo, and was also associated with enhanced syncytium formation in vivo. However, neither pneumococci nor HRSV were found to spread from the upper to the lower respiratory tract, and neither pathogen was transmitted to naive cage mates by direct contact. These results demonstrate

  13. Respiratory Syncytial Virus Inhibitor AZ-27 Differentially Inhibits Different Polymerase Activities at the Promoter

    PubMed Central

    Noton, Sarah L.; Nagendra, Kartikeya; Dunn, Ewan F.; Mawhorter, Michael E.; Yu, Qin

    2015-01-01

    ABSTRACT Respiratory syncytial virus (RSV) is the leading cause of pediatric respiratory disease. RSV has an RNA-dependent RNA polymerase that transcribes and replicates the viral negative-sense RNA genome. The large polymerase subunit (L) has multiple enzymatic activities, having the capability to synthesize RNA and add and methylate a cap on each of the viral mRNAs. Previous studies (H. Xiong et al., Bioorg Med Chem Lett, 23:6789–6793, 2013, http://dx.doi.org/10.1016/j.bmcl.2013.10.018; C. L. Tiong-Yip et al., Antimicrob Agents Chemother, 58:3867–3873, 2014, http://dx.doi.org/10.1128/AAC.02540-14) had identified a small-molecule inhibitor, AZ-27, that targets the L protein. In this study, we examined the effect of AZ-27 on different aspects of RSV polymerase activity. AZ-27 was found to inhibit equally both mRNA transcription and genome replication in cell-based minigenome assays, indicating that it inhibits a step common to both of these RNA synthesis processes. Analysis in an in vitro transcription run-on assay, containing RSV nucleocapsids, showed that AZ-27 inhibits synthesis of transcripts from the 3′ end of the genome to a greater extent than those from the 5′ end, indicating that it inhibits transcription initiation. Consistent with this finding, experiments that assayed polymerase activity on the promoter showed that AZ-27 inhibited transcription and replication initiation. The RSV polymerase also can utilize the promoter sequence to perform a back-priming reaction. Interestingly, addition of AZ-27 had no effect on the addition of up to three nucleotides by back-priming but inhibited further extension of the back-primed RNA. These data provide new information regarding the mechanism of inhibition by AZ-27. They also suggest that the RSV polymerase adopts different conformations to perform its different activities at the promoter. IMPORTANCE Currently, there are no effective antiviral drugs to treat RSV infection. The RSV polymerase is an

  14. [Plasmid construction, expression, immunogenicity and protective efficacy of recombinant protein candidate vaccine of respiratory syncytial virus].

    PubMed

    Zeng, Rui-Hong; Gong, Wei; Fang, Xue-Ping; Zhang, Zhen-Ya; Mei, Xing-Guo

    2005-07-01

    To construct plasmid of recombinant protein candidate vaccine of respiratory syncytial virus, express it in E. coli, and to investigate its immunogenicity and protective efficacy. A CD8+ T cell epitope from respiratory syncytial virus (RSV) M2 protein F/M2:81 - 95 and the G:125-225 (G1) gene fragments from RSV-G protein containing B cell epitopes were amplified by PCR method and then inserted into the prokaryotic expression vector pET-DsbA after bonding to a linker. The fusion protein DsbA-G1-Linker-F/M2:81-95 (D-G1LF/M2) was expressed successfully in E. coli BL21 (DE3). The product was proved to be RSV-specific by Western-blot. After purified by affinity chromatography on Ni+ Sepharose and renatured by gradient dialysis. D-G1LF/M2 was used to immune BALB/c mice. D-G1LF/M2 induced high anti-D-G1LF/M2 IgG, anti-RSV IgG and neutralizing antibody titers in serum and lung of BALB/c mice, and elicied RSV-specific CTL responses. The IgG subclass distribution revealed that IgG1/IgG2a ratio was 2.66. Viral titration indicated that D-G1LF/M2 could protect BALB/c mice against RSV challenge in lung.

  15. Preventing severe respiratory syncytial virus disease: passive, active immunisation and new antivirals.

    PubMed

    Murray, Joanna; Saxena, Sonia; Sharland, Mike

    2014-05-01

    In most high-income countries palivizumab prophylaxis is considered safe, efficacious and cost-effective for preventing respiratory syncytial virus (RSV) hospital admissions among specific subgroups of infants born preterm, with chronic lung disease or with congenital heart disease. Virtually all babies acquire RSV during infancy and previously healthy babies are not eligible to receive palivizumab. Emerging evidence suggests some benefit of palivizumab use in reducing recurrent wheeze among infants born preterm. Better longitudinal studies are needed to examine its clinical and cost-effectiveness on recurrent and chronic respiratory illness and associated healthcare burden on resources in the community and hospitals. Since 99% of child deaths attributed to RSV occur in resource poor countries where expensive prophylaxis is not available or affordable, palivizumab has limited potential to impact on the current global burden of RSV lower respiratory tract infection (LRTI). A range of candidate vaccines for active immunisation against RSV are now in clinical trials. Two promising new antivirals are also currently in phase I/II trials to test their effectiveness in preventing severe RSV LRTI. These agents may be effective in preventing severe disease and phase III studies are in development. In the absence of effective active immunisation against RSV infection, population level approaches to prevent severe RSV LRTI should continue to focus on reducing prenatal and environmental risk factors including prematurity, smoking and improving hygiene practices. PMID:24464977

  16. Central nervous system alterations caused by infection with the human respiratory syncytial virus.

    PubMed

    Bohmwald, Karen; Espinoza, Janyra A; González, Pablo A; Bueno, Susan M; Riedel, Claudia A; Kalergis, Alexis M

    2014-11-01

    Worldwide, the human respiratory syncytial virus (hRSV) is the leading cause of infant hospitalization because of acute respiratory tract infections, including severe bronchiolitis and pneumonia. Despite intense research, to date there is neither vaccine nor treatment available to control hRSV disease burden globally. After infection, an incubation period of 3-5 days is usually followed by symptoms, such as cough and low-grade fever. However, hRSV infection can also produce a larger variety of symptoms, some of which relate to the individual's age at infection. Indeed, infants can display severe symptoms, such as dyspnea and chest wall retractions. Upon examination, crackles and wheezes are also common features that suggest infection by hRSV. Additionally, infection in infants younger than 1 year is associated with several non-specific symptoms, such as failure to thrive, periodic breathing or apnea, and feeding difficulties that usually require hospitalization. Recently, neurological symptoms have also been associated with hRSV respiratory infection and include seizures, central apnea, lethargy, feeding or swallowing difficulties, abnormalities in muscle tone, strabismus, abnormalities in the CSF, and encephalopathy. Here, we discuss recent findings linking the neurological, extrapulmonary effects of hRSV with infection and functional impairment of the CNS.

  17. Respiratory syncytial virus infection: denominator-based studies in Indonesia, Mozambique, Nigeria and South Africa.

    PubMed Central

    Robertson, Susan E.; Roca, Anna; Alonso, Pedro; Simoes, Eric A. F.; Kartasasmita, Cissy B.; Olaleye, David O.; Odaibo, Georgina N.; Collinson, Mark; Venter, Marietjie; Zhu, Yuwei; Wright, Peter F.

    2004-01-01

    OBJECTIVE: To assess the burden of respiratory syncytial virus (RSV)-associated lower respiratory infections (LRI) in children in four developing countries. METHODS: A WHO protocol for prospective population-based surveillance of acute respiratory infections in children aged less than 5 years was used at sites in Indonesia, Mozambique, Nigeria and South Africa. RSV antigen was identified by enzyme-linked immunosorbent assay performed on nasopharyngeal specimens from children meeting clinical case definitions. FINDINGS: Among children aged < 5 years, the incidence of RSV-associated LRI per 1000 child-years was 34 in Indonesia and 94 in Nigeria. The incidence of RSV-associated severe LRI per 1000 child-years was 5 in Mozambique, 10 in Indonesia, and 9 in South Africa. At all study sites, the majority of RSV cases occurred in infants. CONCLUSION: These studies demonstrate that RSV contributes to a substantial but quite variable burden of LRI in children aged < 5 years in four developing countries. The possible explanations for this variation include social factors, such as family size and patterns of seeking health care; the proportion of children infected by human immunodeficiency syndrome (HIV); and differences in clinical definitions used for obtaining samples. The age distribution of cases indicates the need for an RSV vaccine that can protect children early in life. PMID:15654405

  18. Immunobiotic Lactobacillus rhamnosus improves resistance of infant mice against respiratory syncytial virus infection.

    PubMed

    Chiba, Eriko; Tomosada, Yohsuke; Vizoso-Pinto, Maria Guadalupe; Salva, Susana; Takahashi, Takuya; Tsukida, Kohichiro; Kitazawa, Haruki; Alvarez, Susana; Villena, Julio

    2013-10-01

    Previously we showed that orally administered Lactobacillus rhamnosus CRL1505 beneficially regulated the balance between pro- and anti-inflammatory mediators in the lungs of poly(I:C)-challenged mice, allowing an effective inflammatory response against the TLR3/RIG-I agonist but at the same time reducing tissue damage. The aim of the present study was to investigate whether oral administration of the CRL1505 strain was able to improve resistance against respiratory syncytial virus (RSV) infection in infant mice and to evaluate the immunological mechanisms involved in the immunobiotic effect. We demonstrated that treatment of 3-week old BALB/c mice with L. rhamnosus CRL1505 significantly reduce lung viral loads and tissue injuries after the challenge with RSV. Moreover, we showed that the protective effect achieved by the CRL1505 strain is related to its capacity to differentially modulate respiratory antiviral immune response. Our results shows that IFN-γ and IL-10 secreted in response to L. rhamnosus CRL1505 oral stimulation would modulate the pulmonary innate immune microenvironment conducting to the activation of CD103(+) and CD11b(high) dendritic cells and the generation of CD3(+)CD4(+)IFN-γ(+) Th1 cells with the consequent attenuation of the strong and damaging Th2 reactions associated with RSV challenge. Our results indicate that modulation of the common mucosal immune system by immunobiotics could favor protective immunity against respiratory viral pathogens with a high attack rate in early infancy, such as RSV.

  19. Impact of severe disease caused by respiratory syncytial virus in children living in developed countries.

    PubMed

    Simoes, Eric A; Carbonell-Estrany, Xavier

    2003-02-01

    Among industrialized nations, the rate of rehospitalization in the United States for respiratory syncytial virus (RSV) is approximately 30 per 1000, exceptions being noted for American Indians and Alaskan natives, two ethnic groups who tend toward higher rates of RSV hospitalization. In distinction Japan reports an admission rate of 60 per 1000 for RSV disease. Yet Japan ranks considerably lower than many of its western counterparts in premature births. Whether an RSV subtype, a new viral genotype or some other unifying characteristic exists that might explain the severity of adenovirus, parainfluenza and RSV infections in this region of Asia remains to be determined. Outcomes trials in the United States, Canada, United Kingdom, Denmark and Japan all identified crowding and exposure to tobacco smoke as significant and independent risk factors for disease severity of RSV. The epidemiology of RSV is largely consistent throughout Europe, with peak outbreaks occurring in December and January. In Europe RSV accounts for 42 to 45% of hospital admissions for lower respiratory tract infections in children younger than 2 years of age, and inpatient populations tend to be younger and to experience greater disease severity. For RSV bronchiolitis lengths of stay in European hospitals range from a low of 4 days to a high of 10 days. The Infección Respiratoria Infantil por Virus Respiratorio Sincitial Study Group in Spain conducted 2 prospective observational studies in 14 and 26 neonatal units, respectively, on nonprophylaxed neonates to determine hospitalization rates for respiratory syncytial viral illness during 2 consecutive RSV seasons. Throughout each respiratory season the study group followed premature infants of < or =32 weeks gestational age at birth, representing an annual birth cohort of approximately 100 000 infants. A total of 584 infants who were < or =32 weeks gestational age in the first season and 999 in the second season were followed at monthly intervals

  20. Modification of the Respiratory Syncytial Virus F Protein in Virus-Like Particles Impacts Generation of B Cell Memory

    PubMed Central

    Schmidt, Madelyn R.; McGinnes-Cullen, Lori W.; Kenward, Sarah A.; Willems, Kristin N.; Woodland, Robert T.

    2014-01-01

    ABSTRACT Immunization with virus-like particles (VLPs) containing the Newcastle disease virus (NDV) core proteins, NP and M, and two chimera proteins (F/F and H/G) containing the respiratory syncytial virus (RSV) F- and G-protein ectodomains fused to the transmembrane and cytoplasmic domains of NDV F and HN proteins, respectively, stimulated durable RSV-neutralizing antibodies, F-protein-specific long-lived, bone marrow-associated plasma cells (LLPCs), and B cell memory, in striking contrast to RSV infection, which did not (M. R. Schmidt, L. W. McGinnes, S. A. Kenward, K. N. Willems, R. T. Woodland, and T. G. Morrison, J. Virol. 86:11654–11662, 2012). Here we report the characterization of a VLP with an RSV F-protein ectodomain fused to the NDV F-protein heptad repeat 2 (HR2), transmembrane, and cytoplasmic domain sequences, creating a chimera with two tandem HR2 domains, one from the RSV F protein and the other from the NDV F-protein ectodomain (F/HR2F). The F/HR2F chimera protein was efficiently assembled into VLPs along with the H/G chimera protein. This VLP (VLP-H/G+F/HR2F) stimulated anti-F-protein and anti-G-protein IgG, durable RSV-neutralizing antibodies, and anti-RSV F-protein-secreting LLPCs. However, the subtypes of anti-F-protein IgG induced were different from those elicited by VLPs containing the F/F chimera (VLP-H/G+F/F). Most importantly, VLP-H/G+F/HR2F did not induce RSV F-protein-specific B cell memory, as shown by the adoptive transfer of B cells from immunized animals to immunodeficient animals. The VLP did, however, induce B cell memory specific to the RSV G protein. Thus, the form of the F protein has a direct role in inducing anti-F-protein B cell memory. IMPORTANCE The development of vaccines for respiratory syncytial virus (RSV) is hampered by a lack of a clear understanding of the requirements for eliciting protective as well as durable human immune responses to virus antigens. The results of this study indicate that the form of the RSV

  1. Lamb Model of Respiratory Syncytial Virus–Associated Lung Disease: Insights to Pathogenesis and Novel Treatments

    PubMed Central

    Ackermann, Mark R.

    2014-01-01

    Preterm birth is a risk factor for respiratory syncytial virus (RSV) bronchiolitis and hospitalization. The pathogenesis underlying this is not fully understood, and in vivo studies are needed to better clarify essential cellular features and molecular mechanisms. Such studies include analysis of lung tissue from affected human infants and various animal models. The preterm and newborn lamb lung has developmental, structural, cellular, physiologic, and immunologic features similar to that of human infants. Also, the lamb lung is susceptible to various strains of RSV that infect infants and cause similar bronchiolar lesions. Studies in lambs suggest that viral replication in airways (especially bronchioles) is extensive by 4 days after infection, along with bronchiolitis characterized by degeneration and necrosis of epithelial cells, syncytial cell formation, neutrophil infiltration, epithelial cell hypertrophy and hyperplasia, and innate and adaptive immune responses. RSV bronchiolitis greatly affects airflow and gaseous exchange. RSV disease severity is increased in preterm lambs compared with full-term lambs; similar to human infants. The lamb is conducive to experimental assessment of novel, mechanistic therapeutic interventions such as delivery of vascular endothelial growth factor and enhancement of airway epithelial oxidative responses, Club (Clara) cell protein 10, and synthesized compounds such as nanobodies. In contrast, exposure of the fetal ovine lung in vivo to ethanol, a risk factor for preterm birth, reduces pulmonary alveolar development and surfactant protein A expression. Because the formalin-inactivated RSV vaccination enhances some inflammatory responses to RSV infection in lambs, this model has the potential to assess mechanisms of formalin-inactivated RSV enhanced disease as well as newly developed vaccines. PMID:24936027

  2. Molecular epidemiology of human respiratory syncytial virus over three consecutive seasons in Latvia.

    PubMed

    Balmaks, Reinis; Ribakova, Irina; Gardovska, Dace; Kazaks, Andris

    2014-11-01

    Lower respiratory tract infections caused by the human respiratory syncytial virus (HRSV) represent an immense burden of the disease, especially in young children. This study aimed to investigate the evolutionary history of HRSV strains isolated in the Children's Clinical University Hospital (Riga, Latvia) over three consecutive HRSV seasons. Of 207 samples from children hospitalized with lower respiratory tract infections, 88 (42.5%) tested positive for HRSV by RT-PCR. The seasonal activity started and peaked later than the average for the Northern hemisphere. Patients with HRSV lower respiratory tract infection were significantly younger than patients not infected with HRSV. HRSV-A viruses predominated for two consecutive seasons and were followed by an HRSV-B dominant season. Phylogenetic analysis based on glycoprotein G gene partial sequences revealed that viruses of both groups belonged to the worldwide dominant genotypes NA1 (HRSV-A) and BA-IV (HRSV-B). High diversity of this gene was driven only partially by selection pressure, as only two positively selected sites were identified in each group. Two of the HRSV-A isolates in this study contained a 72-nt duplication in the C-terminal end of the G gene (genotype ON1) that was first described in Canada in the 2010-2011 season. Initial spatial and temporal dynamics of this novel genotype were reconstructed by discrete phylogeographic analysis. Fifteen years after acquiring comparable 60-nt duplication in the G gene, genotype BA lineages have replaced all other HRSV-B strains. However, the population size of genotype ON1 plateaued soon and even decreased slightly before the beginning of the 2012-2013 season.

  3. Best practice in the prevention and management of paediatric respiratory syncytial virus infection

    PubMed Central

    Green, Christopher A.; Sande, Charles J.

    2016-01-01

    Respiratory syncytial virus (RSV) infection is ubiquitous with almost all infants having been infected by 2 years of age and lifelong repeated infections common. It is the second largest cause of mortality, after malaria, in infants outside the neonatal period and causes up to 200,000 deaths per year worldwide. RSV results in clinical syndromes that include upper respiratory tract infections, otitis media, bronchiolitis (up to 80% of cases) and lower respiratory tract disease including pneumonia and exacerbations of asthma or viral-induced wheeze. For the purposes of this review we will focus on RSV bronchiolitis in infants in whom the greatest disease burden lies. For infants requiring hospital admission, the identification of the causative respiratory virus is used to direct cohorting or isolation and infection control procedures to minimize nosocomial transmission. Nosocomial RSV infections are associated with poorer clinical outcomes, including increased mortality, the need for mechanical ventilation and longer length of hospital stay. Numerous clinical guidelines for the management of infants with bronchiolitis have been published, although none are specific for RSV bronchiolitis. Ribavirin is the only licensed drug for the specific treatment of RSV infection but due to drug toxicity and minimal clinical benefit it has not been recommended for routine clinical use. There is currently no licensed vaccine to prevent RSV infection but passive immunoprophylaxis using a monoclonal antibody, palivizumab, reduces the risk of hospitalization due to RSV infection by 39–78% in various high-risk infants predisposed to developing severe RSV disease. The current management of RSV bronchiolitis is purely supportive, with feeding support and oxygen supplementation until the infant immune system mounts a response capable of controlling the disease. The development of a successful treatment or prophylactic agent has the potential to revolutionize the care and outcome for

  4. Clinical Presentation and Birth Outcomes Associated with Respiratory Syncytial Virus Infection in Pregnancy

    PubMed Central

    Chu, Helen Y.; Katz, Joanne; Tielsch, James; Khatry, Subarna K.; Shrestha, Laxman; LeClerq, Steven C.; Magaret, Amalia; Kuypers, Jane; Steinhoff, Mark C.; Englund, Janet A.

    2016-01-01

    Background Respiratory syncytial virus (RSV) is the most important cause of viral pneumonia in children worldwide. A maternal vaccine may protect both the mother and infant from RSV illness. The epidemiology and clinical presentation of RSV in pregnant and postpartum women is not well-described. Methods Data were collected from a prospective, randomized trial of influenza immunization in pregnant women in rural southern Nepal. Women were enrolled in their second trimester of pregnancy and followed until six months postpartum. Active weekly home-based surveillance for febrile respiratory illness was performed. Mid-nasal swabs collected with episodes of respiratory illness were tested for RSV by real-time polymerase chain reaction. Results RSV was detected in 14 (0.4%) illness episodes in 3693 women over 3554 person-years of surveillance from 2011–2014. RSV incidence was 3.9/1000 person-years overall, and 11.8/1000 person-years between September and December. Seven (50%) women sought care for RSV illness; none died. Of the 7 (50%) illness episodes during pregnancy, all had live births with 2 (29%) preterm births and a median birthweight of 3060 grams. This compares to 469 (13%) preterm births and a median birthweight of 2790 grams in women without RSV during pregnancy. Of the 7 mothers with postpartum RSV infection, RSV was detected in 4 (57%) of their infants. Conclusions RSV was an uncommon cause of febrile respiratory illness in mothers during pregnancy in Nepal. These data will inform prevention and therapeutic strategies against RSV in resource-limited settings. PMID:27031702

  5. Relationship between meteorological conditions and respiratory syncytial virus in a tropical country.

    PubMed

    Rodriguez-Martinez, C E; Sossa-Briceño, M P; Acuña-Cordero, R

    2015-09-01

    This study aimed to determine which meteorological conditions are associated with respiratory syncytial virus (RSV) isolates in a population of children hospitalized with acute lower respiratory infection (ALRI) in Bogota, Colombia. In an analytical cross-sectional study, links were examined between the number of monthly RSV infections and monthly average climatic variation (temperature, relative humidity, rainfall, wind speed, solar radiation) between 1 January 2010 and 30 April 2011 in a population of hospitalized children aged <3 years with ALRI caused by RSV. Out of a total of 1548 children included in the study (mean age 9·2 ± 8·5 months), 1194 (77·1%) presented RSV infection during the 3-month period from March to May. In the multivariate analysis, after controlling for wind speed, relative humidity, and solar radiation, monthly average temperature [incident rate ratio (IRR) 3·14, 95% confidence interval (CI) 1·56-6·30, P = 0·001] and rainfall (IRR 1·008, 95% CI 1·00-1·01, P = 0·048) were independently associated with the monthly number of RSV infections. In conclusion, in Bogota, a tropical Latin American city, average temperature and rainfall are the meteorological variables most strongly associated with RSV isolation in children hospitalized with ALRI in the city.

  6. Respiratory syncytial virus seasonality in Brazil: implications for the immunisation policy for at-risk populations

    PubMed Central

    Freitas, André Ricardo Ribas; Donalisio, Maria Rita

    2016-01-01

    Respiratory syncytial virus (RSV) infection is the leading cause of hospitalisation for respiratory diseases among children under 5 years old. The aim of this study was to analyse RSV seasonality in the five distinct regions of Brazil using time series analysis (wavelet and Fourier series) of the following indicators: monthly positivity of the immunofluorescence reaction for RSV identified by virologic surveillance system, and rate of hospitalisations per bronchiolitis and pneumonia due to RSV in children under 5 years old (codes CID-10 J12.1, J20.5, J21.0 and J21.9). A total of 12,501 samples with 11.6% positivity for RSV (95% confidence interval 11 - 12.2), varying between 7.1 and 21.4% in the five Brazilian regions, was analysed. A strong trend for annual cycles with a stable stationary pattern in the five regions was identified through wavelet analysis of the indicators. The timing of RSV activity by Fourier analysis was similar between the two indicators analysed and showed regional differences. This study reinforces the importance of adjusting the immunisation period for high risk population with the monoclonal antibody palivizumab taking into account regional differences in seasonality of RSV. PMID:27120006

  7. Stability Characterization of a Vaccine Antigen Based on the Respiratory Syncytial Virus Fusion Glycoprotein

    PubMed Central

    Flynn, Jessica A.; Durr, Eberhard; Swoyer, Ryan; Cejas, Pedro J.; Horton, Melanie S.; Galli, Jennifer D.; Cosmi, Scott A.; Espeseth, Amy S.; Bett, Andrew J.; Zhang, Lan

    2016-01-01

    Infection with Respiratory Syncytial Virus (RSV) causes both upper and lower respiratory tract disease in humans, leading to significant morbidity and mortality in both young children and older adults. Currently, there is no licensed vaccine available, and therapeutic options are limited. During the infection process, the type I viral fusion (F) glycoprotein on the surface of the RSV particle rearranges from a metastable prefusion conformation to a highly stable postfusion form. In people naturally infected with RSV, most potent neutralizing antibodies are directed to the prefusion form of the F protein. Therefore, an engineered RSV F protein stabilized in the prefusion conformation (DS-Cav1) is an attractive vaccine candidate. Long-term stability at 4°C or higher is a desirable attribute for a commercial subunit vaccine antigen. To assess the stability of DS-Cav1, we developed assays using D25, an antibody which recognizes the prefusion F-specific antigenic site Ø, and a novel antibody 4D7, which was found to bind antigenic site I on the postfusion form of RSV F. Biophysical analysis indicated that, upon long-term storage at 4°C, DS-Cav1 undergoes a conformational change, adopting alternate structures that concomitantly lose the site Ø epitope and gain the ability to bind 4D7. PMID:27764150

  8. Respiratory syncytial virus is present in the neonatal intensive care unit.

    PubMed

    Homaira, Nusrat; Sheils, Joanne; Stelzer-Braid, Sacha; Lui, Kei; Oie, Ju-Lee; Snelling, Tom; Jaffe, Adam; Rawlinson, William

    2016-02-01

    Nosocomial transmission of respiratory syncytial virus (RSV) occurs in children within the neonatal intensive care unit (NICU). During peak community RSV transmission, three swabs were collected from the nose, hand and personal clothing of visitors and health care workers (HCW) in NICU once every week for eight weeks. Nasal swabs were collected from every third neonate and from any neonate clinically suspected of having a respiratory infection. Environmental sampling of high touch areas was done once during the study period. All swabs were tested for RSV using real time RT-PCR. There were 173 (519 total) and 109 (327 total) swabs, each of nose, hand and dress from 84 HCWs and 80 visitors respectively and 81 nasal swabs from 55 neonates collected. Thirty five environmental swabs from surfaces of the beds, side tables, counter tops, chairs, tables and computers were collected. Overall 1% of nasal swabs from each of HCWs, visitors and neonates, 4% of dress specimens from visitors and 9% of environmental swabs were positive for RSV-RNA. The results suggest that though the risk for RSV in the NICU remains low, personnel clothing are contaminated with RSV-RNA and may have a role in transmission. PMID:26174559

  9. Pre-fusion F is absent on the surface of formalin-inactivated respiratory syncytial virus

    PubMed Central

    Killikelly, April M.; Kanekiyo, Masaru; Graham, Barney S.

    2016-01-01

    The lack of a licensed vaccine for respiratory syncytial virus (RSV) can be partly attributed to regulatory hurdles resulting from vaccine enhanced respiratory disease (ERD) subsequent to natural RSV infection that was observed in clinical trials of formalin-inactivated RSV (FI-RSV) in antigen-naïve infants. To develop an effective vaccine that does not enhance RSV illness, it is important to understand how formalin and heat inactivation affected the antigenicity and immunogenicity of FI-RSV compared to native virus. Informed by atomic structures of RSV fusion (F) glycoprotein in prefusion (pre-F) and postfusion (post-F) conformations, we demonstrate that FI-RSV predominately presents post-F on the virion surface, whereas infectious RSV presents both pre-F and post-F conformations. This significant antigenic distinction has not been previously appreciated. Thus, a stabilized pre-F antigen is more representative of live RSV than F in its post-F conformation, as displayed on the surface of FI-RSV. This finding has major implications for discriminating current pre-F-based immunogens from FI-RSV used in historical vaccine trials. PMID:27682426

  10. Pulmonary surfactant phosphatidylglycerol inhibits respiratory syncytial virus–induced inflammation and infection

    PubMed Central

    Numata, Mari; Chu, Hong Wei; Dakhama, Azzeddine; Voelker, Dennis R.

    2009-01-01

    Respiratory syncytial virus (RSV) is the most common cause of hospitalization for respiratory tract infection in young children. It is also a significant cause of morbidity and mortality in elderly individuals and in persons with asthma and chronic obstructive pulmonary disease. Currently, no reliable vaccine or simple RSV antiviral therapy is available. Recently, we determined that the minor pulmonary surfactant phospholipid, palmitoyl-oleoyl-phosphatidylglycerol (POPG), could markedly attenuate inflammatory responses induced by lipopolysaccharide through direct interactions with the Toll-like receptor 4 (TLR4) interacting proteins CD14 and MD-2. CD14 and TLR4 have been implicated in the host response to RSV. Treatment of bronchial epithelial cells with POPG significantly inhibited interleukin-6 and -8 production, as well as the cytopathic effects induced by RSV. The phospholipid bound RSV with high affinity and inhibited viral attachment to HEp2 cells. POPG blocked viral plaque formation in vitro by 4 log units, and markedly suppressed the expansion of plaques from cells preinfected with the virus. Administration of POPG to mice, concomitant with viral infection, almost completely eliminated the recovery of virus from the lungs at 3 and 5 days after infection, and abrogated IFN-γ (IFN-γ) production and the enhanced expression of surfactant protein D (SP-D). These findings demonstrate an important approach to prevention and treatment of RSV infections using exogenous administration of a specific surfactant phospholipid. PMID:20080799

  11. Hospitalization for respiratory syncytial virus in the paediatric population in Spain.

    PubMed Central

    Vicente, D.; Montes, M.; Cilla, G.; Perez-Yarza, E. G.; Perez-Trallero, E.

    2003-01-01

    The aim of this population-based retrospective study was to determine the incidence of hospitalization for community-acquired, laboratory-confirmed respiratory syncytial virus (RSV) infection in an unselected paediatric population from southern Europe. The study was performed in an area with 15,700 children aged less than 5 years attended by a single hospital. The presence of RSV in nasopharyngeal aspirates from children with acute respiratory infection treated in the hospital was investigated in four seasons (July 1996-June 2000). A total of 390 episodes of hospitalization for RSV infection were detected and 83.3% of the children were aged less than 1 year old. The annual hospitalization rate was 37/1000 for infants aged less than 6 months and 25/1000 for those aged less than 1 year. During the study period, 2.5% of the infants younger than 1 year and approximately 5% of those younger than 3 months were hospitalized for RSV infection. The mean length of hospital stay was 5.9 days. Seven per cent of the patients required admission to the intensive care unit and more than half of these children were aged less than 1 month. In Spain, community-acquired RSV infection is a highly frequent cause of hospitalization in young children, especially in those aged less than 1 year. Prevention of RSV infection, through the development of vaccines and/or other strategies, should be a public health priority. PMID:14596527

  12. Relationship between meteorological conditions and respiratory syncytial virus in a tropical country.

    PubMed

    Rodriguez-Martinez, C E; Sossa-Briceño, M P; Acuña-Cordero, R

    2015-09-01

    This study aimed to determine which meteorological conditions are associated with respiratory syncytial virus (RSV) isolates in a population of children hospitalized with acute lower respiratory infection (ALRI) in Bogota, Colombia. In an analytical cross-sectional study, links were examined between the number of monthly RSV infections and monthly average climatic variation (temperature, relative humidity, rainfall, wind speed, solar radiation) between 1 January 2010 and 30 April 2011 in a population of hospitalized children aged <3 years with ALRI caused by RSV. Out of a total of 1548 children included in the study (mean age 9·2 ± 8·5 months), 1194 (77·1%) presented RSV infection during the 3-month period from March to May. In the multivariate analysis, after controlling for wind speed, relative humidity, and solar radiation, monthly average temperature [incident rate ratio (IRR) 3·14, 95% confidence interval (CI) 1·56-6·30, P = 0·001] and rainfall (IRR 1·008, 95% CI 1·00-1·01, P = 0·048) were independently associated with the monthly number of RSV infections. In conclusion, in Bogota, a tropical Latin American city, average temperature and rainfall are the meteorological variables most strongly associated with RSV isolation in children hospitalized with ALRI in the city. PMID:25591796

  13. Sirtuin 1 Regulates Dendritic Cell Activation and Autophagy during Respiratory Syncytial Virus-Induced Immune Responses.

    PubMed

    Owczarczyk, Anna B; Schaller, Matthew A; Reed, Michelle; Rasky, Andrew J; Lombard, David B; Lukacs, Nicholas W

    2015-08-15

    Respiratory syncytial virus (RSV) is the major cause of lower respiratory tract infection in children worldwide. Sirtuin 1 (SIRT1), an NAD(+)-dependent deacetylase, has been associated with the induction of autophagy and the regulation of inflammatory mediators. We found that Sirt1 was upregulated in mouse lung after RSV infection. Infected animals that received EX-527, a selective SIRT1 inhibitor, displayed exacerbated lung pathology, with increased mucus production, elevated viral load, and enhanced Th2 cytokine production. Gene expression analysis of isolated cell populations revealed that Sirt1 was most highly upregulated in RSV-treated dendritic cells (DCs). Upon RSV infection, EX-527-treated DCs, Sirt1 small interfering RNA-treated DCs, or DCs from conditional knockout (Sirt1(f/f)-CD11c-Cre(+)) mice showed downregulated inflammatory cytokine gene expression and attenuated autophagy. Finally, RSV infection of Sirt1(f/f)-CD11c-Cre(+) mice resulted in altered lung and lymph node cytokine responses, leading to exacerbated pathology. These data indicate that SIRT1 promotes DC activation associated with autophagy-mediated processes during RSV infection, thereby directing efficient antiviral immune responses. PMID:26157176

  14. Epidemiological and antigenic analysis of respiratory syncytial virus in hospitalised Tunisian children, from 2000 to 2002.

    PubMed

    Fodha, I; Vabret, A; Trabelsi, A; Freymuth, F

    2004-04-01

    Human respiratory syncytial virus (RSV) is the major viral cause of lower respiratory tract disease in children. Infections with the virus occur as annual winter epidemics in temperate climates, placing considerable pressure on the provision of hospital beds. Most molecular epidemiological studies have, until now, focused on isolates from infants in industrialised countries. No data have been available with regard to RSV strains from northern Africa. In this report, a recent RSV outbreak in Tunisia was studied and results showed that 176 of 815 (21.6%) nasopharyngeal aspirates collected from hospitalised children were RSV-positive by immunofluorescence assay. This RSV outbreak showed a temperature-dependent pattern (P=0.026) but no significant association with rainfall. A total of 73 RSV-positive samples were tested by two reverse-transcription-polymerase chain reaction assays (RT-PCR): RT-PCR-1, which amplifies the RNA of all RSV strains, and RT-PCR-2, which allows subgroup classification of RSV. Analysis by hybridisation assay of RT-PCR-2-amplified 1B protein gene products showed a higher prevalence of group B RSV than that of group A (82.5% vs. 17.5% among the typed strains). Knowledge of the variants is important in terms of both diagnosis and definition of a vaccine composition.

  15. Inhibition of respiratory syncytial virus replication and virus-induced p38 kinase activity by berberine.

    PubMed

    Shin, Han-Bo; Choi, Myung-Soo; Yi, Chae-Min; Lee, Jun; Kim, Nam-Jung; Inn, Kyung-Soo

    2015-07-01

    Respiratory syncytial virus (RSV) causes severe lower respiratory tract infection and poses a major public health threat worldwide. No effective vaccines or therapeutics are currently available; berberine, an isoquinoline alkaloid from various medicinal plants, has been shown to exert antiviral and several other biological effects. Recent studies have shown that p38 mitogen-activated protein kinase (MAPK) activity is implicated in infection by and replication of viruses such as RSV and the influenza virus. Because berberine has previously been implicated in modulating the activity of p38 MAPK, its effects on RSV infection and RSV-mediated p38 MAPK activation were examined. Replication of RSV in epithelial cells was significantly reduced by treatment with berberine. Berberine treatment caused decrease in viral protein and mRNA syntheses. Similar to previously reported findings, RSV infection caused phosphorylation of p38 MAPK at a very early time point of infection, and phosphorylation was dramatically reduced by berberine treatment. In addition, production of interleukin-6 mRNA upon RSV infection was significantly suppressed by treatment with berberine, suggesting the anti-inflammatory role of berberine during RSV infection. Taken together, we showed that berberine, a natural compound already proven to be safe for human consumption, suppresses the replication of RSV. In addition, the current study suggests that inhibition of RSV-mediated early p38 MAPK activation, which has been implicated as an early step in viral infection, as a potential molecular mechanism.

  16. Respiratory Syncytial Virus and Cellular Stress Responses: Impact on Replication and Physiopathology

    PubMed Central

    Cervantes-Ortiz, Sandra L.; Zamorano Cuervo, Natalia; Grandvaux, Nathalie

    2016-01-01

    Human respiratory syncytial virus (RSV), a member of the Paramyxoviridae family, is a major cause of severe acute lower respiratory tract infection in infants, elderly and immunocompromised adults. Despite decades of research, a complete integrated picture of RSV-host interaction is still missing. Several cellular responses to stress are involved in the host-response to many virus infections. The endoplasmic reticulum stress induced by altered endoplasmic reticulum (ER) function leads to activation of the unfolded-protein response (UPR) to restore homeostasis. Formation of cytoplasmic stress granules containing translationally stalled mRNAs is a means to control protein translation. Production of reactive oxygen species is balanced by an antioxidant response to prevent oxidative stress and the resulting damages. In recent years, ongoing research has started to unveil specific regulatory interactions of RSV with these host cellular stress responses. Here, we discuss the latest findings regarding the mechanisms evolved by RSV to induce, subvert or manipulate the ER stress, the stress granule and oxidative stress responses. We summarize the evidence linking these stress responses with the regulation of RSV replication and the associated pathogenesis. PMID:27187445

  17. Biophysical characterization of G protein ectodomain of group B human respiratory syncytial virus from E. coli.

    PubMed

    Khan, Wajihul Hasan; Srungaram, V L N Raghuram; Islam, Asimul; Beg, Ilyas; Haider, Md Shakir H; Ahmad, Faizan; Broor, Shobha; Parveen, Shama

    2016-07-01

    Human respiratory syncytial virus (hRSV) is an important pathogen of acute respiratory tract infection. The G protein of hRSV is a transmembrane glycoprotein that is a neutralizing antigen and is thus a vaccine candidate. In this study, synthetic codon optimized ectodomain G protein [G(ΔTM)] of BA genotype of group B hRSV was cloned, expressed, and characterized using biophysical techniques. The molar absorption coefficient and mean residue ellipticity at 222 nm ([θ]222) of G (ΔTM) was found to be 7950 M(-1) cm(-1) and -19701.7 deg cm(2) dmol(-1) respectively. It was concluded that G(ΔTM) mainly consist of α-helix (74.9%) with some amount of β-sheet (4%). The protein was stable up to 85°C without any transition curve. However, heat-induced denaturation of G(ΔTM) resulted in total loss of β-sheet whereas not much change was observed in the α-helix part of the secondary structure. It was concluded that G(ΔTM) is an α-helical protein and it is highly stable at high temperature, but could be easily denatured using high concentrations of GdmCl/urea or acidic condition. This is the first investigation of cloning, expression, and characterization of G(ΔTM) of BA viruses from India. Structural characterization of G protein will assist in drug designing and vaccine development for hRSV.

  18. Exploring the dynamics of respiratory syncytial virus (RSV) transmission in children.

    PubMed

    Hogan, Alexandra B; Glass, Kathryn; Moore, Hannah C; Anderssen, Robert S

    2016-08-01

    Respiratory syncytial virus (RSV) is the main cause of lower respiratory tract infections in children. Whilst highly seasonal, RSV dynamics can have either one-year (annual) or two-year (biennial) cycles. Furthermore, some countries show a 'delayed biennial' pattern, where the epidemic peak in low incidence years is delayed. We develop a compartmental model for RSV infection, driven by a seasonal forcing function, and conduct parameter space and bifurcation analyses to document parameter ranges that give rise to these different seasonal patterns. The model is sensitive to the birth rate, transmission rate, and seasonality parameters, and can replicate RSV dynamics observed in different countries. The seasonality parameter must exceed a threshold for the model to produce biennial cycles. Intermediate values of the birth rate produce the greatest delay in these biennial cycles, while the model reverts to annual cycles if the duration of immunity is too short. Finally, the existence of period doubling and period halving bifurcations suggests robust model dynamics, in agreement with the known regularity of RSV outbreaks. These findings help explain observed RSV data, such as regular biennial dynamics in Western Australia, and delayed biennial dynamics in Finland. From a public health perspective, our findings provide insight into the drivers of RSV transmission, and a foundation for exploring RSV interventions. PMID:27155294

  19. A Cross-Study Biomarker Signature of Human Bronchial Epithelial Cells Infected with Respiratory Syncytial Virus.

    PubMed

    Gardinassi, Luiz Gustavo

    2016-01-01

    Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infections in children, elderly, and immunocompromised individuals. Despite of advances in diagnosis and treatment, biomarkers of RSV infection are still unclear. To understand the host response and propose signatures of RSV infection, previous studies evaluated the transcriptional profile of the human bronchial epithelial cell line-BEAS-2B-infected with different strains of this virus. However, the evolution of statistical methods and functional analysis together with the large amount of expression data provide opportunities to uncover novel biomarkers of inflammation and infections. In view of those facts publicly available microarray datasets from RSV-infected BEAS-2B cells were analyzed with linear model-based statistics and the platform for functional analysis InnateDB. The results from those analyses argue for the reevaluation of previously reported transcription patterns and biological pathways in BEAS-2B cell lines infected with RSV. Importantly, this study revealed a biosignature constituted by genes such as ABCC4, ARMC8, BCLAF1, EZH1, FAM118A, FAM208B, FUS, HSPH1, KAZN, MAP3K2, N6AMT1, PRMT2, S100PBP, SERPINA1, TLK2, ZNF322, and ZNF337 which should be considered in the development of new molecular diagnosis tools. PMID:27274726

  20. Respiratory Syncytial Virus and Cellular Stress Responses: Impact on Replication and Physiopathology.

    PubMed

    Cervantes-Ortiz, Sandra L; Zamorano Cuervo, Natalia; Grandvaux, Nathalie

    2016-01-01

    Human respiratory syncytial virus (RSV), a member of the Paramyxoviridae family, is a major cause of severe acute lower respiratory tract infection in infants, elderly and immunocompromised adults. Despite decades of research, a complete integrated picture of RSV-host interaction is still missing. Several cellular responses to stress are involved in the host-response to many virus infections. The endoplasmic reticulum stress induced by altered endoplasmic reticulum (ER) function leads to activation of the unfolded-protein response (UPR) to restore homeostasis. Formation of cytoplasmic stress granules containing translationally stalled mRNAs is a means to control protein translation. Production of reactive oxygen species is balanced by an antioxidant response to prevent oxidative stress and the resulting damages. In recent years, ongoing research has started to unveil specific regulatory interactions of RSV with these host cellular stress responses. Here, we discuss the latest findings regarding the mechanisms evolved by RSV to induce, subvert or manipulate the ER stress, the stress granule and oxidative stress responses. We summarize the evidence linking these stress responses with the regulation of RSV replication and the associated pathogenesis. PMID:27187445

  1. Alveolar Macrophages Can Control Respiratory Syncytial Virus Infection in the Absence of Type I Interferons.

    PubMed

    Makris, Spyridon; Bajorek, Monika; Culley, Fiona J; Goritzka, Michelle; Johansson, Cecilia

    2016-01-01

    Respiratory syncytial virus (RSV) is a common cause of lower respiratory tract infections. Immunity to RSV is initiated upon detection of the virus by pattern recognition receptors, such as RIG-I-like receptors. RIG-I-like receptors signal via MAVS to induce the synthesis of proinflammatory mediators, including type I interferons (IFNs), which trigger and shape antiviral responses and protect cells from infection. Alveolar macrophages (AMs) are amongst the first cells to encounter invading viruses and the ones producing type I IFNs. However, it is unclear whether IFNs act to prevent AMs from serving as vehicles for viral replication. In this study, primary AMs from MAVS (Mavs-/-)- or type I IFN receptor (Ifnar1-/-)-deficient mice were exposed to RSV ex vivo. Wild-type (wt) AMs but not Mavs-/- and Ifnar1-/- AMs produced inflammatory mediators in response to RSV. Furthermore, Mavs-/- and Ifnar1-/- AMs accumulated more RSV proteins than wt AMs, but the infection was abortive. Thus, RIG-I-like receptor-MAVS and IFNAR signalling are important for the induction of proinflammatory mediators from AMs upon RSV infection, but this signalling is not central for controlling viral replication. The ability to restrict viral replication makes AMs ideal sensors of RSV infection and important initiators of immune responses in the lung. PMID:27423203

  2. Wheezing, asthma, and pulmonary dysfunction 10 years after infection with respiratory syncytial virus in infancy.

    PubMed Central

    Pullan, C R; Hey, E N

    1982-01-01

    Of the 180 children admitted to hospitals in Tyneside in the first year of life with proved respiratory syncytial virus lower respiratory tract infection, 130 were seen for review 10 years later and 34 of the remaining 50 children accounted for. Skin tests, lung function tests, and histamine-challenge and exercise tests for bronchial lability were undertaken in over 100 of the index children and a similar number of control children. A total of 55 (42%) of the 130 index children had had further episodes of wheeze, while only 21 (19%) out of 111 controls had ever wheezed; but few (6.2% v 4.5%) had troublesome symptoms at the age of 10. There was a threefold increase in the incidence of bronchial lability in the index children but no excess of atopy. Maximum expiratory air flow was reduced throughout the vital capacity manoeuvre in the index children, even when those with a history of recurrent wheeze were excluded. Results of single-breath nitrogen washout tests were normal, however, suggesting that ventilation was not appreciably uneven, even though expiratory flow was restricted. These differences might have been caused by infection damaging the growing lung but might also be explained by pre-existing differences in the airway, rendering certain children more susceptible to symptomatic infection when first challenged by the virus in infancy. PMID:6805648

  3. A sensitive real-time PCR for detection and subgrouping of human respiratory syncytial virus

    PubMed Central

    Do, Lien Anh Ha; van Doorn, H. Rogier; Bryant, Juliet E.; Nghiem, My Ngoc; Nguyen Van, Vinh Chau; Vo, Cong Khanh; Nguyen, Minh Dung; Tran, Tinh Hien; Farrar, Jeremy; de Jong, Menno D.

    2012-01-01

    Improved diagnostic tools for rapid detection, quantitation, and subgrouping of human respiratory syncytial virus (RSV) are needed to aid the development and evaluation of novel intervention strategies. A quantitative real-time RT-PCR using specific locked nucleic acid (LNA) probes was developed to identify RSV and to distinguish RSV subgroups A and B (RSV LNA assay). RSV subgroup diversity and the relationship between viral load and disease severity in confirmed RSV infections were also explored. 264 archived respiratory specimens from pediatric patients were tested in parallel using the commercial multiplex Seeplex™ RV detection kit (Seegene) and the novel RSV LNA assay. The LNA assay demonstrated a significantly higher sensitivity than Seeplex, improving overall detection rates from 24% (64/264) to 32% (84/264). Detection limits of 9.0 × 101 and 6.0 × 102 copies/mL were observed for RSV A and B, respectively. RSV A was detected in 53/84 (63%) cases, and 31/84 (37%) were positive for RSV B. This novel method offers a rapid, quantitative, highly specific and sensitive approach to laboratory diagnosis of RSV. PMID:22119628

  4. Sirtuin 1 Regulates Dendritic Cell Activation and Autophagy during Respiratory Syncytial Virus-Induced Immune Responses.

    PubMed

    Owczarczyk, Anna B; Schaller, Matthew A; Reed, Michelle; Rasky, Andrew J; Lombard, David B; Lukacs, Nicholas W

    2015-08-15

    Respiratory syncytial virus (RSV) is the major cause of lower respiratory tract infection in children worldwide. Sirtuin 1 (SIRT1), an NAD(+)-dependent deacetylase, has been associated with the induction of autophagy and the regulation of inflammatory mediators. We found that Sirt1 was upregulated in mouse lung after RSV infection. Infected animals that received EX-527, a selective SIRT1 inhibitor, displayed exacerbated lung pathology, with increased mucus production, elevated viral load, and enhanced Th2 cytokine production. Gene expression analysis of isolated cell populations revealed that Sirt1 was most highly upregulated in RSV-treated dendritic cells (DCs). Upon RSV infection, EX-527-treated DCs, Sirt1 small interfering RNA-treated DCs, or DCs from conditional knockout (Sirt1(f/f)-CD11c-Cre(+)) mice showed downregulated inflammatory cytokine gene expression and attenuated autophagy. Finally, RSV infection of Sirt1(f/f)-CD11c-Cre(+) mice resulted in altered lung and lymph node cytokine responses, leading to exacerbated pathology. These data indicate that SIRT1 promotes DC activation associated with autophagy-mediated processes during RSV infection, thereby directing efficient antiviral immune responses.

  5. Retrospective Parameter Estimation and Forecast of Respiratory Syncytial Virus in the United States

    PubMed Central

    Shaman, Jeffrey

    2016-01-01

    Recent studies have shown that systems combining mathematical modeling and Bayesian inference methods can be used to generate real-time forecasts of future infectious disease incidence. Here we develop such a system to study and forecast respiratory syncytial virus (RSV). RSV is the most common cause of acute lower respiratory infection and bronchiolitis. Advanced warning of the epidemic timing and volume of RSV patient surges has the potential to reduce well-documented delays of treatment in emergency departments. We use a susceptible-infectious-recovered (SIR) model in conjunction with an ensemble adjustment Kalman filter (EAKF) and ten years of regional U.S. specimen data provided by the Centers for Disease Control and Prevention. The data and EAKF are used to optimize the SIR model and i) estimate critical epidemiological parameters over the course of each outbreak and ii) generate retrospective forecasts. The basic reproductive number, R0, is estimated at 3.0 (standard deviation 0.6) across all seasons and locations. The peak magnitude of RSV outbreaks is forecast with nearly 70% accuracy (i.e. nearly 70% of forecasts within 25% of the actual peak), four weeks before the predicted peak. This work represents a first step in the development of a real-time RSV prediction system. PMID:27716828

  6. A Cross-Study Biomarker Signature of Human Bronchial Epithelial Cells Infected with Respiratory Syncytial Virus

    PubMed Central

    Gardinassi, Luiz Gustavo

    2016-01-01

    Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infections in children, elderly, and immunocompromised individuals. Despite of advances in diagnosis and treatment, biomarkers of RSV infection are still unclear. To understand the host response and propose signatures of RSV infection, previous studies evaluated the transcriptional profile of the human bronchial epithelial cell line—BEAS-2B—infected with different strains of this virus. However, the evolution of statistical methods and functional analysis together with the large amount of expression data provide opportunities to uncover novel biomarkers of inflammation and infections. In view of those facts publicly available microarray datasets from RSV-infected BEAS-2B cells were analyzed with linear model-based statistics and the platform for functional analysis InnateDB. The results from those analyses argue for the reevaluation of previously reported transcription patterns and biological pathways in BEAS-2B cell lines infected with RSV. Importantly, this study revealed a biosignature constituted by genes such as ABCC4, ARMC8, BCLAF1, EZH1, FAM118A, FAM208B, FUS, HSPH1, KAZN, MAP3K2, N6AMT1, PRMT2, S100PBP, SERPINA1, TLK2, ZNF322, and ZNF337 which should be considered in the development of new molecular diagnosis tools. PMID:27274726

  7. Mammalian Cell-Derived Respiratory Syncytial Virus-Like Particles Protect the Lower as well as the Upper Respiratory Tract

    PubMed Central

    Walpita, Pramila; Johns, Lisa M.; Tandon, Ravi; Moore, Martin L.

    2015-01-01

    Globally, Respiratory Syncytial Virus (RSV) is a leading cause of bronchiolitis and pneumonia in children less than one year of age and in USA alone, between 85,000 and 144,000 infants are hospitalized every year. To date, there is no licensed vaccine. We have evaluated vaccine potential of mammalian cell-derived native RSV virus-like particles (RSV VLPs) composed of the two surface glycoproteins G and F, and the matrix protein M. Results of in vitro testing showed that the VLPs were functionally assembled and immunoreactive, and that the recombinantly expressed F protein was cleaved intracellularly similarly to the virus-synthesized F protein to produce the F1 and F2 subunits; the presence of the F1 fragment is critical for vaccine development since all the neutralizing epitopes present in the F protein are embedded in this fragment. Additional in vitro testing in human macrophage cell line THP-1 showed that both virus and the VLPs were sensed by TLR-4 and induced a Th1-biased cytokine response. Cotton rats vaccinated with RSV VLPs adjuvanted with alum and monophosphoryl lipid A induced potent neutralizing antibody response, and conferred protection in the lower as well as the upper respiratory tract based on substantial virus clearance from these sites. To the best of our knowledge, this is the first VLP/virosome vaccine study reporting protection of the lower as well as the upper respiratory tract: Prevention from replication in the nose is an important consideration if the target population is infants < 6 months of age. This is because continued virus replication in the nose results in nasal congestion and babies at this age are obligate nose breathers. In conclusion, these results taken together suggest that our VLPs show promise to be a safe and effective vaccine for RSV. PMID:26172453

  8. CT findings in viral lower respiratory tract infections caused by parainfluenza virus, influenza virus and respiratory syncytial virus.

    PubMed

    Kim, Min-Chul; Kim, Mi Young; Lee, Hyun Joo; Lee, Sang-Oh; Choi, Sang-Ho; Kim, Yang Soo; Woo, Jun Hee; Kim, Sung-Han

    2016-06-01

    Viral lower respiratory tract infections (LRTIs) can present with a variety of computed tomography (CT) findings. However, identifying the contribution of a particular virus to CT findings is challenging due to concomitant infections and the limited data on the CT findings in viral LRTIs. We therefore investigate the CT findings in different pure viral LRTIs.All patients who underwent bronchoalveolar lavage (BAL) and were diagnosed with LRTIs caused by parainfluenza virus (PIV), influenza virus, or respiratory syncytial virus (RSV) between 1998 and 2014 were enrolled in a tertiary hospital in Seoul, South Korea. A pure viral LRTI was defined as a positive viral culture from BAL without any positive evidence from respiratory or blood cultures, or from polymerase chain reaction (PCR), or from serologic tests for bacteria, fungi, mycobacteria, or other viruses.CT images of 40 patients with viral LRTIs were analyzed: 14 with PIV, 14 with influenza virus, and 12 with RSV. Patch consolidation (≥1 cm or more than 1 segmental level) was found only in PIV (29%) (P = 0.03), by which CT findings caused by PIV could resemble those seen in bacterial LRTIs. Ground-glass opacities were seen in all cases of influenza virus and were more frequent than in PIV (71%) and RSV (67%) (P = 0.05). Bronchial wall thickening was more common in influenza virus (71%) and RSV (67%) LRTIs than PIV LRTIs (21%) (P = 0.02). With respect to anatomical distribution, PIV infections generally affected the lower lobes (69%), while influenza virus mostly caused diffuse changes throughout the lungs (57%), and RSV frequently formed localized patterns in the upper and mid lobes (44%).The CT findings in LRTIs of PIV, influenza virus, and RSV can be distinguished by certain characteristics. These differences could be useful for early differentiation of these viral LRTIs, and empirical use of appropriate antiviral agents. PMID:27368011

  9. Molecular Characterization of Human Respiratory Syncytial Virus in the Philippines, 2012-2013

    PubMed Central

    Malasao, Rungnapa; Okamoto, Michiko; Chaimongkol, Natthawan; Imamura, Tadatsugu; Tohma, Kentaro; Dapat, Isolde; Dapat, Clyde; Suzuki, Akira; Saito, Mayuko; Saito, Mariko; Tamaki, Raita; Pedrera-Rico, Gay Anne Granada; Aniceto, Rapunzel; Quicho, Reynaldo Frederick Negosa; Segubre-Mercado, Edelwisa; Lupisan, Socorro; Oshitani, Hitoshi

    2015-01-01

    Human respiratory syncytial virus (HRSV) is a major cause of acute lower respiratory tract infections in infants and children worldwide. We performed molecular analysis of HRSV among infants and children with clinical diagnosis of severe pneumonia in four study sites in the Philippines, including Biliran, Leyte, Palawan, and Metro Manila from June 2012 to July 2013. Nasopharyngeal swabs were collected and screened for HRSV using real-time polymerase chain reaction (PCR). Positive samples were tested by conventional PCR and sequenced for the second hypervariable region (2nd HVR) of the G gene. Among a total of 1,505 samples, 423 samples were positive for HRSV (28.1%), of which 305 (72.1%) and 118 (27.9%) were identified as HRSV-A and HRSV-B, respectively. Two genotypes of HRSV-A, NA1 and ON1, were identified during the study period. The novel ON1 genotype with a 72-nucleotide duplication in 2nd HVR of the G gene increased rapidly and finally became the predominant genotype in 2013 with an evolutionary rate higher than the NA1 genotype. Moreover, in the ON1 genotype, we found positive selection at amino acid position 274 (p<0.05) and massive O- and N-glycosylation in the 2nd HVR of the G gene. Among HRSV-B, BA9 was the predominant genotype circulating in the Philippines. However, two sporadic cases of GB2 genotype were found, which might share a common ancestor with other Asian strains. These findings suggest that HRSV is an important cause of severe acute respiratory infection among children in the Philippines and revealed the emergence and subsequent predominance of the ON1 genotype and the sporadic detection of the GB2 genotype. Both genotypes were detected for the first time in the Philippines. PMID:26540236

  10. Molecular Characterization of Human Respiratory Syncytial Virus in the Philippines, 2012-2013.

    PubMed

    Malasao, Rungnapa; Okamoto, Michiko; Chaimongkol, Natthawan; Imamura, Tadatsugu; Tohma, Kentaro; Dapat, Isolde; Dapat, Clyde; Suzuki, Akira; Saito, Mayuko; Saito, Mariko; Tamaki, Raita; Pedrera-Rico, Gay Anne Granada; Aniceto, Rapunzel; Quicho, Reynaldo Frederick Negosa; Segubre-Mercado, Edelwisa; Lupisan, Socorro; Oshitani, Hitoshi

    2015-01-01

    Human respiratory syncytial virus (HRSV) is a major cause of acute lower respiratory tract infections in infants and children worldwide. We performed molecular analysis of HRSV among infants and children with clinical diagnosis of severe pneumonia in four study sites in the Philippines, including Biliran, Leyte, Palawan, and Metro Manila from June 2012 to July 2013. Nasopharyngeal swabs were collected and screened for HRSV using real-time polymerase chain reaction (PCR). Positive samples were tested by conventional PCR and sequenced for the second hypervariable region (2nd HVR) of the G gene. Among a total of 1,505 samples, 423 samples were positive for HRSV (28.1%), of which 305 (72.1%) and 118 (27.9%) were identified as HRSV-A and HRSV-B, respectively. Two genotypes of HRSV-A, NA1 and ON1, were identified during the study period. The novel ON1 genotype with a 72-nucleotide duplication in 2nd HVR of the G gene increased rapidly and finally became the predominant genotype in 2013 with an evolutionary rate higher than the NA1 genotype. Moreover, in the ON1 genotype, we found positive selection at amino acid position 274 (p<0.05) and massive O- and N-glycosylation in the 2nd HVR of the G gene. Among HRSV-B, BA9 was the predominant genotype circulating in the Philippines. However, two sporadic cases of GB2 genotype were found, which might share a common ancestor with other Asian strains. These findings suggest that HRSV is an important cause of severe acute respiratory infection among children in the Philippines and revealed the emergence and subsequent predominance of the ON1 genotype and the sporadic detection of the GB2 genotype. Both genotypes were detected for the first time in the Philippines.

  11. Molecular Epidemiology and Evolution of Human Respiratory Syncytial Virus and Human Metapneumovirus

    PubMed Central

    Gaunt, Eleanor R.; Jansen, Rogier R.; Poovorawan, Yong; Templeton, Kate E.; Toms, Geoffrey L.; Simmonds, Peter

    2011-01-01

    Human respiratory syncytial virus (HRSV) and human metapneumovirus (HMPV) are ubiquitous respiratory pathogens of the Pneumovirinae subfamily of the Paramyxoviridae. Two major surface antigens are expressed by both viruses; the highly conserved fusion (F) protein, and the extremely diverse attachment (G) glycoprotein. Both viruses comprise two genetic groups, A and B. Circulation frequencies of the two genetic groups fluctuate for both viruses, giving rise to frequently observed switching of the predominantly circulating group. Nucleotide sequence data for the F and G gene regions of HRSV and HMPV variants from the UK, the Netherlands, Bangkok and data available from Genbank were used to identify clades of both viruses. Several contemporary circulating clades of HRSV and HMPV were identified by phylogenetic reconstructions. The molecular epidemiology and evolutionary dynamics of clades were modelled in parallel. Times of origin were determined and positively selected sites were identified. Sustained circulation of contemporary clades of both viruses for decades and their global dissemination demonstrated that switching of the predominant genetic group did not arise through the emergence of novel lineages each respiratory season, but through the fluctuating circulation frequencies of pre-existing lineages which undergo proliferative and eclipse phases. An abundance of sites were identified as positively selected within the G protein but not the F protein of both viruses. For HRSV, these were discordant with previously identified residues under selection, suggesting the virus can evade immune responses by generating diversity at multiple sites within linear epitopes. For both viruses, different sites were identified as positively selected between genetic groups. PMID:21390255

  12. Curcumin modified silver nanoparticles for highly efficient inhibition of respiratory syncytial virus infection

    NASA Astrophysics Data System (ADS)

    Yang, Xiao Xi; Li, Chun Mei; Huang, Cheng Zhi

    2016-01-01

    Interactions between nanoparticles and viruses have attracted increasing attention due to the antiviral activity of nanoparticles and the resulting possibility to be employed as biomedical interventions. In this contribution, we developed a very simple route to prepare uniform and stable silver nanoparticles (AgNPs) with antiviral properties by using curcumin, which is a member of the ginger family isolated from rhizomes of the perennial herb Curcuma longa and has a wide range of biological activities like antioxidant, antifungal, antibacterial and anti-inflammatory effects, and acts as reducing and capping agents in this synthetic route. The tissue culture infectious dose (TCID50) assay showed that the curcumin modified silver nanoparticles (cAgNPs) have a highly efficient inhibition effect against respiratory syncytial virus (RSV) infection, giving a decrease of viral titers about two orders of magnitude at the concentration of cAgNPs under which no toxicity was found to the host cells. Mechanism investigations showed that cAgNPs could prevent RSV from infecting the host cells by inactivating the virus directly, indicating that cAgNPs are a novel promising efficient virucide for RSV.Interactions between nanoparticles and viruses have attracted increasing attention due to the antiviral activity of nanoparticles and the resulting possibility to be employed as biomedical interventions. In this contribution, we developed a very simple route to prepare uniform and stable silver nanoparticles (AgNPs) with antiviral properties by using curcumin, which is a member of the ginger family isolated from rhizomes of the perennial herb Curcuma longa and has a wide range of biological activities like antioxidant, antifungal, antibacterial and anti-inflammatory effects, and acts as reducing and capping agents in this synthetic route. The tissue culture infectious dose (TCID50) assay showed that the curcumin modified silver nanoparticles (cAgNPs) have a highly efficient inhibition

  13. Modelling estimates of the burden of respiratory syncytial virus infection in children in the UK

    PubMed Central

    Taylor, Sylvia; Taylor, Robert J; Lustig, Roger L; Schuck-Paim, Cynthia; Haguinet, François; Webb, David J; Logie, John; Matias, Gonçalo; Fleming, Douglas M

    2016-01-01

    Objective The burden of respiratory syncytial virus (RSV) illness is not well characterised in primary care. We estimated the burden of disease attributable to RSV in children in the UK between 1995 and 2009. Design Time-series regression modelling. Setting A multiple linear regression model based on weekly viral surveillance (RSV and influenza, Public Health England), and controlled for non-specific seasonal drivers of disease, estimated the proportion of general practitioner (GP) episodes of care (counted as first visit in a series within 28 days; Clinical Practice Research Datalink, CPRD), hospitalisations (Hospital Episode Statistics, HES) and deaths (Office of National Statistics, ONS) attributable to RSV each season. Participants Children 0–17 years registered with a GP in CPRD, or with a respiratory disease outcome in the HES or ONS databases. Primary outcome measures RSV-attributable burden of GP episodes, hospitalisations and deaths due to respiratory disease by age. RSV-attributable burden associated with selected antibiotic prescriptions. Results RSV-attributable respiratory disease in the UK resulted in an estimated 450 158 GP episodes, 29 160 hospitalisations and 83 deaths per average season in children and adolescents, with the highest proportions in children <6 months of age (14 441/100 000 population, 4184/100 000 and 6/100 000, respectively). In an average season, there were an estimated 125 478 GP episodes for otitis media and 416 133 prescriptions for antibiotics attributable to RSV. More GP episodes, hospitalisations and deaths from respiratory disease were attributable to RSV than to influenza in children under 5 years. Conclusions The burden of RSV in children in the UK exceeds that of influenza. RSV in children and adolescents contributes substantially to GP office visits for a diverse range of illnesses, and was associated with an average 416 133 prescribed antibiotic courses per season. Effective antiviral

  14. Relative frequency, Possible Risk Factors, Viral Codetection Rates, and Seasonality of Respiratory Syncytial Virus Among Children With Lower Respiratory Tract Infection in Northeastern Brazil

    PubMed Central

    Gurgel, Ricardo Queiroz; Bezerra, Patrícia Gomes de Matos; Duarte, Maria do Carmo Menezes Bezerra; Moura, Adriana Ávila; Souza, Edna Lucia; Silva, Luciana Sobral da Silveira; Suzuki, Claudia Eiko; Peixoto, Rodrigo Buzzatti

    2016-01-01

    Abstract Few studies, each limited to a single major city, have investigated the prevalence and seasonal patterns of different viruses among children with low respiratory tract infections (LRTI) in Northeastern Brazil. The aim of this study was to determine the frequency of respiratory syncytial virus (RSV) and of 7 other viruses in children for LRTI in 4 capitals from this region, and investigate their association with several risk factors, including meteorological data. From April 2012 to March 2013, 507 children, aged up to 24 months and hospitalized with LRTI in one of the participating centers at Aracajú, Salvador, Recife, and Maceió, had a sample of nasopharyngeal aspirate collected and analyzed for the following viruses by reverse-transcription polymerase chain reaction followed by hybridization on low-density microarrays: RSV, influenza, parainfluenza, adenovirus, rhinovirus, metapneumovirus, bocavirus, and coronavirus. The result was positive in 66.5% of cases, RSV was the most common virus (40.2%). Except for rhinovirus (17%), all other virus had frequency rates lower than 6%. Viral coinfections were detected in 13.8% of samples. Possible related risk factors for RSV infection were low age upon entry, attendance of daycare, low gestational age, and low educational level of the father. The relative frequency of viral infections was associated with increasing temperature and decreasing humidity separately, but the results also suggested both associated with increased frequency of RSV. Some of these findings differ from those reported for other regions in Brazil and may be used to guide policies that address LRTI. PMID:27082548

  15. Interaction between human BAP31 and respiratory syncytial virus small hydrophobic (SH) protein.

    PubMed

    Li, Yan; Jain, Neeraj; Limpanawat, Suweeraya; To, Janet; Quistgaard, Esben M; Nordlund, Par; Thanabalu, Thirumaran; Torres, Jaume

    2015-08-01

    The small hydrophobic (SH) protein is a short channel-forming polypeptide encoded by the human respiratory syncytial virus (hRSV). Deletion of SH protein leads to the viral attenuation in mice and primates, and delayed apoptosis in infected cells. We have used a membrane-based yeast two-hybrid system (MbY2H) and a library from human lung cDNA to detect proteins that bind SH protein. This led to the identification of a membrane protein, B-cell associated protein 31 (BAP31). Transfected SH protein co-localizes with transfected BAP31 in cells, and pulls down endogenous BAP31. Titration of purified C-terminal endodomain of BAP31 against isotopically labeled SH protein in detergent micelles suggests direct interaction between the two proteins. Given the key role of BAP31 in protein trafficking and its critical involvement in pro- and anti-apoptotic pathways, this novel interaction may constitute a potential drug target.

  16. Atomic force microscopic investigation of respiratory syncytial virus infection in HEp-2 cells.

    PubMed

    Tiwari, P M; Eroglu, E; Boyoglu-Barnum, S; He, Q; Willing, G A; Vig, K; Dennis, V A; Singh, S R

    2014-01-01

    Respiratory syncytial virus (RSV) primarily causes bronchiolitis and pneumonia in infants. In spite of intense research, no safe and effective vaccine has been developed yet. For understanding its pathogenesis and development of anti-RSV drugs/therapeutics, it is indispensable to study the RSV-host interaction. Although, there are limited studies using electron microscopy to elucidate the infection process of RSV, to our knowledge, no study has reported the morphological impact of RSV infection using atomic force microscopy. We report the cytoplasmic and nuclear changes in human epidermoid cell line type 2 using atomic force microscopy. Human epidermoid cell line type 2 cells, grown on cover slips, were infected with RSV and fixed after various time periods, processed and observed for morphological changes using atomic force microscopy. RSV infected cells showed loss of membrane integrity, with degeneration in the cellular content and cytoskeleton. Nuclear membrane was disintegrated and nuclear volume was decreased. The chromatin of the RSV infected cells was condensed, progressing towards degeneration via pyknosis and apoptosis. Membrane protrusions of ~150-200 nm diameter were observed on RSV infected cells after 6 h, suggestive of prospective RSV budding sites. To our knowledge, this is the first study of RSV infection process using atomic force microscopy. Such morphological studies could help explore viral infection process aiding the development of anti-RSV therapies.

  17. Intranasal nanoemulsion-based inactivated respiratory syncytial virus vaccines protect against viral challenge in cotton rats

    PubMed Central

    O'Konek, Jessica J; Makidon, Paul E; Landers, Jeffrey J; Cao, Zhengyi; Malinczak, Carrie-Anne; Pannu, Jessie; Sun, Jennifer; Bitko, Vira; Ciotti, Susan; Hamouda, Tarek; Wojcinski, Zbigniew W; Lukacs, Nicholas W; Fattom, Ali; Baker, James R

    2015-01-01

    Respiratory Syncytial Virus is a leading cause of bronchiolitis and pneumonia in infants, the elderly and individuals with compromised immune systems. Despite decades of research, there is currently no available vaccine for RSV. Our group has previously demonstrated that intranasal immunization of mice with RSV inactivated by and adjuvanted with W805EC nanoemulsion elicits robust humoral and cellular immune responses, resulting in protection against RSV infection. This protection was achieved without the induction of airway hyper-reactivity or a Th2-skewed immune response. The cotton rat Sigmodon hispidus has been used for years as an excellent small animal model of RSV disease. Thus, we extended these rodent studies to the more permissive cotton rat model. Intranasal immunization of the nanoemulsion-adjuvanted RSV vaccines induced high antibody titers and a robust Th1-skewed cellular response. Importantly, vaccination provided sterilizing cross-protective immunity against a heterologous RSV challenge and did not induce marked or severe histological effects or eosinophilia in the lung after viral challenge. Overall, these data demonstrate that nanoemulsion-formulated whole RSV vaccines are both safe and effective for immunization in multiple animal models. PMID:26307915

  18. Titanium dioxide nanoparticles exacerbate pneumonia in respiratory syncytial virus (RSV)-infected mice.

    PubMed

    Hashiguchi, Seiko; Yoshida, Hiroki; Akashi, Toshi; Komemoto, Keiji; Ueda, Tomoyuki; Ikarashi, Yoshiaki; Miyauchi, Aki; Konno, Katsuhiko; Yamanaka, Sayoko; Hirose, Akihiko; Kurokawa, Masahiko; Watanabe, Wataru

    2015-03-01

    To reveal the effects of TiO2 nanoparticles, used in cosmetics and building materials, on the immune response, a respiratory syncytial virus (RSV) infection mouse model was used. BALB/c mice were exposed once intranasally to TiO2 at 0.5mg/kg and infected intranasally with RSV five days later. The levels of IFN-γ and chemokine CCL5, representative markers of pneumonia, in the bronchoalveolar lavage fluids of RSV-infected mice had increased significantly in TiO2-exposed mice compared with the control on day 5 post-infection, but not in uninfected mice. While pulmonary viral titers were not affected by TiO2 exposure, an increase in the infiltration of lymphocytes into the alveolar septa in lung tissues was observed. Immunohistochemical analysis revealed aggregation of TiO2 nanoparticles near inflammatory cells in the severely affected region. Thus, a single exposure to TiO2 nanoparticles affected the immune system and exacerbated pneumonia in RSV-infected mice.

  19. An attraction-repulsion point process model for respiratory syncytial virus infections.

    PubMed

    Goldstein, Joshua; Haran, Murali; Simeonov, Ivan; Fricks, John; Chiaromonte, Francesca

    2015-06-01

    How is the progression of a virus influenced by properties intrinsic to individual cells? We address this question by studying the susceptibility of cells infected with two strains of the human respiratory syncytial virus (RSV-A and RSV-B) in an in vitro experiment. Spatial patterns of infected cells give us insight into how local conditions influence susceptibility to the virus. We observe a complicated attraction and repulsion behavior, a tendency for infected cells to lump together or remain apart. We develop a new spatial point process model to describe this behavior. Inference on spatial point processes is difficult because the likelihood functions of these models contain intractable normalizing constants; we adapt an MCMC algorithm called double Metropolis-Hastings to overcome this computational challenge. Our methods are computationally efficient even for large point patterns consisting of over 10,000 points. We illustrate the application of our model and inferential approach to simulated data examples and fit our model to various RSV experiments. Because our model parameters are easy to interpret, we are able to draw meaningful scientific conclusions from the fitted models. PMID:25660222

  20. Phosphorylation of human respiratory syncytial virus P protein at serine 54 regulates viral uncoating

    SciTech Connect

    Asenjo, Ana; Gonzalez-Armas, Juan C.; Villanueva, Nieves

    2008-10-10

    The human respiratory syncytial virus (HRSV) structural P protein, phosphorylated at serine (S) and threonine (T) residues, is a co-factor of viral RNA polymerase. The phosphorylation of S54 is controlled by the coordinated action of two cellular enzymes: a lithium-sensitive kinase, probably glycogen synthetase kinase (GSK-3) {beta} and protein phosphatase 2A (PP2A). Inhibition of lithium-sensitive kinase, soon after infection, blocks the viral growth cycle by inhibiting synthesis and/or accumulation of viral RNAs, proteins and extracellular particles. P protein phosphorylation at S54 is required to liberate viral ribonucleoproteins (RNPs) from M protein, during the uncoating process. Kinase inhibition, late in infection, produces a decrease in genomic RNA and infectious viral particles. LiCl, intranasally applied to mice infected with HRSV A2 strain, reduces the number of mice with virus in their lungs and the virus titre. Administration of LiCl to humans via aerosol should prevent HRSV infection, without secondary effects.

  1. Ginseng protects against respiratory syncytial virus by modulating multiple immune cells and inhibiting viral replication.

    PubMed

    Lee, Jong Seok; Lee, Yu-Na; Lee, Young-Tae; Hwang, Hye Suk; Kim, Ki-Hye; Ko, Eun-Ju; Kim, Min-Chul; Kang, Sang-Moo

    2015-01-01

    Ginseng has been used in humans for thousands of years but its effects on viral infection have not been well understood. We investigated the effects of red ginseng extract (RGE) on respiratory syncytial virus (RSV) infection using in vitro cell culture and in vivo mouse models. RGE partially protected human epithelial (HEp2) cells from RSV-induced cell death and viral replication. In addition, RGE significantly inhibited the production of RSV-induced pro-inflammatory cytokine (TNF-α) in murine dendritic and macrophage-like cells. More importantly, RGE intranasal pre-treatment prevented loss of mouse body weight after RSV infection. RGE treatment improved lung viral clearance and enhanced the production of interferon (IFN-γ) in bronchoalveolar lavage cells upon RSV infection of mice. Analysis of cellular phenotypes in bronchoalveolar lavage fluids showed that RGE treatment increased the populations of CD8+ T cells and CD11c+ dendritic cells upon RSV infection of mice. Taken together, these results provide evidence that ginseng has protective effects against RSV infection through multiple mechanisms, which include improving cell survival, partial inhibition of viral replication and modulation of cytokine production and types of immune cells migrating into the lung. PMID:25658239

  2. Preparation of respiratory syncytial virus subgroup A and B antigens for enzyme immunoassay antibody detection.

    PubMed

    Mlinarić-Galinović, G; Chonmaintree, T; Abraham, R; Garofalo, R; Patel, J A; Ogra, P L

    1994-01-01

    A simplified method was described for purification of respiratory syncytial virus (RSV) subgroup A and B aimed to be used as antigens in enzyme immunoassay (EIA). The titer of each RSV subgroup and the amount of protein was determined from the visible band in 45% sucrose gradient. The quality of prepared RSV subgroup antigens for EIA was described in terms of the achievable final titer, the amount of protein, and EIA criss-cross titration. The RSV subgroup A and B antigens, diluted as 1:100 (low opalescent band in 45% sucrose layer) or 1:800 (high opalescent band in 45% sucrose layer) produced a positive reaction in EIA criss-cross titration with IgG antibodies from the patient's serum (convalescent phase) diluted as 1:25,600 (for RSV A) and 1:6,400 (for RSV B). This method offers shorter and more simplified steps of viral antigen purification, and provides acceptable quantity and quality of viral antigens appropriate for use in EIA.

  3. Systemic immunoprophylaxis of nasal respiratory syncytial virus infection in cotton rats.

    PubMed

    Sami, I R; Piazza, F M; Johnson, S A; Darnell, M E; Ottolini, M G; Hemming, V G; Prince, G A

    1995-02-01

    The cotton rat model was used to test whether systemically administered immunoglobulin could protect nasal tissues against low challenge doses of respiratory syncytial virus (RSV). Animals were pretreated by intraperitoneal injection of human immunoglobulin with moderate (1:2226) or high (1:15,000) neutralizing antibody titers to RSV (day 0), challenged intranasally with RSV Long at doses ranging from 10(1) to 10(5) pfu (day 1), and sacrificed for virus titration (day 5). Pretreatment with moderate-titer immunoglobulin effected complete or near complete nasal protection against low to moderate (10(1)-10(3) pfu) RSV challenge doses and a significant reduction in nasal RSV titers at high (10(4)-10(5) pfu) challenge doses. Pretreatment with high-titer immunoglobulin effected near complete nasal protection at an RSV challenge dose of 10(3) pfu and highly significant and significant reductions in nasal RSV titers at challenge doses of 10(4) and 10(5) pfu, respectively. Immunoprophylaxis effected complete or near complete pulmonary protection at all RSV challenge doses.

  4. A cotton rat model of effectors of immunity to respiratory syncytial virus other than serum antibody.

    PubMed

    Piazza, F M; Schmidt, H J; Johnson, S A; Dotson, D L; Darnell, M E; Ottolini, M G; Porter, D D; Prince, G A

    1995-06-01

    A model for studying effectors of immunity to respiratory syncytial virus (RSV) was developed. Paris of inbred cotton rats (Sigmodon hispidus) were joined surgically using the technique of parabiosis. One week later, one animal of each pair was primed intranasally with a small volume of RSV suspension. Fourteen days after priming, both animals of each pair were bled for determination of serum neutralizing antibody titers, and challenged intranasally with a standard dose of RSV suspension. Single, unprimed cotton rats were challenged concomitantly and served as controls. Four days after challenge, all animals were sacrificed for virus titration of nasal tissues and lungs. Parabiosed cotton rats were surgically separated at varying intervals between priming and challenge (days 7, 9, 12, or 14 after priming) or were kept joined until sacrificed (day 18). Significant transfer of nasal and pulmonary immunity from primed to unprimed parabionts began 9 days after priming, gradually increasing through 18 days. Resistance to RSV challenge in spite of low levels of serum neutralizing antibody suggests that non-antibody immunologic mediators were responsible for the transferred immunity. Evidence is presented for three broad categories of RSV immunologic effectors: systemic, local with a transient systemic phase, and local without a systemic phase. These categories are now amenable to further study using the described model.

  5. Validity of laboratory-based surveillance for detection of respiratory syncytial virus seasons.

    PubMed

    Hampp, Christian; Asal, Nabih; Lipowski, Earlene; Kauf, Teresa; Schneider, Eileen; Kubilis, Paul; Winterstein, Almut

    2013-04-15

    In this study, we validated the Centers for Disease Control and Prevention's use of a 10% threshold of median proportion of positive laboratory tests (median proportion positive (MPP)) to identify respiratory syncytial virus (RSV) seasons against a standard based on hospitalization claims. Medicaid fee-for-service recipients under 2 years of age from California, Florida, Illinois, and Texas (1999-2004), continuously eligible since birth, were categorized for each week as high-risk or low-risk with regard to RSV-related hospitalization based on medical and pharmacy claims data and birth certificates. Weeks were categorized as on-season if the RSV hospitalization incidence rate in high-risk children exceeded the seasonal peak of the incidence rate in low-risk children. Receiver operating characteristic (ROC) curves were used to measure the ability of MPP to discriminate between on-season and off-season weeks as determined from hospitalization data. Areas under the ROC curve ranged from 0.88 (95% confidence interval: 0.83, 0.92) in Illinois to 0.96 (95% confidence interval: 0.94, 0.98) in California. Requiring at least 5 positive tests in addition to the 10% MPP threshold optimized accuracy, as indicated by minimized root mean square errors. The 10% MPP with the added requirement of at least 5 positive tests is a valid method for identifying clinically significant RSV seasons across geographically diverse states.

  6. Structure of Respiratory Syncytial Virus Fusion Glycoprotein in the Postfusion Conformation Reveals Preservation of Neutralizing Epitopes

    SciTech Connect

    McLellan, Jason S.; Yang, Yongping; Graham, Barney S.; Kwong, Peter D.

    2011-09-16

    Respiratory syncytial virus (RSV) invades host cells via a type I fusion (F) glycoprotein that undergoes dramatic structural rearrangements during the fusion process. Neutralizing monoclonal antibodies, such as 101F, palivizumab, and motavizumab, target two major antigenic sites on the RSV F glycoprotein. The structures of these sites as peptide complexes with motavizumab and 101F have been previously determined, but a structure for the trimeric RSV F glycoprotein ectodomain has remained elusive. To address this issue, we undertook structural and biophysical studies on stable ectodomain constructs. Here, we present the 2.8-{angstrom} crystal structure of the trimeric RSV F ectodomain in its postfusion conformation. The structure revealed that the 101F and motavizumab epitopes are present in the postfusion state and that their conformations are similar to those observed in the antibody-bound peptide structures. Both antibodies bound the postfusion F glycoprotein with high affinity in surface plasmon resonance experiments. Modeling of the antibodies bound to the F glycoprotein predicts that the 101F epitope is larger than the linear peptide and restricted to a single protomer in the trimer, whereas motavizumab likely contacts residues on two protomers, indicating a quaternary epitope. Mechanistically, these results suggest that 101F and motavizumab can bind to multiple conformations of the fusion glycoprotein and can neutralize late in the entry process. The structural preservation of neutralizing epitopes in the postfusion state suggests that this conformation can elicit neutralizing antibodies and serve as a useful vaccine antigen.

  7. A recombinant varicella vaccine harboring a respiratory syncytial virus gene induces humoral immunity.

    PubMed

    Murakami, Kouki; Matsuura, Masaaki; Ota, Megumi; Gomi, Yasuyuki; Yamanishi, Koichi; Mori, Yasuko

    2015-11-01

    The varicella-zoster virus (VZV) Oka vaccine strain (vOka) is highly efficient and causes few adverse events; therefore, it is used worldwide. We previously constructed recombinant vOka (rvOka) harboring the mumps virus gene. Immunizing guinea pigs with rvOka induced the production of neutralizing antibodies against the mumps virus and VZV. Here, we constructed recombinant vOka viruses containing either the respiratory syncytial virus (RSV) subgroup A fusion glycoprotein (RSV A-F) gene or RSV subgroup B fusion glycoprotein (RSV B-F) gene (rvOka-RSV A-F or rvOka-RSV B-F). Indirect immunofluorescence and Western blot analyses confirmed the expression of each recombinant RSV protein in virus-infected cells. Immunizing guinea pigs with rvOka-RSV A-F or rvOka-RSV B-F led to the induction of antibodies against RSV proteins. These results suggest that the current varicella vaccine genome can be used to generate custom-made vaccine vectors to develop the next generation of live vaccines.

  8. Experimental animal model for analyzing immunobiological responses following vaccination with formalin-inactivated respiratory syncytial virus.

    PubMed

    Sawada, Akihito; Nakayama, Tetsuo

    2016-04-01

    Formalin-inactivated respiratory syncytial virus (FI-RSV) vaccine was developed in the 1960s. However, this vaccine does not prevent infection in RSV-naïve recipients and has the paradoxical effect of increasing the severity of RSV illness following natural infection, which has been a major obstacle to developing RSV vaccines. Several experimental animal models for determining the cause of the severe symptoms in FI-RSV recipients have been developed. In the present study, cotton rats immunized with FI-RSV were challenged with RSV and histopathological findings and recovery of infectious virus were studied. Copy numbers of mRNA of Th1 and Th2 cytokines were measured in lung tissues to gain better understanding of their immune responses. Infiltration of inflammatory cells and prominent interstitial pneumonitis were observed in the FI-RSV group, as was induction of mRNA of Th2 cytokines such as IL-4, IL-10, IL-13 and RANTES. Rats immunized with recombinant measles virus expressing the RSV F protein (MVAIK/RSV/F) and those treated with anti-RSV mAb (palivizumab) showed very mild interstitial pneumonitis. Amounts of mRNA of IL-1β, IFN-γ and IL-4 were higher in the MVAIK/RSV/F group. Administration of palivizumab before RSV challenge decreased the severity of interstitial pneumonitis in the FI-RSV group. FI-RSV induced skewed Th2 responses, resulting in severe inflammatory responses. PMID:26865035

  9. Intranasal nanoemulsion-based inactivated respiratory syncytial virus vaccines protect against viral challenge in cotton rats.

    PubMed

    O'Konek, Jessica J; Makidon, Paul E; Landers, Jeffrey J; Cao, Zhengyi; Malinczak, Carrie-Anne; Pannu, Jessie; Sun, Jennifer; Bitko, Vira; Ciotti, Susan; Hamouda, Tarek; Wojcinski, Zbigniew W; Lukacs, Nicholas W; Fattom, Ali; Baker, James R

    2015-01-01

    Respiratory Syncytial Virus is a leading cause of bronchiolitis and pneumonia in infants, the elderly and individuals with compromised immune systems. Despite decades of research, there is currently no available vaccine for RSV. Our group has previously demonstrated that intranasal immunization of mice with RSV inactivated by and adjuvanted with W805EC nanoemulsion elicits robust humoral and cellular immune responses, resulting in protection against RSV infection. This protection was achieved without the induction of airway hyper-reactivity or a Th2-skewed immune response. The cotton rat Sigmodon hispidus has been used for years as an excellent small animal model of RSV disease. Thus, we extended these rodent studies to the more permissive cotton rat model. Intranasal immunization of the nanoemulsion-adjuvanted RSV vaccines induced high antibody titers and a robust Th1-skewed cellular response. Importantly, vaccination provided sterilizing cross-protective immunity against a heterologous RSV challenge and did not induce marked or severe histological effects or eosinophilia in the lung after viral challenge. Overall, these data demonstrate that nanoemulsion-formulated whole RSV vaccines are both safe and effective for immunization in multiple animal models. PMID:26307915

  10. Development of Electrochemiluminescent Serology Assays to Measure the Humoral Response to Antigens of Respiratory Syncytial Virus

    PubMed Central

    Maifeld, Sarah V.; Ro, Bodrey; Mok, Hoyin; Chu, Marla; Yu, Li; Yamagata, Ryan; Leonardson, Tansy; Chio, Vera; Parhy, Bandita; Park, Samuel; Carlson, Marcia; Machhi, Shushil; Ulbrandt, Nancy; Falsey, Ann R.; Walsh, Edward E.; Wang, C. Kathy; Esser, Mark T.; Zuo, Fengrong

    2016-01-01

    Sensitive and precise serology assays are needed to measure the humoral response to antigens of respiratory syncytial virus (RSV) following natural infection or vaccination. We developed and evaluated a collection of electrochemiluminescent (ECL) serology assays using four RSV antigens (F, N, Ga and Gb). To assess the merits of ECL technology, the four ECL serology assays were evaluated using a well-characterized “gold standard” panel of acute and convalescent serum samples from fifty-nine RSV-positive and thirty RSV-negative elderly subjects (≥65 years old). The combined results from the four ECL assays demonstrated good concordance to the “gold standard” diagnosis, reaching 95% diagnostic sensitivity and 100% diagnostic specificity. Additionally, a combination of ECL assays provided higher diagnostic sensitivity than a commercially available diagnostic ELISA or cell-based microneutralization assay. In summary, these data demonstrate the advantages of using ECL-based serology assays and highlight their use as a sensitive diagnostic approach to detect recent RSV infection in an elderly population. PMID:27070145

  11. Development of Electrochemiluminescent Serology Assays to Measure the Humoral Response to Antigens of Respiratory Syncytial Virus.

    PubMed

    Maifeld, Sarah V; Ro, Bodrey; Mok, Hoyin; Chu, Marla; Yu, Li; Yamagata, Ryan; Leonardson, Tansy; Chio, Vera; Parhy, Bandita; Park, Samuel; Carlson, Marcia; Machhi, Shushil; Ulbrandt, Nancy; Falsey, Ann R; Walsh, Edward E; Wang, C Kathy; Esser, Mark T; Zuo, Fengrong

    2016-01-01

    Sensitive and precise serology assays are needed to measure the humoral response to antigens of respiratory syncytial virus (RSV) following natural infection or vaccination. We developed and evaluated a collection of electrochemiluminescent (ECL) serology assays using four RSV antigens (F, N, Ga and Gb). To assess the merits of ECL technology, the four ECL serology assays were evaluated using a well-characterized "gold standard" panel of acute and convalescent serum samples from fifty-nine RSV-positive and thirty RSV-negative elderly subjects (≥65 years old). The combined results from the four ECL assays demonstrated good concordance to the "gold standard" diagnosis, reaching 95% diagnostic sensitivity and 100% diagnostic specificity. Additionally, a combination of ECL assays provided higher diagnostic sensitivity than a commercially available diagnostic ELISA or cell-based microneutralization assay. In summary, these data demonstrate the advantages of using ECL-based serology assays and highlight their use as a sensitive diagnostic approach to detect recent RSV infection in an elderly population.

  12. Respiratory syncytial virus increases lung cellular bioenergetics in neonatal C57BL/6 mice

    SciTech Connect

    Alsuwaidi, Ahmed R.; Albawardi, Alia; Almarzooqi, Saeeda; Benedict, Sheela; Othman, Aws R.; Hartwig, Stacey M.; Varga, Steven M.; Souid, Abdul-Kader

    2014-04-15

    We have previously reported that lung cellular bioenergetics (cellular respiration and ATP) increased in 4–10 week-old BALB/c mice infected with respiratory syncytial virus (RSV). This study examined the kinetics and changes in cellular bioenergetics in ≤2-week-old C57BL/6 mice following RSV infection. Mice (5–14 days old) were inoculated intranasally with RSV and the lungs were examined on days 1–10 post-infection. Histopathology and electron microscopy revealed preserved pneumocyte architectures and organelles. Increased lung cellular bioenergetics was noted from days 1–10 post-infection. Cellular GSH remained unchanged. These results indicate that the increased lung cellular respiration (measured by mitochondrial O{sub 2} consumption) and ATP following RSV infection is independent of either age or genetic background of the host. - Highlights: • RSV infection increases lung cellular respiration and ATP in neonatal C57BL/6 mice. • Increased lung cellular bioenergetics is a biomarker of RSV infection. • Lung cellular glutathione remains unchanged in RSV infection.

  13. Bronchiectasis exacerbations: The role of atypical bacteria and respiratory syncytial virus

    PubMed Central

    Metaxas, Eugenios I; Balis, Evangelos; Papaparaskevas, Joseph; Spanakis, Nicholas E; Tatsis, Georgios; Tsakris, Athanasios

    2015-01-01

    BACKGROUND: Aside from the known role of common bacteria, there is a paucity of data regarding the possible role of atypical bacteria and viruses in exacerbations of non-cystic fibrosis bronchiectasis. OBJECTIVE: To explore the possible role of atypical bacteria (namely, Mycoplasma pneumoniae and Chlamydophila pneumoniae) and respiratory syncytial virus (RSV) as causative agents of bronchiectasis exacerbations. METHODS: A cohort of 33 patients was studied over a two-year period (one year follow-up for each patient). Polymerase chain reaction for the detection of M pneumoniae, C pneumoniae and RSV in bronchoalveolar lavage samples were performed during all visits. Antibody titres (immunoglobulin [Ig]M and IgG) against the aforementioned pathogens were also measured. In addition, cultures for common bacteria and mycobacteria were performed from the bronchoalveolar lavage samples. RESULTS: Fifteen patients experienced a total of 19 exacerbations during the study period. Although RSV was detected by polymerase chain reaction during stable visits in four patients, it was never detected during an exacerbation. M pneumoniae and C pneumoniae were never detected at stable visits or during exacerbations. IgM antibody titres for these three pathogens were negative in all patient visits. CONCLUSIONS: Atypical pathogens and RSV did not appear to be causative agents of bronchiectasis exacerbations. PMID:25874735

  14. Influence of Respiratory Syncytial Virus Strain Differences on Pathogenesis and Immunity

    PubMed Central

    Melero, José A.; Moore, Martin L.

    2016-01-01

    Molecular epidemiology studies have provided convincing evidence of antigenic and sequence variability among respiratory syncytial virus (RSV) isolates. Circulating viruses have been classified into two antigenic groups (A and B) that correlate with well-delineated genetic groups. Most sequence and antigenic differences (both inter- and intra-groups) accumulate in two hypervariable segments of the G protein gene. Sequences of the G gene have been used for phylogenetic analyses. These studies have shown a worldwide distribution of RSV strains with both local and global replacement of dominant viruses with time. Although data are still limited, there is evidence that strain variation may contribute to differences in pathogenicity. In addition, there is some but limited evidence that RSV variation may be, at least partially, immune (antibody) driven. However, there is the paradox in RSV that, in contrast to other viruses (e.g., influenza viruses) the epitopes recognized by the most effective RSV-neutralizing antibodies are highly conserved. In contrast, antibodies that recognize strain-specific epitopes are poorly neutralizing. It is likely that this apparent contradiction is due to the lack of a comprehensive knowledge of the duration and specificities of the human antibody response against RSV antigens. Since there are some data supporting a group- (or clade-) specific antibody response after a primary infection in humans, it may be wise to consider the incorporation of strains representative of groups A and B (or their antigens) in future RSV vaccine development. PMID:24362684

  15. Nosocomial respiratory syncytial virus infections: the "Cold War" has not ended.

    PubMed

    Hall, C B

    2000-08-01

    Respiratory syncytial virus (RSV) is a major nosocomial hazard on pediatric wards during its annual outbreaks. It produces significant morbidity in young children and is most severe in those with underlying conditions, especially cardiopulmonary and immunosuppressive diseases. In older patients, RSV may exacerbate an underlying condition or pulmonary and cardiac manifestations. On transplant units, of RSV may be occult and is associated with high mortality rates. The manifestations of nosocomial RSV infections may be atypical, especially in neonates and immunosuppressed patients, resulting in delayed or missed diagnosis and adding appreciably to the costs of hospitalization. RSV is primarily spread by close contact with infectious secretions, either by large-particle aerosols or by fomites and subsequent self-inoculation, and medical staff are often instrumental in its transmission. Thus, integral to any infection control program is the education of personnel about the modes of transmission, the manifestations, and the importance of RSV nosocomial infections. Hand washing is probably the most important infection control procedure. The choice of barrier controls should be decided by individual institutions depending on the patients, the type of ward, and the benefit relative to cost.

  16. Morbidity and mortality among patients with respiratory syncytial virus infection: a 2-year retrospective review.

    PubMed

    Anderson, Neil W; Binnicker, Matthew J; Harris, Dana M; Chirila, Razvan M; Brumble, Lisa; Mandrekar, Jay; Hata, D Jane

    2016-07-01

    Previous studies have demonstrated high morbidity and mortality for adult patients with respiratory syncytial virus (RSV) infection. We performed a retrospective, multicenter, two-year chart review of all patients (n = 334) testing positive for RSV by the ProFlu + (®) Influenza A/B and RSV assay (Hologic, Bedford, MA). We analyzed indicators of morbidity and mortality in children <6 years old, immunocompetent and immunosuppressed adults, and transplant patients. Significant morbidity and mortality was observed among hematopoietic stem cell transplant patients (7.3%, 60-day mortality), solid organ transplant patients (13.3%, 60-day mortality), and COPD patients (12.8%, 60-day mortality). Of the patients positive for RSV, 144 (43.1%) of 334 received antibacterials or antifungals following diagnosis. Of these patients, a bacterial or fungal pathogen was not recovered from 60% of cases. Despite advances in RSV treatment, certain populations appear to be inadequately treated, while others appear to be inappropriately treated with unnecessary antimicrobials. PMID:27179369

  17. Respiratory syncytial virus M2-1 protein induces the activation of nuclear factor kappa B

    SciTech Connect

    Reimers, Kerstin . E-mail: reimers.kerstin@mh-hannover.de; Buchholz, Katja; Werchau, Hermann

    2005-01-20

    Respiratory syncytial virus (RSV) induces the production of a number of cytokines and chemokines by activation of nuclear factor kappa B (NF-{kappa}B). The activation of NF-{kappa}B has been shown to depend on viral replication in the infected cells. In this study, we demonstrate that expression of RSV M2-1 protein, a transcriptional processivity and anti-termination factor, is sufficient to activate NF-{kappa}B in A549 cells. Electromobility shift assays show increased NF-{kappa}B complexes in the nuclei of M2-1-expressing cells. M2-1 protein is found in nuclei of M2-1-expressing cells and in RSV-infected cells. Co-immunoprecipitations of nuclear extracts of M2-1-expressing cells and of RSV-infected cells revealed an association of M2-1 with Rel A protein. Furthermore, the activation of NF-{kappa}B depends on the C-terminus of the RSV M2-1 protein, as shown by NF-{kappa}B-induced gene expression of a reporter gene construct.

  18. Development of Acquired Immunity following Repeated Respiratory Syncytial Virus Infections in Cotton Rats.

    PubMed

    Yamaji, Yoshiaki; Yasui, Yosuke; Nakayama, Tetsuo

    2016-01-01

    Respiratory syncytial virus (RSV) infections occur every year worldwide. Most infants are infected with RSV by one year of age and are reinfected because immune responses after the first infection are too weak to protect against subsequent infections. In the present study, immune responses against RSV were investigated in order to obtain a better understanding of repetitive RSV infections in cotton rats. No detectable neutralizing antibody (NT) was developed after the first infection, and the second infection was not prevented. The results of histological examinations revealed severe inflammation, viral antigens were detected around bronchial epithelial cells, and infectious viruses were recovered from lung homogenates. Following the second infection neutralizing antibodies were significantly elevated, and CD8+ cells were activated in response to RSV-F253-265. No viral antigens was detected thereafter in lung tissues and infectious viruses were not recovered. Similar results were obtained in the present study using the subgroups A and B. These results support the induction of humoral and cellular immune responses following repetitive infections with RSV; however, these responses were insufficient to eliminate viruses in the first and second infections. PMID:27224021

  19. Molecular evolution of the fusion protein gene in human respiratory syncytial virus subgroup A.

    PubMed

    Kimura, Hirokazu; Nagasawa, Koo; Tsukagoshi, Hiroyuki; Matsushima, Yuki; Fujita, Kiyotaka; Yoshida, Lay Myint; Tanaka, Ryota; Ishii, Haruyuki; Shimojo, Naoki; Kuroda, Makoto; Ryo, Akihide

    2016-09-01

    We studied the molecular evolution of the fusion protein (F) gene in the human respiratory syncytial virus subgroup A (HRSV-A). We performed time-scaled phylogenetic analyses using the Bayesian Markov chain Monte Carlo (MCMC) method. We also conducted genetic distance (p-distance), positive/negative selection, and Bayesian skyline plot analyses. Furthermore, we mapped the amino acid substitutions of the protein. The MCMC-constructed tree indicated that the HRSV F gene diverged from the bovine RSV (BRSV) gene approximately 550years ago and had a relatively low substitution rate (7.59×10(-4) substitutions/site/year). Moreover, a common ancestor of HRSV-A and -B diverged approximately 280years ago, which has since formed four distinct clusters. The present HRSV-A strains were assigned six genotypes based on F gene sequences and attachment glycoprotein gene sequences. The present strains exhibited high F gene sequence similarity values and low genetic divergence. No positive selection sites were identified; however, 50 negative selection sites were identified. F protein amino acid substitutions at 17 sites were distributed in the F protein. The effective population size of the gene has remained relatively constant, but the population size of the prevalent genotype (GA2) has increased in the last 10years. These results suggest that the HRSV-AF gene has evolved independently and formed some genotypes. PMID:27291709

  20. Human CD8(+) T Cells Target Multiple Epitopes in Respiratory Syncytial Virus Polymerase.

    PubMed

    Burbulla, Daniel; Günther, Patrick S; Peper, Janet K; Jahn, Gerhard; Dennehy, Kevin M

    2016-06-01

    Respiratory syncytial virus (RSV) infection is a serious health problem in young children, immunocompromised patients, and the elderly. The development of novel prevention strategies, such as a vaccine to RSV, is a high priority. One strategy is to design a peptide-based vaccine that activates appropriate CD8(+) T-cell responses. However, this approach is limited by the low number of RSV peptide epitopes defined to date that activate CD8(+) T cells. We aimed to identify peptide epitopes that are presented by common human leukocyte antigen types (HLA-A*01, -A*02, and -B*07). We identify one novel HLA-A*02-restricted and two novel HLA-A*01-restricted peptide epitopes from RSV polymerase. Peptide-HLA multimer staining of specific T cells from healthy donor peripheral blood mononuclear cell, the memory phenotype of such peptide-specific T cells ex vivo, and functional IFNγ responses in short-term stimulation assays suggest that these peptides are recognized during RSV infection. Such peptides are candidates for inclusion into a peptide-based RSV vaccine designed to stimulate defined CD8(+) T-cell responses.

  1. Transcription elongation factor of respiratory syncytial virus, a nonsegmented negative-strand RNA virus.

    PubMed Central

    Collins, P L; Hill, M G; Cristina, J; Grosfeld, H

    1996-01-01

    RNA synthesis by the paramyxovirus respiratory syncytial virus, a ubiquitous human pathogen, was found to be more complex than previously appreciated for the nonsegmented negative-strand RNA viruses. Intracellular RNA replication of a plasmid-encoded "minigenome" analog of viral genomic RNA was directed by coexpression of the N, P, and L proteins. But, under these conditions, the greater part of mRNA synthesis terminated prematurely. This difference in processivity between the replicase and the transcriptase was unanticipated because the two enzymes ostensively shared the same protein subunits and template. Coexpression of the M2 gene at a low level of input plasmid resulted in the efficient production of full-length mRNA and, in the case of a dicistronic minigenome, sequential transcription. At a higher level, coexpression of the M2 gene inhibited transcription and RNA replication. The M2 mRNA contains two overlapping translational open reading frames (ORFs), which were segregated for further analysis. Expression of the upstream ORF1, which encoded the previously described 22-kDa M2 protein, was associated with transcription elongation. A model involving this protein in the balance between transcription and replication is proposed. ORF2, which lacks an assigned protein, was associated with inhibition of RNA synthesis. We propose that this activity renders nucleocapsids synthetically quiescent prior to incorporation into virions. Images Fig. 2 Fig. 3 Fig. 5 Fig. 6 Fig. 7 Fig. 8 PMID:8552680

  2. Induction of long-term immunity against respiratory syncytial virus glycoprotein by an osmotic polymeric nanocarrier.

    PubMed

    Firdous, Jannatul; Islam, Mohammad Ariful; Park, Sung-Moo; Cheon, In-Su; Shim, Byoung-Shik; Yoon, Hyo-Shin; Song, Manki; Chang, Jun; Choi, Yun-Jaie; Park, Yeong-Min; Boraschi, Diana; Han, Seung-Hyun; Cho, Chong-Su; Yun, Cheol-Heui

    2014-11-01

    Respiratory syncytial virus (RSV) is one of the most common causes of viral deaths in infants worldwide, yet no effective vaccines are available. Here, we report an osmotically active polysaccharide-based polysorbitol transporter (PST) prepared from sorbitol diacrylate and low-molecular-weight polyethylenimine (PEI) showing a potent, yet safe, adjuvant activity and acting as an effective delivery tool for RSV glycoprotein (RGp) antigen. PST showed no toxicity in vitro or in vivo, unlike PEI and the well-known experimental mucosal adjuvant cholera toxin (CT). PST formed nano-sized complexes with RGp by simple mixing, without affecting antigenic stability. The complexes exhibited negative surface charges that made them highly efficient in the selective activation of phagocytic cells and enhancement of phagocytic uptake. This resulted in an improved cytokine production and in the significant augmentation of RGp-specific antibody production, which persisted for over 200 days. Interestingly, PST/RGp enhanced phagocytic uptake owing to the osmotic property of PST and its negative zeta potential, suggesting that PST could selectively stimulate phagocytic cells, thereby facilitating a long-lived antigen-specific immune response, which was presumably further enhanced by the polysaccharide properties of PST. PMID:25110285

  3. Interaction between human BAP31 and respiratory syncytial virus small hydrophobic (SH) protein

    SciTech Connect

    Li, Yan; Jain, Neeraj; Limpanawat, Suweeraya; To, Janet; Quistgaard, Esben M.; Nordlund, Par; Thanabalu, Thirumaran; Torres, Jaume

    2015-08-15

    The small hydrophobic (SH) protein is a short channel-forming polypeptide encoded by the human respiratory syncytial virus (hRSV). Deletion of SH protein leads to the viral attenuation in mice and primates, and delayed apoptosis in infected cells. We have used a membrane-based yeast two-hybrid system (MbY2H) and a library from human lung cDNA to detect proteins that bind SH protein. This led to the identification of a membrane protein, B-cell associated protein 31 (BAP31). Transfected SH protein co-localizes with transfected BAP31 in cells, and pulls down endogenous BAP31. Titration of purified C-terminal endodomain of BAP31 against isotopically labeled SH protein in detergent micelles suggests direct interaction between the two proteins. Given the key role of BAP31 in protein trafficking and its critical involvement in pro- and anti-apoptotic pathways, this novel interaction may constitute a potential drug target. - Highlights: • A yeast two-hybrid system (MbY2H) detected BAP31 as a binder of RSV SH protein. • Transfected SH and BAP31 co-localize in lung epithelial cells. • Endogenous BAP31 is pulled down by RSV SH protein. • BAP31 endodomain interacts with the N-terminal α-helix of SH protein in micelles. • This interaction is proposed to be a potential drug target.

  4. The respiratory syncytial virus nucleoprotein-RNA complex forms a left-handed helical nucleocapsid.

    PubMed

    Bakker, Saskia E; Duquerroy, Stéphane; Galloux, Marie; Loney, Colin; Conner, Edward; Eléouët, Jean-François; Rey, Félix A; Bhella, David

    2013-08-01

    Respiratory syncytial virus (RSV) is an important human pathogen. Its nucleocapsid (NC), which comprises the negative sense RNA viral genome coated by the viral nucleoprotein N, is a critical assembly that serves as template for both mRNA synthesis and genome replication. We have previously described the X-ray structure of an NC-like structure: a decameric ring formed of N-RNA that mimics one turn of the helical NC. In the absence of experimental data we had hypothesized that the NC helix would be right-handed, as the N-N contacts in the ring appeared to more easily adapt to that conformation. We now unambiguously show that the RSV NC is a left-handed helix. We further show that the contacts in the ring can be distorted to maintain key N-N-protein interactions in a left-handed helix, and discuss the implications of the resulting atomic model of the helical NC for viral replication and transcription.

  5. Respiratory syncytial virus alters electrophysiologic properties in cotton rat airway epithelium.

    PubMed

    Cloutier, M M; Wong, D; Ogra, P L

    1989-01-01

    The effect of respiratory syncytial virus (RSV) infection on the electrophysiologic properties of the airway epithelium was studied in tracheas obtained from cotton rats, after in vivo exposure to the virus. RSV infection was documented by tissue culture infectivity and immunofluorescent antibody techniques. Light microscopic studies of the tracheas 72 hours after exposure to RSV revealed normal epithelial morphology. RSV infection produced a 28% decrease in short-circuit current (Isc) and an almost 100% increase in tissue resistance. The decrease in Isc was partially attributed to a decrease in Cl secretion, while the increase in tissue resistance was associated with significant restriction to sodium and chloride movement through the paracellular pathway in RSV-infected tracheas. We conclude from these studies that RSV infection in the cotton rat trachea produces significant changes in active and passive ion flows. Alterations in ion transport across the airway epithelium may result in changes in secondary water transport across the airways and may contribute to the pathophysiology of RSV bronchiolitis and other locally invasive mucosal viral infections in children.

  6. Direct Inhibition of Cellular Fatty Acid Synthase Impairs Replication of Respiratory Syncytial Virus and Other Respiratory Viruses.

    PubMed

    Ohol, Yamini M; Wang, Zhaoti; Kemble, George; Duke, Gregory

    2015-01-01

    Fatty acid synthase (FASN) catalyzes the de novo synthesis of palmitate, a fatty acid utilized for synthesis of more complex fatty acids, plasma membrane structure, and post-translational palmitoylation of host and viral proteins. We have developed a potent inhibitor of FASN (TVB-3166) that reduces the production of respiratory syncytial virus (RSV) progeny in vitro from infected human lung epithelial cells (A549) and in vivo from mice challenged intranasally with RSV. Addition of TVB-3166 to the culture medium of RSV-infected A549 cells reduces viral spread without inducing cytopathic effects. The antiviral effect of the FASN inhibitor is a direct consequence of reducing de novo palmitate synthesis; similar doses are required for both antiviral activity and inhibition of palmitate production, and the addition of exogenous palmitate to TVB-3166-treated cells restores RSV production. TVB-3166 has minimal effect on RSV entry but significantly reduces viral RNA replication, protein levels, viral particle formation and infectivity of released viral particles. TVB-3166 substantially impacts viral replication, reducing production of infectious progeny 250-fold. In vivo, oral administration of TVB-3166 to RSV-A (Long)-infected BALB/c mice on normal chow, starting either on the day of infection or one day post-infection, reduces RSV lung titers 21-fold and 9-fold respectively. Further, TVB-3166 also inhibits the production of RSV B, human parainfluenza 3 (PIV3), and human rhinovirus 16 (HRV16) progeny from A549, HEp2 and HeLa cells respectively. Thus, inhibition of FASN and palmitate synthesis by TVB-3166 significantly reduces RSV progeny both in vitro and in vivo and has broad-spectrum activity against other respiratory viruses. FASN inhibition may alter the composition of regions of the host cell membrane where RSV assembly or replication occurs, or change the membrane composition of RSV progeny particles, decreasing their infectivity.

  7. Direct Inhibition of Cellular Fatty Acid Synthase Impairs Replication of Respiratory Syncytial Virus and Other Respiratory Viruses

    PubMed Central

    Ohol, Yamini M.; Wang, Zhaoti; Kemble, George; Duke, Gregory

    2015-01-01

    Fatty acid synthase (FASN) catalyzes the de novo synthesis of palmitate, a fatty acid utilized for synthesis of more complex fatty acids, plasma membrane structure, and post-translational palmitoylation of host and viral proteins. We have developed a potent inhibitor of FASN (TVB-3166) that reduces the production of respiratory syncytial virus (RSV) progeny in vitro from infected human lung epithelial cells (A549) and in vivo from mice challenged intranasally with RSV. Addition of TVB-3166 to the culture medium of RSV-infected A549 cells reduces viral spread without inducing cytopathic effects. The antiviral effect of the FASN inhibitor is a direct consequence of reducing de novo palmitate synthesis; similar doses are required for both antiviral activity and inhibition of palmitate production, and the addition of exogenous palmitate to TVB-3166-treated cells restores RSV production. TVB-3166 has minimal effect on RSV entry but significantly reduces viral RNA replication, protein levels, viral particle formation and infectivity of released viral particles. TVB-3166 substantially impacts viral replication, reducing production of infectious progeny 250-fold. In vivo, oral administration of TVB-3166 to RSV-A (Long)-infected BALB/c mice on normal chow, starting either on the day of infection or one day post-infection, reduces RSV lung titers 21-fold and 9-fold respectively. Further, TVB-3166 also inhibits the production of RSV B, human parainfluenza 3 (PIV3), and human rhinovirus 16 (HRV16) progeny from A549, HEp2 and HeLa cells respectively. Thus, inhibition of FASN and palmitate synthesis by TVB-3166 significantly reduces RSV progeny both in vitro and in vivo and has broad-spectrum activity against other respiratory viruses. FASN inhibition may alter the composition of regions of the host cell membrane where RSV assembly or replication occurs, or change the membrane composition of RSV progeny particles, decreasing their infectivity. PMID:26659560

  8. The impact of viral dynamics on the clinical severity of infants with respiratory syncytial virus bronchiolitis.

    PubMed

    Zhou, Lili; Xiao, Qiuyan; Zhao, Yao; Huang, Ailong; Ren, Luo; Liu, Enmei

    2015-08-01

    The impact of dynamic respiratory syncytial virus (RSV) load on the clinical severity of hospitalized infants with bronchiolitis has not been clarified. Nasopharyngeal aspirates were obtained from 60 infants who were diagnosed with bronchiolitis within 96 hr of wheezing onset upon admission and on days 3, 5, and 7 in the hospital, and 17 respiratory viruses were detected. The RSV load was quantified by real-time qPCR for RSV subtypes A and B at different time points. Scoring criteria were used to evaluate the degree of severity. A total of 40 infants were determined to be RSV-positive, nine were identified as RSV subtype A (RSVA), and 31 were RSV subtype B (RSVB). The peak RSV load was observed upon admission, and the RSV load decreased significantly over time; in addition, this decrease began to have significant differences on day 5. There was a positive correlation between the RSV load and the clinical score (r(2)  = 0.121 and P < 0.001). According to the clinical scores, the infants in the severe group tended to have higher RSV loads than those in the moderate and mild groups. Multivariate logistic regression models revealed that the viral load on day 3 was independently associated with the degree of severity. This study elucidated that a higher mean RSV load was associated with a more severe disease and a longer duration of hospitalization and symptoms. This study also clarified RSV replication in infants and provides a theoretical basis for specifying an anti-RSV therapy strategy.

  9. Chimpanzee adenovirus- and MVA-vectored respiratory syncytial virus vaccine is safe and immunogenic in adults.

    PubMed

    Green, Christopher A; Scarselli, Elisa; Sande, Charles J; Thompson, Amber J; de Lara, Catherine M; Taylor, Kathryn S; Haworth, Kathryn; Del Sorbo, Mariarosaria; Angus, Brian; Siani, Loredana; Di Marco, Stefania; Traboni, Cinzia; Folgori, Antonella; Colloca, Stefano; Capone, Stefania; Vitelli, Alessandra; Cortese, Riccardo; Klenerman, Paul; Nicosia, Alfredo; Pollard, Andrew J

    2015-08-12

    Respiratory syncytial virus (RSV) causes respiratory infection in annual epidemics, with infants and the elderly at particular risk of developing severe disease and death. However, despite its importance, no vaccine exists. The chimpanzee adenovirus, PanAd3-RSV, and modified vaccinia virus Ankara, MVA-RSV, are replication-defective viral vectors encoding the RSV fusion (F), nucleocapsid (N), and matrix (M2-1) proteins for the induction of humoral and cellular responses. We performed an open-label, dose escalation, phase 1 clinical trial in 42 healthy adults in which four different combinations of prime/boost vaccinations were investigated for safety and immunogenicity, including both intramuscular (IM) and intranasal (IN) administration of the adenovirus-vectored vaccine. The vaccines were safe and well tolerated, with the most common reported adverse events being mild injection site reactions. No vaccine-related serious adverse events occurred. RSV neutralizing antibody titers rose in response to IM prime with PanAd3-RSV and after IM boost for individuals primed by the IN route. Circulating anti-F immunoglobulin G (IgG) and IgA antibody-secreting cells (ASCs) were observed after the IM prime and IM boost. RSV-specific T cell responses were increased after the IM PanAd3-RSV prime and were most efficiently boosted by IM MVA-RSV. Interferon-γ (IFN-γ) secretion after boost was from both CD4(+) and CD8(+) T cells, without detectable T helper cell 2 (TH2) cytokines that have been previously associated with immune pathogenesis following exposure to RSV after the formalin-inactivated RSV vaccine. In conclusion, PanAd3-RSV and MVA-RSV are safe and immunogenic in healthy adults. These vaccine candidates warrant further clinical evaluation of efficacy to assess their potential to reduce the burden of RSV disease. PMID:26268313

  10. Epidemic activity of respiratory syncytial virus is related to temperature and rainfall in equatorial tropical countries.

    PubMed

    Gamba-Sanchez, N; Rodriguez-Martinez, C E; Sossa-Briceño, M P

    2016-07-01

    Although viral acute lower respiratory infections (ALRIs) are a major public health problem in tropical low- and middle-income countries (LMICs), and there is growing evidence showing their relationship with meteorological parameters, studies performed in these countries are scarce. In an analytical cross-sectional study, we determined which of the main meteorological parameters (temperature, absolute humidity, rainfall, wind speed, and solar radiation) predicted respiratory syncytial virus (RSV) activity in a population of hospitalized children with ALRI during a 5-year period, from January 2009 to December 2013. Out of a total of 4559 children included in the study (mean age 9·2 ± 8·5 months), 2953 (64·8%) presented RSV infection during the 3-month period from March to May. In the multivariate analysis, after controlling for absolute humidity, wind speed, and solar radiation, temperature [incidence rate ratio (IRR) 2·25, 95% confidence interval (CI) 1·11-4·53, P = 0·024], and rainfall (IRR 1·01, 95% CI 1·00-1·02, P = 0·044) were independently associated with the monthly number of RSV infections. In conclusion, in Bogota, the capital of a tropical LMIC lying slightly above the equator, RSV activity peaks in the 3-month period from March to May, the main rainy period of the year in the city. In addition, rainfall and temperature are the two most important meteorological parameters that are independently associated with RSV activity in hospitalized children with ALRI in the city. PMID:26888544

  11. Marked induction of matrix metalloproteinase-10 by respiratory syncytial virus infection in human nasal epithelial cells.

    PubMed

    Hirakawa, Satoshi; Kojima, Takashi; Obata, Kazufumi; Okabayashi, Tamaki; Yokota, Shin-Ichi; Nomura, Kazuaki; Obonai, Toshimasa; Fuchimoto, Jun; Himi, Tetsuo; Tsutsumi, Hiroyuki; Sawada, Norimasa

    2013-12-01

    Respiratory syncytial virus (RSV) is an important pathogen of bronchiolitis, asthma, and severe lower respiratory tract disease in infants and young children. Matrix metalloproteinases (MMPs) play key roles in viral infection, inflammation and remodeling of the airway. However, the roles and regulation of MMPs in human nasal epithelial cells (HNECs) after RSV infection remain unclear. To investigate the regulation of MMP induced after RSV infection in HNECs, an RSV-infected model of HNECs in vitro was used. It was found that mRNA of MMP-10 was markedly increased in HNECs after RSV infection, together with induction of mRNAs of MMP-1, -7, -9, and -19. The amount of MMP-10 released from HNECs was also increased in a time-dependent manner after RSV infection as was that of chemokine RANTES. The upregulation of MMP-10 in HNECs after RSV infection was prevented by inhibitors of NF-κB and pan-PKC with inhibition of RSV replication, whereas it was prevented by inhibitors of JAK/STAT, MAPK, and EGF receptors without inhibition of RSV replication. In lung tissue of an infant with severe RSV infection in which a few RSV antibody-positive macrophages were observed, MMP-10 was expressed at the apical side of the bronchial epithelial cells and alveolar epithelial cells. In conclusion, MMP-10 induced by RSV infection in HNECs is regulated via distinct signal transduction pathways with or without relation to RSV replication. MMP-10 may play an important role in the pathogenesis of RSV diseases and it has the potential to be a novel marker and therapeutic target for RSV infection.

  12. Chimpanzee adenovirus- and MVA-vectored respiratory syncytial virus vaccine is safe and immunogenic in adults.

    PubMed

    Green, Christopher A; Scarselli, Elisa; Sande, Charles J; Thompson, Amber J; de Lara, Catherine M; Taylor, Kathryn S; Haworth, Kathryn; Del Sorbo, Mariarosaria; Angus, Brian; Siani, Loredana; Di Marco, Stefania; Traboni, Cinzia; Folgori, Antonella; Colloca, Stefano; Capone, Stefania; Vitelli, Alessandra; Cortese, Riccardo; Klenerman, Paul; Nicosia, Alfredo; Pollard, Andrew J

    2015-08-12

    Respiratory syncytial virus (RSV) causes respiratory infection in annual epidemics, with infants and the elderly at particular risk of developing severe disease and death. However, despite its importance, no vaccine exists. The chimpanzee adenovirus, PanAd3-RSV, and modified vaccinia virus Ankara, MVA-RSV, are replication-defective viral vectors encoding the RSV fusion (F), nucleocapsid (N), and matrix (M2-1) proteins for the induction of humoral and cellular responses. We performed an open-label, dose escalation, phase 1 clinical trial in 42 healthy adults in which four different combinations of prime/boost vaccinations were investigated for safety and immunogenicity, including both intramuscular (IM) and intranasal (IN) administration of the adenovirus-vectored vaccine. The vaccines were safe and well tolerated, with the most common reported adverse events being mild injection site reactions. No vaccine-related serious adverse events occurred. RSV neutralizing antibody titers rose in response to IM prime with PanAd3-RSV and after IM boost for individuals primed by the IN route. Circulating anti-F immunoglobulin G (IgG) and IgA antibody-secreting cells (ASCs) were observed after the IM prime and IM boost. RSV-specific T cell responses were increased after the IM PanAd3-RSV prime and were most efficiently boosted by IM MVA-RSV. Interferon-γ (IFN-γ) secretion after boost was from both CD4(+) and CD8(+) T cells, without detectable T helper cell 2 (TH2) cytokines that have been previously associated with immune pathogenesis following exposure to RSV after the formalin-inactivated RSV vaccine. In conclusion, PanAd3-RSV and MVA-RSV are safe and immunogenic in healthy adults. These vaccine candidates warrant further clinical evaluation of efficacy to assess their potential to reduce the burden of RSV disease.

  13. Respiratory syncytial virus promotes Moraxella catarrhalis-induced ascending experimental otitis media.

    PubMed

    Brockson, M Elizabeth; Novotny, Laura A; Jurcisek, Joseph A; McGillivary, Glen; Bowers, Martha R; Bakaletz, Lauren O

    2012-01-01

    Otitis media (OM) is a polymicrobial disease wherein prior or concurrent infection with an upper respiratory tract virus plays an essential role, predisposing the middle ear to bacterial invasion. In episodes of acute bacterial OM, respiratory syncytial virus (RSV) is the most commonly isolated virus and thus serves as an important co-pathogen. Of the predominant bacterial agents of OM, the pathogenesis of disease due to Moraxella catarrhalis is the least well understood. Rigorous study of M. catarrhalis in the context of OM has been significantly hindered by lack of an animal model. To bridge this gap, we assessed whether co-infection of chinchillas with M. catarrhalis and RSV would facilitate ascension of M. catarrhalis from the nasopharynx into the middle ear. Chinchillas were challenged intranasally with M. catarrhalis followed 48 hours later by intranasal challenge with RSV. Within 7 days, 100% of nasopharynges were colonized with M. catarrhalis and homogenates of middle ear mucosa were also culture-positive. Moreover, within the middle ear space, the mucosa exhibited hemorrhagic foci, and a small volume of serosanguinous effusion was present in one of six ears. To improve upon this model, and based on epidemiologic data, nontypeable Haemophilus influenzae (NTHI) was included as an additional bacterial co-pathogen via intranasal administration four days before M. catarrhalis challenge. With this latter protocol, M. catarrhalis was cultured from the nasopharynx and middle ear homogenates of a maximum of 88% and 79% animals, respectively, for up to 17 days after intranasal challenge with M. catarrhalis. Additionally, hemorrhagic foci were observed in 79% of middle ears upon sacrifice. Thus, these data demonstrated that co-infection with RSV and NTHI predisposed to M. catarrhalis-induced ascending experimental OM. This model can be used both in studies of pathogenesis as well as to investigate strategies to prevent or treat OM due to M. catarrhalis.

  14. A Randomized Placebo Controlled Trial of Ibuprofen for Respiratory Syncytial Virus Infection in a Bovine Model

    PubMed Central

    Walsh, Paul; Behrens, Nicole; Carvallo Chaigneau, Francisco R.; McEligot, Heather; Agrawal, Karan; Newman, John W.; Anderson, Mark; Gershwin, Laurel J.

    2016-01-01

    Background Respiratory syncytial virus (RSV) is the most common cause of bronchiolitis and hospital admission in infants. An analogous disease occurs in cattle and costs US agriculture a billion dollars a year. RSV causes much of its morbidity indirectly via adverse effects of the host response to the virus. RSV is accompanied by elevated prostaglandin E2 (PGE2) which is followed by neutrophil led inflammation in the lung. Ibuprofen is a prototypical non-steroidal anti-inflammatory drug that decreases PGE2 levels by inhibiting cyclooxygenase. Hypotheses We hypothesized that treatment of RSV with ibuprofen would decrease PGE2 levels, modulate the immune response, decrease clinical illness, and decrease the histopathological lung changes in a bovine model of RSV. We further hypothesized that viral replication would be unaffected. Methods We performed a randomized placebo controlled trial of ibuprofen in 16 outbred Holstein calves that we infected with RSV. We measured clinical scores, cyclooxygenase, lipoxygenase and endocannabinoid products in plasma and mediastinal lymph nodes and interleukin (Il)-4, Il-13, Il-17 and interferon-γ in mediastinal lymph nodes. RSV shedding was measured daily and nasal Il-6, Il-8 and Il-17 every other day. The calves were necropsied on Day 10 post inoculation and histology performed. Results One calf in the ibuprofen group required euthanasia on Day 8 of infection for respiratory distress. Clinical scores (p<0.01) and weight gain (p = 0.08) seemed better in the ibuprofen group. Ibuprofen decreased cyclooxygenase, lipoxygenase, and cytochrome P450 products, and increased monoacylglycerols in lung lymph nodes. Ibuprofen modulated the immune response as measured by narrowed range of observed Il-13, Il-17 and IFN-γ gene expression in mediastinal lymph nodes. Lung histology was not different between groups, and viral shedding was increased in calves randomized to ibuprofen. Conclusions Ibuprofen decreased PGE2, modulated the immune

  15. Revealing the binding mode between respiratory syncytial virus fusion protein and benzimidazole-based inhibitors.

    PubMed

    Ji, Dingjue; Ye, Wei; Chen, HaiFeng

    2015-07-01

    Human respiratory syncytial virus (HRSV) is a major respiratory pathogen in newborn infants and young children and can also be a threat to some elderly and high-risk adults with chronic pulmonary disease and the severely immunocompromised. The RSV fusion (RSVF) protein has been an attractive target for vaccine and drug development. Experimental results indicate a series of benzimidazole-based inhibitors which target RSVF protein to inhibit the viral entry of RSV. To reveal the binding mode between these inhibitors and RSVF protein, molecular docking and molecular dynamics simulations were used to investigate the interactions between the inhibitors and the core domain of RSVF protein. MD results suggest that the active molecules have stronger π-π stacking, cation-π, and other interactions than less active inhibitors. The binding free energy between the active inhibitor and RSVF protein is also found to be significantly lower than that of the less active one using MM/GBSA. Then, Comparative Molecular Field Analysis (CoMFA) and Comparative Molecular Similarity Indices Analysis (CoMSIA) methods were used to construct three dimensional quantitative structure-activity (3D-QSAR) models. The cross-validated q(2) values are found to be 0.821 and 0.795 for CoMFA and CoMSIA, respectively. And the non-cross-validated r(2) values are 0.973 and 0.961. Ninety-two test set compounds validated these models. The results suggest that these models are robust with good prediction abilities. Furthermore, these models reveal possible methods to improve the bioactivity of inhibitors. PMID:25872614

  16. Quantitative proteomic analysis of A549 cells infected with human respiratory syncytial virus.

    PubMed

    Munday, Diane C; Emmott, Edward; Surtees, Rebecca; Lardeau, Charles-Hugues; Wu, Weining; Duprex, W Paul; Dove, Brian K; Barr, John N; Hiscox, Julian A

    2010-11-01

    Human respiratory syncytial virus (HRSV) is a major cause of pediatric lower respiratory tract disease to which there is no vaccine or efficacious chemotherapeutic strategy. Although RNA synthesis and virus assembly occur in the cytoplasm, HRSV is known to induce nuclear responses in the host cell as replication alters global gene expression. Quantitative proteomics was used to take an unbiased overview of the protein changes in transformed human alveolar basal epithelial cells infected with HRSV. Underpinning this was the use of stable isotope labeling with amino acids in cell culture coupled to LC-MS/MS, which allowed the direct and simultaneous identification and quantification of both cellular and viral proteins. To reduce sample complexity and increase data return on potential protein localization, cells were fractionated into nuclear and cytoplasmic extracts. This resulted in the identification of 1,140 cellular proteins and six viral proteins. The proteomics data were analyzed using Ingenuity Pathways Analysis to identify defined canonical pathways and functional groupings. Selected data were validated using Western blot, direct and indirect immunofluorescence confocal microscopy, and functional assays. The study served to validate and expand upon known HRSV-host cell interactions, including those associated with the antiviral response and alterations in subnuclear structures such as the nucleolus and ND10 (promyelocytic leukemia bodies). In addition, novel changes were observed in mitochondrial proteins and functions, cell cycle regulatory molecules, nuclear pore complex proteins and nucleocytoplasmic trafficking proteins. These data shed light into how the cell is potentially altered to create conditions more favorable for infection. Additionally, the study highlights the application and advantage of stable isotope labeling with amino acids in cell culture coupled to LC-MS/MS for the analysis of virus-host interactions.

  17. Progress in understanding and controlling respiratory syncytial virus: still crazy after all these years

    PubMed Central

    Collins, Peter L.; Melero, José A.

    2011-01-01

    Human respiratory syncytial virus (RSV) is a ubiquitous pathogen that infects everyone worldwide early in life and is a leading cause of severe lower respiratory tract disease in the pediatric population as well as in the elderly and in profoundly immunosuppressed individuals. RSV is an enveloped, nonsegmented negative-sense RNA virus that is classified in Family Paramyxoviridae and is one of its more complex members. Although the replicative cycle of RSV follows the general pattern of the Paramyxoviridae, it encodes additional proteins. Two of these (NS1 and NS2) inhibit the host type I and type III interferon (IFN) responses, among other functions, and another gene encodes two novel RNA synthesis factors (M2-1 and M2-2). The attachment (G) glycoprotein also exhibits unusual features, such as high sequence variability, extensive glycosylation, cytokine mimicry, and a shed form that helps the virus evade neutralizing antibodies. RSV is notable for being able to efficiently infect early in life, with the peak of hospitalization at 2–3 months of age. It also is notable for the ability to reinfect symptomatically throughout life without need for significant antigenic change, although immunity from prior infection reduces disease. It is widely thought that re-infection is due to an ability of RSV to inhibit or subvert the host immune response. Mechanisms of viral pathogenesis remain controversial. RSV is notable for a historic, tragic pediatric vaccine failure involving a formalin-inactivated virus preparation that was evaluated in the 1960’s and that was poorly protective and paradoxically primed for enhanced RSV disease. RSV also is notable for the development of a successful strategy for passive immunoprophylaxis of high-risk infants using RSV-neutralizing antibodies. Vaccines and new antiviral drugs are in pre-clinical and clinical development, but controlling RSV remains a formidable challenge. PMID:21963675

  18. Risk Factors for Severe Respiratory Syncytial Virus Lower Respiratory Tract Infection

    PubMed Central

    Sommer, Constanze; Resch, Bernhard; Simões, Eric A.F

    2011-01-01

    RSV infection is a leading cause of lower respiratory tract infection, especially in High-risk infants with a history of prematurity, bronchopulmonary dysplasia (BPD), congenital heart disease (CHD), neuromusculair impairment, immunodeficiency, and Down syndrome. Host related risk factors that have been identified to be associated with severe RSV related lower respiratory tract infection include young age below 6 months at the beginning of RSV season, multiple birth, male sex, low socioeconomic status and parental education, crowded living conditions, young siblings, maternal smoking and indoor smoke pollution, malnutrition/small for gestational age, family history of atopy or asthma, low cord serum RSV antibody titers, and living at altitude. Risk factors increasing the risk of acquisition of RSV have been identified to be birth before and/or during RSV season, day care attendance, presence of older siblings in school or day-care, and lack of breast feeding. Some of these risk factors are discussed controversially and some of them are found continuously throughout the literature. Given the high cost of RSV prophylaxis, especially for the large population of late preterm infants, algorithms and risk score systems have been published that could identify high-risk infants for treatment with palivizumab out of this gestational age group. Several models reported on an average sensitivity and specificity of 70 percent and, thus, are helpful to identify infants at high risk for severe RSV infection and need for prophylaxis with palivizumab. PMID:22262987

  19. Differential expression of cytokines in response to respiratory syncytial virus infection of calves with high or low circulating 25-hydroxyvitamin D3

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Deficiency of serum levels of 25-hydroxyvitamin D3 has been related to increased risk of lower respiratory tract infections (LRTI) in children. Respiratory syncytial virus (RSV) is the leading cause of LRTI in infants and young children. The neonatal calf model of RSV infection shares many features ...

  20. A comprehensive proteomic view of responses of A549 type II alveolar epithelial cells to human respiratory syncytial virus infection.

    PubMed

    Dave, Keyur A; Norris, Emma L; Bukreyev, Alexander A; Headlam, Madeleine J; Buchholz, Ursula J; Singh, Toshna; Collins, Peter L; Gorman, Jeffrey J

    2014-12-01

    Human respiratory syncytial virus is a major respiratory pathogen for which there are no suitable antivirals or vaccines. A better understanding of the host cell response to this virus may redress this problem. The present report concerns analysis of multiple independent biological replicates of control and 24 h infected lysates of A549 cells by two different proteomic workflows. One workflow involved fractionation of lysates by in-solution protein IEF and individual fractions were digested using trypsin prior to capillary HPLC-LTQ-OrbitrapXL-MS/MS. A second workflow involved digestion of whole cell lysates and analysis by nanoUltraHPLC-LTQ-OrbitrapElite-MS/MS. Both workflows resulted in the quantification of viral proteins exclusively in lysates of infected cells in the relative abundances anticipated from previous studies. Unprecedented numbers (3247 - 5010) of host cell protein groups were also quantified and the infection-specific regulation of a large number (191) of these protein groups was evident based on a stringent false discovery rate cut-off (<1%). Bioinformatic analyses revealed that most of the regulated proteins were potentially regulated by type I, II, and III interferon, TNF-α and noncanonical NF-κB2 mediated antiviral response pathways. Regulation of specific protein groups by infection was validated by quantitative Western blotting and the cytokine-/key regulator-specific nature of their regulation was confirmed by comparable analyses of cytokine treated A549 cells. Overall, it is evident that the workflows described herein have produced the most comprehensive proteomic characterization of host cell responses to human respiratory syncytial virus published to date. These workflows will form the basis for analysis of the impacts of specific genes of human respiratory syncytial virus responses of A549 and other cell lines using a gene-deleted version of the virus. They should also prove valuable for the analysis of the impact of other infectious

  1. A Comprehensive Proteomic View of Responses of A549 Type II Alveolar Epithelial Cells to Human Respiratory Syncytial Virus Infection*

    PubMed Central

    Dave, Keyur A.; Norris, Emma L.; Bukreyev, Alexander A.; Headlam, Madeleine J.; Buchholz, Ursula J.; Singh, Toshna; Collins, Peter L.; Gorman, Jeffrey J.

    2014-01-01

    Human respiratory syncytial virus is a major respiratory pathogen for which there are no suitable antivirals or vaccines. A better understanding of the host cell response to this virus may redress this problem. The present report concerns analysis of multiple independent biological replicates of control and 24 h infected lysates of A549 cells by two different proteomic workflows. One workflow involved fractionation of lysates by in-solution protein IEF and individual fractions were digested using trypsin prior to capillary HPLC-LTQ-OrbitrapXL-MS/MS. A second workflow involved digestion of whole cell lysates and analysis by nanoUltraHPLC-LTQ-OrbitrapElite-MS/MS. Both workflows resulted in the quantification of viral proteins exclusively in lysates of infected cells in the relative abundances anticipated from previous studies. Unprecedented numbers (3247 - 5010) of host cell protein groups were also quantified and the infection-specific regulation of a large number (191) of these protein groups was evident based on a stringent false discovery rate cut-off (<1%). Bioinformatic analyses revealed that most of the regulated proteins were potentially regulated by type I, II, and III interferon, TNF-α and noncanonical NF-κB2 mediated antiviral response pathways. Regulation of specific protein groups by infection was validated by quantitative Western blotting and the cytokine-/key regulator-specific nature of their regulation was confirmed by comparable analyses of cytokine treated A549 cells. Overall, it is evident that the workflows described herein have produced the most comprehensive proteomic characterization of host cell responses to human respiratory syncytial virus published to date. These workflows will form the basis for analysis of the impacts of specific genes of human respiratory syncytial virus responses of A549 and other cell lines using a gene-deleted version of the virus. They should also prove valuable for the analysis of the impact of other infectious

  2. Respiratory Syncytial Virus Bronchiolitis in Children up to 5 Years of Age in Spain: Epidemiology and Comorbidities

    PubMed Central

    Gil-Prieto, Ruth; Gonzalez-Escalada, Alba; Marín-García, Patricia; Gallardo-Pino, Carmen; Gil-de-Miguel, Angel

    2015-01-01

    Abstract This epidemiological survey in Spain estimates the burden of respiratory syncytial virus (RSV) infection in children up to 5 year of age during a 15-year period (1997–2011). Observational retrospective survey was conducted by reviewing data of the National Surveillance System for Hospital Data, including >98% of Spanish hospitals. All hospitalizations related to RSV infection for children up to 5 years, reported during 1997–2011 period, were analyzed. Codes were selected by using the International Classification of Diseases 9th Clinical Modification 466.0–466.19, 480.1, and 079.6. A total of 326,175 and 286,007 hospital discharges for children up to 5 and 2 years of age were reported during the study period. The annual incidence was 1072 and 2413 patients per 100,000, respectively. The average length of hospital stay was 5.7 (standard deviation 8.2) days. Four hundred forty-six deaths were reported; of those, 403 occurred in children <2 years and 355 (80%) occurred in children <12 months of age. Hospitalization and mortality rates were significantly higher in boys and decrease significantly with age. The higher rate of hospitalization and mortality rates were found in the first year of life. Annual average cost for National Health Care System was € 47 M with a mean hospitalization cost of €2162. The average length of hospitalization and costs were significantly higher in high-risk children. RSV infections in children up to 5 year of age still pose a significant health threat in Spain, especially in the infants. The development of preventive, diagnostic, and therapeutic guidelines focused in children with comorbidities may help reduce the hospital and economic burden of the disease. PMID:26020386

  3. Respiratory syncytial virus genotypes circulating in urban Ghana: february to november 2006

    PubMed Central

    Obodai, Evangeline; Asmah, Richard; Boamah, Isaac; Goka, Bamenla; Odoom, John Kofi; Adiku, Theophilus

    2014-01-01

    Introduction Respiratory syncytial virus (RSV) is the major cause of acute lower respiratory tract infection (ALRI) in young children. RSV strains have been divided into 2 major antigenic groups (A and B), which are further divided into several genotypes, but very little is known about its circulating genotypes in Ghana. This study characterized RSV genotypes detected in children with ALRI in Accra between February and November 2006. Methods Nasopharyngeal aspirates (NPA) were obtained from children diagnosed with ALRI between February and November 2006. The NPA were screened for RSV using a nested multiplex reverse transcriptase polymerase chain reaction (RT-PCR) method for genotyping RSV. Viral RNA was extracted from the NPA using guanidinium isothiocyanate method and purified with an RNAID commercial kit. Care-givers gave their consent prior to specimen collection. Administered questionnaires captured information on patient demographic and clinical history. Results A total of 53 children were enrolled in the study with a male to female ratio of 3:1. Of the 53 NPA analyzed, 60.4% (32/53) were positive for RSV. Subsequent genotypic analysis showed that 72% (23/32) of the 60.4% RSV infections were RSV B only and 28% (9/32) were co-infections of both RSV A and B. Children between the ages of 2 - 12 months were the most affected age group per an RSV infection rate of 37.5% (12/32). No significant difference was detected in the recovery rate of ALRI (98.1%) and RSV (96.9%) positive patients from the infection. One patient died resulting in a mortality rate of 3.1%. Bronchopneumonia (20 out of 32 cases) was the major diagnosis on admission. RSV infection was seasonal dependent, described by 2 peaks in October and April-May. Conclusion Both RSV A and RSV B genotypes co-circulated during the study period with RSV B predominating. RSV may possibly be the main pathogen of lower respiratory tract illness during epidemics in the wet seasons. Genotyping by the multiplex RT

  4. Systemic inflammatory response syndrome and prolonged hypoperfusion lesions in an infant with respiratory syncytial virus encephalopathy.

    PubMed

    Miyamoto, Kenji; Fujisawa, Masahide; Hozumi, Hajime; Tsuboi, Tatsuo; Kuwashima, Shigeko; Hirao, Jun-ichi; Sugita, Kenichi; Arisaka, Osamu

    2013-10-01

    Respiratory syncytial virus (RSV) is a cause of neurological complications in infants. We report a rare case of RSV encephalopathy in an infant who presented with poor sucking and hypothermia at 17 days of age after suffering from rhinorrhea and a cough for several days. After hospitalization, the patient presented with stupor and hypotonia lasting for at least 24 h, and was intubated, sedated, and ventilated for treatment of pneumonia. These symptoms led to diagnosis of pediatric systemic inflammatory response syndrome (SIRS) caused by RSV infection. High-dose steroid therapy was combined with artificial ventilation because the initial ventilation therapy was ineffective. Interleukin (IL)-6 levels in spinal fluid were markedly increased upon admission, and serum IL-6 and IL-8 levels showed even greater elevation. The patient was diagnosed with RSV encephalopathy. On day 5, high signal intensity in the bilateral hippocampus was observed on diffusion-weighted magnetic resonance imaging (MRI). On day 14, the patient presented with delayed partial seizure and an electroencephalogram showed occasional unilateral spikes in the parietal area, but the hippocampal abnormality had improved to normal on MRI. (99m)Tc-labeled ethylcysteinate dimer single-photon emission computed tomography (SPECT) on day 18 showed hypoperfusion of the bilateral frontal and parietal regions and the unilateral temporal region. SPECT at 3 months after onset still showed hypoperfusion of the bilateral frontal region and unilateral temporal region, but hypoperfusion of the bilateral parietal region had improved. The patient has no neurological deficit at 6 months. These findings suggest that RSV encephalopathy with cytokine storm induces several symptoms and complications, including SIRS and prolonged brain hypoperfusion on SPECT.

  5. Roflumilast Inhibits Respiratory Syncytial Virus Infection in Human Differentiated Bronchial Epithelial Cells

    PubMed Central

    Mata, Manuel; Martinez, Isidoro; Melero, Jose A.; Tenor, Herman; Cortijo, Julio

    2013-01-01

    Respiratory syncytial virus (RSV) causes acute exacerbations in COPD and asthma. RSV infects bronchial epithelial cells (HBE) that trigger RSV associated lung pathology. This study explores whether the phosphodiesterase 4 (PDE4) inhibitor Roflumilast N-oxide (RNO), alters RSV infection of well-differentiated HBE (WD-HBE) in vitro. WD-HBE were RSV infected in the presence or absence of RNO (0.1-100 nM). Viral infection (staining of F and G proteins, nucleoprotein RNA level), mRNA of ICAM-1, ciliated cell markers (digital high speed videomicroscopy, β-tubulin immunofluorescence, Foxj1 and Dnai2 mRNA), Goblet cells (PAS), mRNA of MUC5AC and CLCA1, mRNA and protein level of IL-13, IL-6, IL-8, TNFα, formation of H2O2 and the anti-oxidative armamentarium (mRNA of Nrf2, HO-1, GPx; total antioxidant capacity (TAC) were measured at day 10 or 15 post infection. RNO inhibited RSV infection of WD-HBE, prevented the loss of ciliated cells and markers, reduced the increase of MUC5AC and CLCA1 and inhibited the increase of IL-13, IL-6, IL-8, TNFα and ICAM-1. Additionally RNO reversed the reduction of Nrf2, HO-1 and GPx mRNA levels and consequently restored the TAC and reduced the H2O2 formation. RNO inhibits RSV infection of WD-HBE cultures and mitigates the cytopathological changes associated to this virus. PMID:23936072

  6. Respiratory Syncytial Virus Infections Enhance Cigarette Smoke Induced COPD in Mice

    PubMed Central

    Foronjy, Robert F.; Dabo, Abdoulaye J.; Taggart, Clifford C.; Weldon, Sinead; Geraghty, Patrick

    2014-01-01

    Respiratory syncytial viral (RSV) infections are a frequent cause of chronic obstructive pulmonary disease (COPD) exacerbations, which are a major factor in disease progression and mortality. RSV is able to evade antiviral defenses to persist in the lungs of COPD patients. Though RSV infection has been identified in COPD, its contribution to cigarette smoke-induced airway inflammation and lung tissue destruction has not been established. Here we examine the long-term effects of cigarette smoke exposure, in combination with monthly RSV infections, on pulmonary inflammation, protease production and remodeling in mice. RSV exposures enhanced the influx of macrophages, neutrophils and lymphocytes to the airways of cigarette smoke exposed C57BL/6J mice. This infiltration of cells was most pronounced around the vasculature and bronchial airways. By itself, RSV caused significant airspace enlargement and fibrosis in mice and these effects were accentuated with concomitant smoke exposure. Combined stimulation with both smoke and RSV synergistically induced cytokine (IL-1α, IL-17, IFN-γ, KC, IL-13, CXCL9, RANTES, MIF and GM-CSF) and protease (MMP-2, -8, -12, -13, -16 and cathepsins E, S, W and Z) expression. In addition, RSV exposure caused marked apoptosis within the airways of infected mice, which was augmented by cigarette smoke exposure. RSV and smoke exposure also reduced protein phosphatase 2A (PP2A) and protein tyrosine phosphates (PTP1B) expression and activity. This is significant as these phosphatases counter smoke-induced inflammation and protease expression. Together, these findings show for the first time that recurrent RSV infection markedly enhances inflammation, apoptosis and tissue destruction in smoke-exposed mice. Indeed, these results indicate that preventing RSV transmission and infection has the potential to significantly impact on COPD severity and progression. PMID:24587397

  7. Predicting the start week of respiratory syncytial virus outbreaks using real time weather variables

    PubMed Central

    2010-01-01

    Background Respiratory Syncytial Virus (RSV), a major cause of bronchiolitis, has a large impact on the census of pediatric hospitals during outbreak seasons. Reliable prediction of the week these outbreaks will start, based on readily available data, could help pediatric hospitals better prepare for large outbreaks. Methods Naïve Bayes (NB) classifier models were constructed using weather data from 1985-2008 considering only variables that are available in real time and that could be used to forecast the week in which an RSV outbreak will occur in Salt Lake County, Utah. Outbreak start dates were determined by a panel of experts using 32,509 records with ICD-9 coded RSV and bronchiolitis diagnoses from Intermountain Healthcare hospitals and clinics for the RSV seasons from 1985 to 2008. Results NB models predicted RSV outbreaks up to 3 weeks in advance with an estimated sensitivity of up to 67% and estimated specificities as high as 94% to 100%. Temperature and wind speed were the best overall predictors, but other weather variables also showed relevance depending on how far in advance the predictions were made. The weather conditions predictive of an RSV outbreak in our study were similar to those that lead to temperature inversions in the Salt Lake Valley. Conclusions We demonstrate that Naïve Bayes (NB) classifier models based on weather data available in real time have the potential to be used as effective predictive models. These models may be able to predict the week that an RSV outbreak will occur with clinical relevance. Their clinical usefulness will be field tested during the next five years. PMID:21044325

  8. Respiratory Syncytial Virus Aggravates Renal Injury through Cytokines and Direct Renal Injury

    PubMed Central

    Zhai, Songhui; Hu, Lijuan; Zhong, Lin; Guo, Yannan; Dong, Liqun; Jia, Ruizhen; Wang, Zheng

    2016-01-01

    The purpose of this study was to investigate the relationship between renal injury and reinfection that is caused by respiratory syncytial virus (RSV) and to analyze the mechanism of renal injury. Rats were repeatedly infected with RSV on days 4, 8, 14, and 28, then sacrificed and examined on day 56 after the primary infection. Renal injury was examined by transmission electron microscopy and histopathology. The F protein of RSV was detected in the renal tissue by indirect immunofluorescence. Proteinuria and urinary glycosaminoglycans (GAGs), serum levels of albumin, urea nitrogen, and creatinine, secretion of cytokines, T lymphocyte population and subsets, and dendritic cell (DC) activation state were examined. The results showed that renal injury was more serious in the reinfection group than in the primary infection group. At a higher infection dose, 6 × 106 PFU, the renal injury was more severe, accompanied by higher levels of proteinuria and urinary GAGs excretion, and lower levels of serum albumin. Podocyte foot effacement was more extensive, and hyperplasia of mesangial cells and proliferation of mesangial matrix were observed. The maturation state of DCs was specific, compared with the primary infection. There was also a decrease in the ratio of CD4+ to CD8+ T lymphocytes, due to an increase in the percentage of CD8+ T lymphocytes and a decrease in the percentage of CD4+ T lymphocytes, and a dramatic increase in the levels of IL-6 and IL-17. In terms of the different reinfection times, the day 14 reinfection group yielded the most serious renal injury and the most significant change in immune function. RSV F protein was still expressed in the glomeruli 56 days after RSV infection. Altogether, these results reveal that RSV infection could aggravate renal injury, which might be due to direct renal injury caused by RSV and the inflammatory lesions caused by the anti-virus response induced by RSV. PMID:27747195

  9. Quantification of respiratory syncytial virus polypeptides in nasal secretions by monoclonal antibodies.

    PubMed

    Hendry, R M; Godfrey, E; Anderson, L J; Fernie, B F; McIntosh, K

    1985-08-01

    An indirect enzyme-linked immunosorbent assay (ELISA) which uses monoclonal antibody as solid-phase immunosorbent was developed to measure specific polypeptides of respiratory syncytial virus (RSV). The assay was used to examine 43 nasopharyngeal (NP) aspirates from RSV-positive infants that had been examined previously for RSV by culture, direct immunofluorescence, and polyclonal antibody ELISA. Frozen NP aspirates were serially diluted and examined for the 66K mol. wt. fusion glycoprotein (F), the 84K large surface glycoprotein (G) and the 41K nucleoprotein (N) by monoclonal capture ELISA. F protein was detected in all 43 specimens, G protein was detectable in 20 (47%) and N protein in 22 (51%) of 43 NP aspirates. In specimens with detectable G and N proteins, F was detected by endpoint titration at approximately tenfold greater dilutions than either G or N. In 19 sequential NP aspirates from five patients with RSV infection, F was present in higher titre throughout infection. In 20 cases, matching cell culture isolates were examined by immunofluorescence with strain-specific monoclonal antibodies. Three of 20 isolates showed strain-specific differences by their lack of reaction with anti-G monoclonal antibody. Titration of the 20 cell culture isolates by monoclonal antibody capture ELISA showed the relative amount of F and N proteins to be equal in all cases, whereas levels of G protein tended to be slightly lower. Reconstruction experiments with NP aspirates demonstrated that degradation of F and N proteins did not occur in NP aspirates, but that G protein antigenicity appeared to be affected by nasal secretions. When compared with cell culture-grown material, nasal secretions contained abundant F protein but a surprisingly low concentration of N protein.

  10. Retrospective Multicenter Study of Respiratory Syncytial Virus Prophylaxis in Korean Children with Congenital Heart Diseases

    PubMed Central

    Kim, Ah Young; Jung, Se Yong; Choi, Jae Young; Kim, Gi Beom; Kim, Young-Hwue; Shim, Woo Sup; Kang, I-Seok

    2016-01-01

    Background and Objectives We conducted a review of current data on respiratory syncytial virus (RSV) prophylaxis with palivizumab, in Korean children with congenital heart diseases (CHD). In 2009, the Korean guideline for RSV prophylaxis had established up to five shots monthly per RSV season, only for children <1 year of age with hemodynamic significance CHD (HS-CHD). Subjects and Methods During the RSV seasons in 2009-2015, we performed a retrospective review of data for 466 infants with CHD, examined at six centers in Korea. Results Infants received an average of 3.7±1.9 (range, 1-10) injections during the RSV season. Fifty-seven HS-CHD patients (12.2%) were hospitalized with breakthrough RSV bronchiolitis, with a recurrence in three patients, one year after the initial check-up. Among patients with simple CHD, only five (1.1%) patients received one additional dose postoperatively, as per the limitations set by the Korean guideline. Among the 30 deaths (6.4%), five (1.1%) were attributed to RSV infection; three to simple CHD, one to Tetralogy of Fallot, and one to hypertrophic cardiomyopathy (HCM). Of the three HCM patients that exceeded guidelines for RSV prophylaxis, two (66.6%) were hospitalized, and one died of RSV infection (33.3%). Conclusion In accordance to the Korean guideline, minimal injections of palivizumab were administered to patients having HS-CHD

  11. Respiratory Syncytial Virus Persistence in Murine Macrophages Impairs IFN-β Response but Not Synthesis

    PubMed Central

    Rivera-Toledo, Evelyn; Torres-González, Laura; Gómez, Beatriz

    2015-01-01

    Type-I interferon (IFN-I) production is an early response to viral infection and pathogenic viruses have evolved multiple strategies to evade this cellular defense. Some viruses can establish and maintain persistent infections by altering the IFN-I signaling pathway. Here, we studied IFN-I synthesis and response in an in vitro model of persistent infection by respiratory syncytial virus (RSV) in a murine macrophage-like cell line. In this model, interferon regulatory factor 3 was constitutively active and located at nuclei of persistently infected cells, inducing expression of IFN-beta mRNA and protein. However, persistently infected macrophages did not respond in an autocrine manner to the secreted-IFN-beta or to recombinant-IFN-beta, since phosphorylated-STAT1 was not detected by western blot and transcription of the interferon-stimulated genes (ISGs) Mx1 and ISG56 was not induced. Treatment of non-infected macrophages with supernatants from persistently infected cells induced STAT1 phosphorylation and ISGs expression, mediated by the IFN-I present in the supernatants, because blocking the IFN-I receptor inhibited STAT1 phosphorylation. Results suggest that the lack of autocrine response to IFN-I by the host cell may be one mechanism for maintenance of RSV persistence. Furthermore, STAT1 phosphorylation and ISGs expression induced in non-infected cells by supernatants from persistently infected macrophages suggest that RSV persistence may trigger a proinflammatory phenotype in non-infected cells as part of the pathogenesis of RSV infection. PMID:26501312

  12. Macrophages are required for dendritic cell uptake of respiratory syncytial virus from an infected epithelium.

    PubMed

    Ugonna, Kelechi; Bingle, Colin D; Plant, Karen; Wilson, Kirsty; Everard, Mark L

    2014-01-01

    We have previously shown that the respiratory syncytial virus [RSV] can productively infect monocyte derived dendritic cells [MoDC] and remain dormant within the same cells for prolonged periods. It is therefore possible that infected dendritic cells act as a reservoir within the airways of individuals between annual epidemics. In the present study we explored the possibility that sub-epithelial DCs can be infected with RSV from differentiated bronchial epithelium and that in turn RSV from DCs can infect the epithelium. A dual co-culture model was established in which a differentiated primary airway epithelium on an Air Liquid Interface (ALI) was cultured on a transwell insert and MoDCs were subsequently added to the basolateral membrane of the insert. Further experiments were undertaken using a triple co-culture model in which in which macrophages were added to the apical surface of the differentiated epithelium. A modified RSV [rr-RSV] expressing a red fluorescent protein marker of replication was used to infect either the MoDCs or the differentiated epithelium and infection of the reciprocal cell type was assessed using confocal microscopy. Our data shows that primary epithelium became infected when rr-RSV infected MoDCs were introduced onto the basal surface of the transwell insert. MoDCs located beneath the epithelium did not become infected with virus from infected epithelial cells in the dual co-culture model. However when macrophages were present on the apical surface of the primary epithelium infection of the basal MoDCs occurred. Our data suggests that RSV infected dendritic cells readily transmit infection to epithelial cells even when they are located beneath the basal layer. However macrophages appear to be necessary for the transmission of infection from epithelial cells to basal dendritic cells.

  13. Analysis of respiratory syncytial virus genetic variability with amplified cDNAs.

    PubMed Central

    Sullender, W M; Sun, L; Anderson, L J

    1993-01-01

    Antigenic and genetic heterogeneities exist within the two major antigenic groups of respiratory syncytial (RS) virus. We developed a polymerase chain reaction (PCR)-based assay that not only differentiates the two RS virus groups but allows distinctions within groups on the basis of changes in the nucleotide sequences, as revealed by restriction fragment analysis. In this assay, viral RNA served as a template for cDNA synthesis with extension from a synthetic oligonucleotide primer complementary to bases 164 to 186 in the F protein mRNA. For PCR amplification, two group-specific 5' primers were added. The two primers corresponded to the G protein mRNA sequence of group B (bases 10 to 30) or group A (bases 247 to 267) RS virus. Agarose gel electrophoresis readily discriminated the 1.1-kb group B and the 0.9-kb group A virus amplification products. All 47 viruses tested were assigned to the same group by both PCR and monoclonal antibody reaction pattern analysis. Restriction fragment analysis of the amplified DNAs revealed 12 restriction patterns for group A viruses and 7 restriction patterns for group B viruses, while the monoclonal antibody reaction patterns revealed seven patterns for group A viruses and 3 patterns for group B viruses. Most viruses with the same monoclonal antibody reaction patterns had different restriction patterns, and some viruses with the same restriction patterns had different monoclonal antibody reaction patterns. Thus, the results of the PCR assay concurred with the monoclonal antibody reaction pattern analysis for group classification of RS viruses, while the restriction fragment analysis identified greater diversity within groups than was seen with the monoclonal antibody analysis. Images PMID:8099086

  14. A Cysteine Zipper Stabilizes a Pre-Fusion F Glycoprotein Vaccine for Respiratory Syncytial Virus.

    PubMed

    Stewart-Jones, Guillaume B E; Thomas, Paul V; Chen, Man; Druz, Aliaksandr; Joyce, M Gordon; Kong, Wing-Pui; Sastry, Mallika; Soto, Cinque; Yang, Yongping; Zhang, Baoshan; Chen, Lei; Chuang, Gwo-Yu; Georgiev, Ivelin S; McLellan, Jason S; Srivatsan, Sanjay; Zhou, Tongqing; Baxa, Ulrich; Mascola, John R; Graham, Barney S; Kwong, Peter D

    2015-01-01

    Recombinant subunit vaccines should contain minimal non-pathogen motifs to reduce potential off-target reactivity. We recently developed a vaccine antigen against respiratory syncytial virus (RSV), which comprised the fusion (F) glycoprotein stabilized in its pre-fusion trimeric conformation by "DS-Cav1" mutations and by an appended C-terminal trimerization motif or "foldon" from T4-bacteriophage fibritin. Here we investigate the creation of a cysteine zipper to allow for the removal of the phage foldon, while maintaining the immunogenicity of the parent DS-Cav1+foldon antigen. Constructs without foldon yielded RSV F monomers, and enzymatic removal of the phage foldon from pre-fusion F trimers resulted in their dissociation into monomers. Because the native C terminus of the pre-fusion RSV F ectodomain encompasses a viral trimeric coiled-coil, we explored whether introduction of cysteine residues capable of forming inter-protomer disulfides might allow for stable trimers. Structural modeling indicated the introduced cysteines to form disulfide "rings", with each ring comprising a different set of inward facing residues of the coiled-coil. Three sets of rings could be placed within the native RSV F coiled-coil, and additional rings could be added by duplicating portions of the coiled-coil. High levels of neutralizing activity in mice, equivalent to that of the parent DS-Cav1+foldon antigen, were elicited by a 4-ring stabilized RSV F trimer with no foldon. Structure-based alteration of a viral coiled-coil to create a cysteine zipper thus allows a phage trimerization motif to be removed from a candidate vaccine antigen. PMID:26098893

  15. Gene Sequence Variability of the Three Surface Proteins of Human Respiratory Syncytial Virus (HRSV) in Texas

    PubMed Central

    Tapia, Lorena I.; Shaw, Chad A.; Aideyan, Letisha O.; Jewell, Alan M.; Dawson, Brian C.; Haq, Taha R.; Piedra, Pedro A.

    2014-01-01

    Human respiratory syncytial virus (HRSV) has three surface glycoproteins: small hydrophobic (SH), attachment (G) and fusion (F), encoded by three consecutive genes (SH-G-F). A 270-nt fragment of the G gene is used to genotype HRSV isolates. This study genotyped and investigated the variability of the gene and amino acid sequences of the three surface proteins of HRSV strains collected from 1987 to 2005 from one center. Sixty original clinical isolates and 5 prototype strains were analyzed. Sequences containing SH, F and G genes were generated, and multiple alignments and phylogenetic trees were analyzed. Genetic variability by protein domains comparing virus genotypes was assessed. Complete sequences of the SH-G-F genes were obtained for all 65 samples: HRSV-A = 35; HRSV-B = 30. In group A strains, genotypes GA5 and GA2 were predominant. For HRSV-B strains, the genotype GB4 was predominant from 1992 to 1994 and only genotype BA viruses were detected in 2004–2005. Different genetic variability at nucleotide level was detected between the genes, with G gene being the most variable and the highest variability detected in the 270-nt G fragment that is frequently used to genotype the virus. High variability (>10%) was also detected in the signal peptide and transmembrane domains of the F gene of HRSV A strains. Variability among the HRSV strains resulting in non-synonymous changes was detected in hypervariable domains of G protein, the signal peptide of the F protein, a not previously defined domain in the F protein, and the antigenic site Ø in the pre-fusion F. Divergent trends were observed between HRSV -A and -B groups for some functional domains. A diverse population of HRSV -A and -B genotypes circulated in Houston during an 18 year period. We hypothesize that diverse sequence variation of the surface protein genes provide HRSV strains a survival advantage in a partially immune-protected community. PMID:24625544

  16. Multiple CD4+ T cell subsets produce immunomodulatory interleukin-10 during respiratory syncytial virus infection

    PubMed Central

    Weiss, Kayla A.; Christiaansen, Allison F.; Fulton, Ross B.; Meyerholz, David K.; Varga, Steven M.

    2011-01-01

    The host immune response is believed to contribute to the severity of pulmonary disease induced by acute respiratory syncytial virus (RSV) infection. Because RSV-induced pulmonary disease is associated with immunopathology, we evaluated the role of IL-10 in modulating the RSV-specific immune response. We found that IL-10 protein levels in the lung were increased following acute RSV infection with maximum production corresponding to the peak of the virus-specific T cell response. The majority of IL-10 producing cells in the lung during acute RSV infection were CD4+ T cells. The IL-10-producing CD4+ T cells included Foxp3+Tregs, Foxp3− CD4+ T cells that co-produce IFN-γ, and Foxp3− CD4+ T cells that do not co-produce IFN-γ. RSV infection of IL-10-deficient mice resulted in more severe disease, as measured by increased weight loss and airway resistance, as compared to control mice. We also observed an increase in the magnitude of the RSV-induced CD8+ and CD4+ T cell response that correlated with increased disease severity in the absence of IL-10 or following IL-10R blockade. Interestingly, IL-10R blockade during acute RSV infection altered CD4+ T cell subset distribution, resulting in a significant increase in IL-17A-producing CD4+ T cells and a concomitant decrease in Foxp3+ regulatory T cells. These results demonstrate that IL-10 plays a critical role in modulating the adaptive immune response to RSV by limiting T-cell-mediated pulmonary inflammation and injury. PMID:21844390

  17. Ozone effect on respiratory syncytial virus infectivity and cytokine production by human alveolar macrophages

    SciTech Connect

    Soukup, J.; Koren, H.S.; Becker, S.

    1993-01-01

    The study was performed to evaluate the effect of ozone (O3) exposure at 1 ppm for 2 hr on the susceptibility/resistance of adult human alveolar macrophages (AM) to infection with respiratory syncytial virus (RSV) in vitro and on RSV-induced cytokine production by the AM. AM were first exposed to O3 or to filtered air and then infected with RSV at multiplicities of infection (m.o.i.) of 0.1 1.0 and 10. The percentage RSV-infected AM and the amount of infectious virus released by the cells were determined at Days 2 and 4 after infection. Interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF) levels in the supernatants were determined on Day 2. No difference in the percentage infected AM or in the amount of infectious RSV produced was found between control and O3-exposed cultures. However, O3-exposed AM infected with RSV at m.o.i. 1 produced less IL-1 in response to RSV infection than control AM:63.6 pg/ml compared with 98.5 pg/ml. No difference in IL-1 was seen with m.o.i. 10. IL-6 levels were also decreased, but only after infection with m.o.i. 0.1. At this level of infection 830 pg/ml was produced by control AM as compared to 468.2 pg/ml by O3-exposed AM. TNF production was unaffected by O3 at all multiplicities of infection. (Copyright (c) 1993 by Academic Press, Inc.)

  18. The Human Respiratory Syncytial Virus Matrix Protein Is Required for Maturation of Viral Filaments

    PubMed Central

    Mitra, Ruchira; Baviskar, Pradyumna; Duncan-Decocq, Rebecca R.; Patel, Darshna

    2012-01-01

    An experimental system was developed to generate infectious human respiratory syncytial virus (HRSV) lacking matrix (M) protein expression (M-null virus) from cDNA. The role of the M protein in virus assembly was then examined by infecting HEp-2 and Vero cells with the M-null virus and assessing the impact on infectious virus production and viral protein trafficking. In the absence of M, the production of infectious progeny was strongly impaired. Immunofluorescence (IF) microscopy analysis using antibodies against the nucleoprotein (N), attachment protein (G), and fusion protein (F) failed to detect the characteristic virus-induced cell surface filaments, which are believed to represent infectious virions. In addition, a large proportion of the N protein was detected in viral replication factories termed inclusion bodies (IBs). High-resolution analysis of the surface of M-null virus-infected cells by field emission scanning electron microscopy (SEM) revealed the presence of large areas with densely packed, uniformly short filaments. Although unusually short, these filaments were otherwise similar to those induced by an M-containing control virus, including the presence of the viral G and F proteins. The abundance of the short, stunted filaments in the absence of M indicates that M is not required for the initial stages of filament formation but plays an important role in the maturation or elongation of these structures. In addition, the absence of mature viral filaments and the simultaneous increase in the level of the N protein within IBs suggest that the M protein is involved in the transport of viral ribonucleoprotein (RNP) complexes from cytoplasmic IBs to sites of budding. PMID:22318136

  19. Immunomodulation with IL-4R alpha antisense oligonucleotide prevents respiratory syncytial virus-mediated pulmonary disease.

    PubMed

    Ripple, Michael J; You, Dahui; Honnegowda, Srinivasa; Giaimo, Joseph D; Sewell, Andrew B; Becnel, David M; Cormier, Stephania A

    2010-10-15

    Respiratory syncytial virus (RSV) causes significant morbidity and mortality in infants worldwide. Severe RSV infections in infants cause bronchiolitis, wheeze, and/or cough and significantly increase the risk for developing asthma. RSV pathogenesis is thought to be due to a Th2-type immune response initiated in response to RSV infection, specifically in the infant. Using a neonatal mouse system as an appropriate model for human infants, we sought to determine whether local inhibition of IL-4Rα expression during primary RSV infection in the neonate would prevent Th2-skewed responses to secondary RSV infection and improve long-term pulmonary function. To reduce IL-4Rα expression, antisense oligonucleotides (ASOs) specific for IL-4Rα were administered intranasally to neonatal mice at the time of primary infection. Mice were initially infected with RSV at 1 wk of age and were reinfected at 6 wk of age. Administration of IL-4Rα ASOs during primary RSV infection in neonatal mice abolished the pulmonary dysfunction normally observed following reinfection in the adult. This ablation of pulmonary dysfunction correlated with a persistent rebalancing of the Th cell compartment with decreased Th2 responses (i.e., reduced goblet cell hyperplasia, Th2 cells, and cytokine secretion) and increased Th1 responses (i.e., elevated Th1 cell numbers and type I Abs and cytokines). Our data support our hypothesis that a reduction in the Th2 immune response during primary infection in neonates prevents Th2-mediated pulmonary pathology initially and upon reinfection and further suggest that vaccine strategies incorporating IL-4Rα ASOs may be of significant benefit to infants.

  20. A Cysteine Zipper Stabilizes a Pre-Fusion F Glycoprotein Vaccine for Respiratory Syncytial Virus.

    PubMed

    Stewart-Jones, Guillaume B E; Thomas, Paul V; Chen, Man; Druz, Aliaksandr; Joyce, M Gordon; Kong, Wing-Pui; Sastry, Mallika; Soto, Cinque; Yang, Yongping; Zhang, Baoshan; Chen, Lei; Chuang, Gwo-Yu; Georgiev, Ivelin S; McLellan, Jason S; Srivatsan, Sanjay; Zhou, Tongqing; Baxa, Ulrich; Mascola, John R; Graham, Barney S; Kwong, Peter D

    2015-01-01

    Recombinant subunit vaccines should contain minimal non-pathogen motifs to reduce potential off-target reactivity. We recently developed a vaccine antigen against respiratory syncytial virus (RSV), which comprised the fusion (F) glycoprotein stabilized in its pre-fusion trimeric conformation by "DS-Cav1" mutations and by an appended C-terminal trimerization motif or "foldon" from T4-bacteriophage fibritin. Here we investigate the creation of a cysteine zipper to allow for the removal of the phage foldon, while maintaining the immunogenicity of the parent DS-Cav1+foldon antigen. Constructs without foldon yielded RSV F monomers, and enzymatic removal of the phage foldon from pre-fusion F trimers resulted in their dissociation into monomers. Because the native C terminus of the pre-fusion RSV F ectodomain encompasses a viral trimeric coiled-coil, we explored whether introduction of cysteine residues capable of forming inter-protomer disulfides might allow for stable trimers. Structural modeling indicated the introduced cysteines to form disulfide "rings", with each ring comprising a different set of inward facing residues of the coiled-coil. Three sets of rings could be placed within the native RSV F coiled-coil, and additional rings could be added by duplicating portions of the coiled-coil. High levels of neutralizing activity in mice, equivalent to that of the parent DS-Cav1+foldon antigen, were elicited by a 4-ring stabilized RSV F trimer with no foldon. Structure-based alteration of a viral coiled-coil to create a cysteine zipper thus allows a phage trimerization motif to be removed from a candidate vaccine antigen.

  1. A Cysteine Zipper Stabilizes a Pre-Fusion F Glycoprotein Vaccine for Respiratory Syncytial Virus

    PubMed Central

    Stewart-Jones, Guillaume B. E.; Thomas, Paul V.; Chen, Lei; Chuang, Gwo-Yu; Georgiev, Ivelin S.; McLellan, Jason S.; Srivatsan, Sanjay; Zhou, Tongqing; Baxa, Ulrich; Mascola, John R.; Graham, Barney S.; Kwong, Peter D.

    2015-01-01

    Recombinant subunit vaccines should contain minimal non-pathogen motifs to reduce potential off-target reactivity. We recently developed a vaccine antigen against respiratory syncytial virus (RSV), which comprised the fusion (F) glycoprotein stabilized in its pre-fusion trimeric conformation by “DS-Cav1” mutations and by an appended C-terminal trimerization motif or “foldon” from T4-bacteriophage fibritin. Here we investigate the creation of a cysteine zipper to allow for the removal of the phage foldon, while maintaining the immunogenicity of the parent DS-Cav1+foldon antigen. Constructs without foldon yielded RSV F monomers, and enzymatic removal of the phage foldon from pre-fusion F trimers resulted in their dissociation into monomers. Because the native C terminus of the pre-fusion RSV F ectodomain encompasses a viral trimeric coiled-coil, we explored whether introduction of cysteine residues capable of forming inter-protomer disulfides might allow for stable trimers. Structural modeling indicated the introduced cysteines to form disulfide “rings”, with each ring comprising a different set of inward facing residues of the coiled-coil. Three sets of rings could be placed within the native RSV F coiled-coil, and additional rings could be added by duplicating portions of the coiled-coil. High levels of neutralizing activity in mice, equivalent to that of the parent DS-Cav1+foldon antigen, were elicited by a 4-ring stabilized RSV F trimer with no foldon. Structure-based alteration of a viral coiled-coil to create a cysteine zipper thus allows a phage trimerization motif to be removed from a candidate vaccine antigen. PMID:26098893

  2. Respiratory syncytial virus infection in children admitted to hospital but ventilated mechanically for other reasons.

    PubMed

    von Renesse, Anja; Schildgen, Oliver; Klinkenberg, Dennis; Müller, Andreas; von Moers, Arpad; Simon, Arne

    2009-01-01

    One thousand five hundred sixty-eight RSV infections were documented prospectively in 1,541 pediatric patients. Of these, 20 (1.3%) had acquired the RSV infection while treated by mechanical ventilation for reasons other than the actual RSV infection (group ventilated mechanically). The clinical characteristics of children who were infected with respiratory syncytial virus (RSV) infection while ventilated mechanically for other reasons are described and compared with a matched control group. Sixty percent of the group ventilated mechanically had at least one additional risk factor for a severe course of infection (prematurity 50%, chronic lung disease 20%, congenital heart disease 35%, immunodeficiency 20%). The median age at diagnosis in the group ventilated mechanically was 4.2 months. The matched pairs analysis (group ventilated mechanically vs. control group) revealed a higher proportion of patients with hypoxemia and apnoea in the group ventilated mechanically; more patients in the control group showed symptoms of airway obstruction (wheezing). At least one chest radiography was performed in 95% of the patients (n = 19) in the group ventilated mechanically versus 45% (n = 9) in the control group (P = 0.001). The frequency of pneumonia was 40% in the group ventilated mechanically and 20% in the control group. Despite existing consensus recommendations, only two patients (10%) of the group ventilated mechanically had received palivizumab previously. Significantly more patients in the group ventilated mechanically received antibiotic treatment (85% vs. 45%, P = 0.008), and attributable mortality was higher in the group ventilated mechanically (15% [n = 3] vs. 0% in the control group, P = 0.231). Children treated by long term mechanical ventilation may acquire RSV infection by transmission by droplets or caregivers and face an increased risk of a severe course of RSV infection. The low rate of immunoprophylaxis in this particular risk group should be improved.

  3. The role of dendritic cells in innate and adaptive immunity to respiratory syncytial virus, and implications for vaccine development.

    PubMed

    Garg, Ravendra; Shrivastava, Pratima; van Drunen Littel-van den Hurk, Sylvia

    2012-12-01

    Respiratory syncytial virus (RSV) is a common human pathogen that causes cold-like symptoms in most healthy adults and children. However, RSV often moves into the lower respiratory tract in infants and young children predisposed to respiratory illness, making it the most common cause of pediatric broncheolitis and pneumonia. The development of an appropriate balanced immune response is critical for recovery from RSV, while an unbalanced and/or excessively vigorous response may lead to immunopathogenesis. Different dendritic cell (DC) subsets influence the magnitude and quality of the host response to RSV infection, with myeloid DCs mediating and plasmacytoid DCs modulating immunopathology. Furthermore, stimulation of DCs through Toll-like receptors is essential for induction of protective immunity to RSV. These characteristics have implications for the rational design of a RSV vaccine.

  4. Induction of protective effector immunity to prevent pathogenesis caused by the respiratory syncytial virus. Implications on therapy and vaccine design

    PubMed Central

    Espinoza, Janyra A; Bueno, Susan M; Riedel, Claudia A; Kalergis, Alexis M

    2014-01-01

    Human respiratory syncytial virus (hRSV) is the leading cause of respiratory illness in infants and young children around the globe. This pathogen, which was discovered in 1956, continues to cause a huge number of hospitalizations due to respiratory disease and it is considered a health and economic burden worldwide, especially in developing countries. The immune response elicited by hRSV infection leads to lung and systemic inflammation, which results in lung damage but is not efficient at preventing viral replication. Indeed, natural hRSV infection induces a poor immune memory that allows recurrent infections. Here, we review the most recent knowledge about the lifecycle of hRSV, the immune response elicited by this virus and the subsequent pathology induced in response to infection in the airways. Novel findings about the alterations that this virus causes in the central nervous system and potential therapies and vaccines designed to treat or prevent hRSV infection are discussed. PMID:24801878

  5. Novel diversity-oriented synthesis-derived respiratory syncytial virus inhibitors identified via a high throughput replicon-based screen.

    PubMed

    Duvall, Jeremy R; VerPlank, Lynn; Ludeke, Barbara; McLeod, Sarah M; Lee, Maurice D; Vishwanathan, Karthick; Mulrooney, Carol A; Le Quement, Sebastian; Yu, Qin; Palmer, Michelle A; Fleming, Paul; Fearns, Rachel; Foley, Michael A; Scherer, Christina A

    2016-07-01

    Respiratory syncytial virus (RSV) infections affect millions of children and adults every year. Despite the significant disease burden, there are currently no safe and effective vaccines or therapeutics. We employed a replicon-based high throughput screen combined with live-virus triaging assays to identify three novel diversity-oriented synthesis-derived scaffolds with activity against RSV. One of these small molecules is shown to target the RSV polymerase (L protein) to inhibit viral replication and transcription; the mechanisms of action of the other small molecules are currently unknown. The compounds described herein may provide attractive inhibitors for lead optimization campaigns. PMID:27059228

  6. Successive Respiratory Syncytial Virus Epidemics in Local Populations Arise from Multiple Variant Introductions, Providing Insights into Virus Persistence

    PubMed Central

    Otieno, James R.; Ngama, Mwanajuma; Mwihuri, Alexander G.; Medley, Graham F.; Cane, Patricia A.; Nokes, D. James

    2015-01-01

    ABSTRACT Respiratory syncytial virus (RSV) is a global respiratory pathogen of humans, with infection occurring characteristically as recurrent seasonal epidemics. Unlike influenza viruses, little attention has been paid to the mechanism underlying worldwide spread and persistence of RSV and how this may be discerned through an improved understanding of the introduction and persistence of RSV in local communities. We analyzed 651 attachment (G) glycoprotein nucleotide sequences of RSV B collected over 11 epidemics (2002 to 2012) in Kilifi, Kenya, and contemporaneous data collected elsewhere in Kenya and 18 other countries worldwide (2002 to 2012). Based on phylogeny, genetic distance and clustering patterns, we set out pragmatic criteria to classify local viruses into distinct genotypes and variants, identifying those newly introduced and those locally persisting. Three genotypes were identified in the Kilifi data set: BA (n = 500), SAB1 (n = 148), and SAB4 (n = 3). Recurrent RSV epidemics in the local population were composed of numerous genetic variants, most of which have been newly introduced rather than persisting in the location from season to season. Global comparison revealed that (i) most Kilifi variants do not cluster closely with strains from outside Kenya, (ii) some Kilifi variants were closely related to those observed outside Kenya (mostly Western Europe), and (iii) many variants were circulating elsewhere but were never detected in Kilifi. These results are consistent with the hypothesis that year-to-year presence of RSV at the local level (i.e., Kilifi) is achieved primarily, but not exclusively, through introductions from a pool of variants that are geographically restricted (i.e., to Kenya or to the region) rather than global. IMPORTANCE The mechanism by which RSV persists and reinvades local populations is poorly understood. We investigated this by studying the temporal patterns of RSV variants in a rural setting in tropical Africa and comparing

  7. Sequencing and Analysis of Globally Obtained Human Respiratory Syncytial Virus A and B Genomes

    PubMed Central

    Bose, Michael E.; He, Jie; Shrivastava, Susmita; Nelson, Martha I.; Bera, Jayati; Halpin, Rebecca A.; Town, Christopher D.; Lorenzi, Hernan A.; Noyola, Daniel E.; Falcone, Valeria; Gerna, Giuseppe; De Beenhouwer, Hans; Videla, Cristina; Kok, Tuckweng; Venter, Marietjie; Williams, John V.; Henrickson, Kelly J.

    2015-01-01

    Background Human respiratory syncytial virus (RSV) is the leading cause of respiratory tract infections in children globally, with nearly all children experiencing at least one infection by the age of two. Partial sequencing of the attachment glycoprotein gene is conducted routinely for genotyping, but relatively few whole genome sequences are available for RSV. The goal of our study was to sequence the genomes of RSV strains collected from multiple countries to further understand the global diversity of RSV at a whole-genome level. Methods We collected RSV samples and isolates from Mexico, Argentina, Belgium, Italy, Germany, Australia, South Africa, and the USA from the years 1998-2010. Both Sanger and next-generation sequencing with the Illumina and 454 platforms were used to sequence the whole genomes of RSV A and B. Phylogenetic analyses were performed using the Bayesian and maximum likelihood methods of phylogenetic inference. Results We sequenced the genomes of 34 RSVA and 23 RSVB viruses. Phylogenetic analysis showed that the RSVA genome evolves at an estimated rate of 6.72 × 10-4 substitutions/site/year (95% HPD 5.61 × 10-4 to 7.6 × 10-4) and for RSVB the evolutionary rate was 7.69 × 10-4 substitutions/site/year (95% HPD 6.81 × 10-4 to 8.62 × 10-4). We found multiple clades co-circulating globally for both RSV A and B. The predominant clades were GA2 and GA5 for RSVA and BA for RSVB. Conclusions Our analyses showed that RSV circulates on a global scale with the same predominant clades of viruses being found in countries around the world. However, the distribution of clades can change rapidly as new strains emerge. We did not observe a strong spatial structure in our trees, with the same three main clades of RSV co-circulating globally, suggesting that the evolution of RSV is not strongly regionalized. PMID:25793751

  8. Structure-Based Design of Head-Only Fusion Glycoprotein Immunogens for Respiratory Syncytial Virus.

    PubMed

    Boyington, Jeffrey C; Joyce, M Gordon; Sastry, Mallika; Stewart-Jones, Guillaume B E; Chen, Man; Kong, Wing-Pui; Ngwuta, Joan O; Thomas, Paul V; Tsybovsky, Yaroslav; Yang, Yongping; Zhang, Baoshan; Chen, Lei; Druz, Aliaksandr; Georgiev, Ivelin S; Ko, Kiyoon; Zhou, Tongqing; Mascola, John R; Graham, Barney S; Kwong, Peter D

    2016-01-01

    Respiratory syncytial virus (RSV) is a significant cause of severe respiratory illness worldwide, particularly in infants, young children, and the elderly. Although no licensed vaccine is currently available, an engineered version of the metastable RSV fusion (F) surface glycoprotein-stabilized in the pre-fusion (pre-F) conformation by "DS-Cav1" mutations-elicits high titer RSV-neutralizing responses. Moreover, pre-F-specific antibodies, often against the neutralization-sensitive antigenic site Ø in the membrane-distal head region of trimeric F glycoprotein, comprise a substantial portion of the human response to natural RSV infection. To focus the vaccine-elicited response to antigenic site Ø, we designed a series of RSV F immunogens that comprised the membrane-distal head of the F glycoprotein in its pre-F conformation. These "head-only" immunogens formed monomers, dimers, and trimers. Antigenic analysis revealed that a majority of the 70 engineered head-only immunogens displayed reactivity to site Ø-targeting antibodies, which was similar to that of the parent RSV F DS-Cav1 trimers, often with increased thermostability. We evaluated four of these head-only immunogens in detail, probing their recognition by antibodies, their physical stability, structure, and immunogenicity. When tested in naïve mice, a head-only trimer, half the size of the parent RSV F trimer, induced RSV titers, which were statistically comparable to those induced by DS-Cav1. When used to boost DS-Cav1-primed mice, two head-only RSV F immunogens, a dimer and a trimer, boosted RSV-neutralizing titers to levels that were comparable to those boosted by DS-Cav1, although with higher site Ø-directed responses. Our results provide proof-of-concept for the ability of the smaller head-only RSV F immunogens to focus the vaccine-elicited response to antigenic site Ø. Decent primary immunogenicity, enhanced physical stability, potential ease of manufacture, and potent immunogenicity upon boosting

  9. Structure-Based Design of Head-Only Fusion Glycoprotein Immunogens for Respiratory Syncytial Virus.

    PubMed

    Boyington, Jeffrey C; Joyce, M Gordon; Sastry, Mallika; Stewart-Jones, Guillaume B E; Chen, Man; Kong, Wing-Pui; Ngwuta, Joan O; Thomas, Paul V; Tsybovsky, Yaroslav; Yang, Yongping; Zhang, Baoshan; Chen, Lei; Druz, Aliaksandr; Georgiev, Ivelin S; Ko, Kiyoon; Zhou, Tongqing; Mascola, John R; Graham, Barney S; Kwong, Peter D

    2016-01-01

    Respiratory syncytial virus (RSV) is a significant cause of severe respiratory illness worldwide, particularly in infants, young children, and the elderly. Although no licensed vaccine is currently available, an engineered version of the metastable RSV fusion (F) surface glycoprotein-stabilized in the pre-fusion (pre-F) conformation by "DS-Cav1" mutations-elicits high titer RSV-neutralizing responses. Moreover, pre-F-specific antibodies, often against the neutralization-sensitive antigenic site Ø in the membrane-distal head region of trimeric F glycoprotein, comprise a substantial portion of the human response to natural RSV infection. To focus the vaccine-elicited response to antigenic site Ø, we designed a series of RSV F immunogens that comprised the membrane-distal head of the F glycoprotein in its pre-F conformation. These "head-only" immunogens formed monomers, dimers, and trimers. Antigenic analysis revealed that a majority of the 70 engineered head-only immunogens displayed reactivity to site Ø-targeting antibodies, which was similar to that of the parent RSV F DS-Cav1 trimers, often with increased thermostability. We evaluated four of these head-only immunogens in detail, probing their recognition by antibodies, their physical stability, structure, and immunogenicity. When tested in naïve mice, a head-only trimer, half the size of the parent RSV F trimer, induced RSV titers, which were statistically comparable to those induced by DS-Cav1. When used to boost DS-Cav1-primed mice, two head-only RSV F immunogens, a dimer and a trimer, boosted RSV-neutralizing titers to levels that were comparable to those boosted by DS-Cav1, although with higher site Ø-directed responses. Our results provide proof-of-concept for the ability of the smaller head-only RSV F immunogens to focus the vaccine-elicited response to antigenic site Ø. Decent primary immunogenicity, enhanced physical stability, potential ease of manufacture, and potent immunogenicity upon boosting

  10. Elevated temperature triggers human respiratory syncytial virus F protein six-helix bundle formation

    SciTech Connect

    Yunus, Abdul S.; Jackson, Trent P.; Crisafi, Katherine; Burimski, Irina; Kilgore, Nicole R.; Zoumplis, Dorian; Allaway, Graham P.; Wild, Carl T.; Salzwedel, Karl

    2010-01-20

    Human respiratory syncytial virus (RSV) is a major cause of severe lower respiratory tract infection in infants, immunocompromised patients, and the elderly. The RSV fusion (F) protein mediates fusion of the viral envelope with the target cell membrane during virus entry and is a primary target for antiviral drug and vaccine development. The F protein contains two heptad repeat regions, HR1 and HR2. Peptides corresponding to these regions form a six-helix bundle structure that is thought to play a critical role in membrane fusion. However, characterization of six-helix bundle formation in native RSV F protein has been hindered by the fact that a trigger for F protein conformational change has yet to be identified. Here we demonstrate that RSV F protein on the surface of infected cells undergoes a conformational change following exposure to elevated temperature, resulting in the formation of the six-helix bundle structure. We first generated and characterized six-helix bundle-specific antibodies raised against recombinant peptides modeling the RSV F protein six-helix bundle structure. We then used these antibodies as probes to monitor RSV F protein six-helix bundle formation in response to a diverse array of potential triggers of conformational changes. We found that exposure of 'membrane-anchored' RSV F protein to elevated temperature (45-55 deg. C) was sufficient to trigger six-helix bundle formation. Antibody binding to the six-helix bundle conformation was detected by both flow cytometry and cell-surface immunoprecipitation of the RSV F protein. None of the other treatments, including interaction with a number of potential receptors, resulted in significant binding by six-helix bundle-specific antibodies. We conclude that native, untriggered RSV F protein exists in a metastable state that can be converted in vitro to the more stable, fusogenic six-helix bundle conformation by an increase in thermal energy. These findings help to better define the mechanism of

  11. Structure-Based Design of Head-Only Fusion Glycoprotein Immunogens for Respiratory Syncytial Virus

    PubMed Central

    Boyington, Jeffrey C.; Chen, Man; Kong, Wing-Pui; Ngwuta, Joan O.; Thomas, Paul V.; Tsybovsky, Yaroslav; Yang, Yongping; Zhang, Baoshan; Chen, Lei; Druz, Aliaksandr; Georgiev, Ivelin S.; Ko, Kiyoon; Zhou, Tongqing; Mascola, John R.; Graham, Barney S.; Kwong, Peter D.

    2016-01-01

    Respiratory syncytial virus (RSV) is a significant cause of severe respiratory illness worldwide, particularly in infants, young children, and the elderly. Although no licensed vaccine is currently available, an engineered version of the metastable RSV fusion (F) surface glycoprotein—stabilized in the pre-fusion (pre-F) conformation by “DS-Cav1” mutations—elicits high titer RSV-neutralizing responses. Moreover, pre-F-specific antibodies, often against the neutralization-sensitive antigenic site Ø in the membrane-distal head region of trimeric F glycoprotein, comprise a substantial portion of the human response to natural RSV infection. To focus the vaccine-elicited response to antigenic site Ø, we designed a series of RSV F immunogens that comprised the membrane-distal head of the F glycoprotein in its pre-F conformation. These “head-only” immunogens formed monomers, dimers, and trimers. Antigenic analysis revealed that a majority of the 70 engineered head-only immunogens displayed reactivity to site Ø-targeting antibodies, which was similar to that of the parent RSV F DS-Cav1 trimers, often with increased thermostability. We evaluated four of these head-only immunogens in detail, probing their recognition by antibodies, their physical stability, structure, and immunogenicity. When tested in naïve mice, a head-only trimer, half the size of the parent RSV F trimer, induced RSV titers, which were statistically comparable to those induced by DS-Cav1. When used to boost DS-Cav1-primed mice, two head-only RSV F immunogens, a dimer and a trimer, boosted RSV-neutralizing titers to levels that were comparable to those boosted by DS-Cav1, although with higher site Ø-directed responses. Our results provide proof-of-concept for the ability of the smaller head-only RSV F immunogens to focus the vaccine-elicited response to antigenic site Ø. Decent primary immunogenicity, enhanced physical stability, potential ease of manufacture, and potent immunogenicity

  12. Measles–mumps–rubella vaccination and respiratory syncytial virus-associated hospital contact

    PubMed Central

    Sørup, Signe; Benn, Christine Stabell; Stensballe, Lone Graff; Aaby, Peter; Ravn, Henrik

    2015-01-01

    Background The live measles vaccine has been associated with lower non-measles mortality and admissions in low-income countries. The live measles–mumps–rubella vaccine has also been associated with lower rate of admissions with any type of infection in Danish children; the association was strongest for admissions with lower respiratory infections. Objective To examine whether measles, mumps, and rubella (MMR) vaccination was associated with reduced rate of hospital contact related to respiratory syncytial virus (RSV) in a high-income country. Methods Nationwide cohort study of laboratory-confirmed RSV hospital contacts at age 14–23 months in all children born in Denmark 1997–2002 who had already received the vaccine against diphtheria, tetanus, pertussis (acellular), polio, and Haemophilus influenzae type b (DTaP-IPV-Hib) at the recommended ages of 3, 5, and 12 months. Results The study included 888 RSV hospital contacts in 128,588 person years of follow up (rate 6.8/1000 person years). Having MMR as the most recent vaccine was associated with a reduced rate of RSV hospital contacts compared with having DTaP-IPV-Hib as the most recent vaccine (Incidence rate ratio (IRR), 0.75; 95% confidence interval (CI), 0.63–0.89). After adjustment for potential confounders including exact age in days the IRR was 0.78 (95% CI, 0.66–0.93). The adjusted IRR was 0.74 (95% CI, 0.60–0.92) in males and 0.84 (95% CI, 0.66–1.06) in females (P Interaction, 0.42). There was no association in the first month after MMR vaccination (adjusted IRR, 0.97; 95% CI, 0.76–1.24) but the adjusted IRR was 0.70 (95% CI, 0.58–0.85) from one month after MMR vaccination. Conclusions MMR vaccination was associated with reduced rate of hospital contacts related to laboratory-confirmed RSV infection. Further research on the association between MMR vaccination and other unrelated pathogens are warranted. PMID:25446818

  13. Comparison of Cepheid Xpert Flu/RSV XC and BioFire FilmArray for Detection of Influenza A, Influenza B, and Respiratory Syncytial Virus.

    PubMed

    Wahrenbrock, Mark G; Matushek, Scott; Boonlayangoor, Sue; Tesic, Vera; Beavis, Kathleen G; Charnot-Katsikas, Angella

    2016-07-01

    The Xpert Flu/RSV XC was compared to the FilmArray respiratory panel for detection of influenza (Flu) A, Flu B, and respiratory syncytial virus (RSV), using 128 nasopharyngeal swabs. Positive agreements were 100% for Flu A and RSV and 92.3% for Flu B. The Xpert may be useful in clinical situations when extensive testing is not required and may serve an important role in laboratories already performing broader respiratory panel testing.

  14. Respiratory syncytial virus: the virus, the disease and the immune response.

    PubMed

    Ogra, Pearay L

    2004-01-01

    and respiratory mucosa. Both CD(4)- and CD(8)-specific as well as Th(1)- and Th(2)-cell specific immune responses have been observed during human infection. In addition, proinflammatory as well as immunoregulatory cytokines and chemokines are induced in the respiratory tract after natural and induced (in vitro) infection. Significant progress has been made in understanding the role of Th(1) vs. Th(2), IgE, viral induced cytokines and chemokines in the mechanisms of pathogenesis of the disease, development of wheezing and in the prevention and treatment of the infection and its sequelae. Respiratory syncytial virus (RSV) is one of the commonest human viral infections, and virtually every child is infected by the third birthday. Because of its restricted mucosal immunopathology, and frequent association with bronchial hyperreactivity and development of wheezing, RSV has served as an important model to investigate mechanisms of mucosal immune responses and development of mucosal disease following infection. The importance of RSV in bronchopulmonary disease and development of bronchial hyperreactivity has been the focus of several recent symposia [Kimpen JL, Simoes EAF. Am J Respir Crit Care Med 2001; 163:S1-S6]. This brief report will only summarise, based on selected references, the historical landmarks of its discovery and current understanding of the mechanisms of immunity, and their possible role in the pathogenesis of bronchopulmonary disease.

  15. Incidence of Medically Attended Respiratory Syncytial Virus and Influenza Illnesses in Children 6–59 Months Old During Four Seasons

    PubMed Central

    Simpson, Melissa D.; Kieke, Burney A.; Sundaram, Maria E.; McClure, David L.; Meece, Jennifer K.; Sifakis, Frangiscos; Gasser, Robert A.; Belongia, Edward A.

    2016-01-01

    Background. Respiratory syncytial virus (RSV) and influenza are significant causes of seasonal respiratory illness in children. The incidence of influenza and RSV hospitalization is well documented, but the incidence of medically attended, laboratory-confirmed illness has not been assessed in a well defined community cohort. Methods. Children aged 6–59 months with medically attended acute respiratory illness were prospectively enrolled during the 2006–2007 through 2009–2010 influenza seasons in a Wisconsin community cohort. Nasal swabs were tested for RSV and influenza by multiplex reverse-transcription polymerase chain reaction. The population incidence of medically attended RSV and influenza was estimated separately and standardized to weeks 40 through 18 of each season. Results. The cohort included 2800–3073 children each season. There were 2384 children enrolled with acute respiratory illness; 627 (26%) were positive for RSV and 314 (13%) for influenza. The mean age was 28 months (standard deviation [SD] = 15) for RSV-positive and 38 months (SD = 16) for influenza-positive children. Seasonal incidence (cases per 10 000) was 1718 (95% confidence interval [CI], 1602–1843) for RSV and 768 (95% CI, 696–848) for influenza. Respiratory syncytial virus incidence was highest among children 6–11 (2927) and 12–23 months old (2377). Influenza incidence was highest (850) in children 24–59 months old. The incidence of RSV was higher than influenza across all seasons and age groups. Conclusions. The incidence of medically attended RSV was highest in children 6–23 months old, and it was consistently higher than influenza. The burden of RSV remains high throughout the first 2 years of life. PMID:27419158

  16. Incidence of Medically Attended Respiratory Syncytial Virus and Influenza Illnesses in Children 6-59 Months Old During Four Seasons.

    PubMed

    Simpson, Melissa D; Kieke, Burney A; Sundaram, Maria E; McClure, David L; Meece, Jennifer K; Sifakis, Frangiscos; Gasser, Robert A; Belongia, Edward A

    2016-04-01

    Background.  Respiratory syncytial virus (RSV) and influenza are significant causes of seasonal respiratory illness in children. The incidence of influenza and RSV hospitalization is well documented, but the incidence of medically attended, laboratory-confirmed illness has not been assessed in a well defined community cohort. Methods.  Children aged 6-59 months with medically attended acute respiratory illness were prospectively enrolled during the 2006-2007 through 2009-2010 influenza seasons in a Wisconsin community cohort. Nasal swabs were tested for RSV and influenza by multiplex reverse-transcription polymerase chain reaction. The population incidence of medically attended RSV and influenza was estimated separately and standardized to weeks 40 through 18 of each season. Results.  The cohort included 2800-3073 children each season. There were 2384 children enrolled with acute respiratory illness; 627 (26%) were positive for RSV and 314 (13%) for influenza. The mean age was 28 months (standard deviation [SD] = 15) for RSV-positive and 38 months (SD = 16) for influenza-positive children. Seasonal incidence (cases per 10 000) was 1718 (95% confidence interval [CI], 1602-1843) for RSV and 768 (95% CI, 696-848) for influenza. Respiratory syncytial virus incidence was highest among children 6-11 (2927) and 12-23 months old (2377). Influenza incidence was highest (850) in children 24-59 months old. The incidence of RSV was higher than influenza across all seasons and age groups. Conclusions.  The incidence of medically attended RSV was highest in children 6-23 months old, and it was consistently higher than influenza. The burden of RSV remains high throughout the first 2 years of life. PMID:27419158

  17. Real-Time Detection of Influenza A, Influenza B, and Respiratory Syncytial Virus A and B in Respiratory Specimens by Use of Nanoparticle Probes ▿ †

    PubMed Central

    Jannetto, Paul J.; Buchan, Blake W.; Vaughan, Kimberly A.; Ledford, Joellen S.; Anderson, Dennis K.; Henley, Donald C.; Quigley, Neil B.; Ledeboer, Nathan A.

    2010-01-01

    Seasonal epidemics of influenza and respiratory syncytial virus are responsible for significant morbidity and mortality worldwide. Infrequently, novel or reemergent strains of influenza A virus have caused rapid, severe global pandemics resulting in millions of fatalities. The ability to efficiently and accurately detect and differentiate respiratory viruses is paramount for effective treatment, infection control, and epidemiological surveillance. We evaluated the ability of two FDA-cleared nucleic acid-based tests, the semiautomated respiratory virus nucleic acid test (VRNAT) and the fully automated respiratory virus nucleic acid test SP (RVNATSP) (Nanosphere Inc., Northbrook, IL) to detect influenza A virus, influenza B virus, and respiratory syncytial virus A and B (RSV A/B) from clinical nasopharyngeal swab specimens. Detection of viral RNA in both tests is based on nucleic acid amplification followed by hybridization to capture probes immobilized on a glass slide. A novel technology utilizing gold nanoparticle-conjugated probes is utilized to detect the presence of captured target DNA. This microarray-based approach to detection has proven to be more sensitive than the traditional culture/direct fluorescent-antibody assay (DFA) method for detecting RSV and influenza viruses in clinical specimens, including the novel 2009 H1N1 strain. Specifically, we report 98.0% sensitivity and 96.5% specificity for the VRNAT compared to culture/DFA. Further, the VRNAT detected virus in an additional 58% of specimens that were culture negative. These data were confirmed using bidirectional sequencing. Evaluation of the fully automated RVNATSP, which is built on the same detection technology as the VRNAT but contains an updated processor enabling complete automation, revealed the two tests to be functionally equivalent. Thus, the RVNATSP is a fully automated sample-to-result test capable of reliable detection of select respiratory viruses directly from clinical specimens in 3

  18. Pneumonia associated with infection with pneumocystis, respiratory syncytial virus, chlamydia, mycoplasma, and cytomegalovirus in children in Papua New Guinea.

    PubMed Central

    Shann, F; Walters, S; Pifer, L L; Graham, D M; Jack, I; Uren, E; Birch, D; Stallman, N D

    1986-01-01

    Paired serum samples were collected from 94 children with pneumonia admitted to Goroka Hospital, Papua New Guinea. All but three of the children were aged 1-24 months. Only nine children were malnourished, with weight for age less than 70% of the Harvard median (three had weight for age less than 60% of the Harvard median). Pneumocystis carinii antigen was detected in the serum of 23 children. Twenty two children had serological evidence of recent infection with respiratory syncytial virus. Five children were probably infected with Chlamydia trachomatis at the time of the study, and there was less convincing serological evidence of current infection in a further 11 children. Five children showed a fourfold rise in antibody to Mycoplasma pneumoniae. Although only one child showed a fourfold rise in antibody to cytomegalovirus, 86 children had this antibody. No child showed a fourfold rise in antibody to Ureaplasma urealyticum or Legionella pneumophila. P carinii, respiratory syncytial virus, C trachomatis, M pneumoniae, and cytomegalovirus may be important causes of pneumonia in children in developing countries. PMID:3002538

  19. Oral efficacy of a respiratory syncytial virus inhibitor in rodent models of infection.

    PubMed

    Cianci, Christopher; Genovesi, Eugene V; Lamb, Lucinda; Medina, Ivette; Yang, Zheng; Zadjura, Lisa; Yang, Hyekyung; D'Arienzo, Celia; Sin, Ny; Yu, Kuo-Long; Combrink, Keith; Li, Zhufang; Colonno, Richard; Meanwell, Nicholas; Clark, Junius; Krystal, Mark

    2004-07-01

    BMS-433771 is a potent inhibitor of respiratory syncytial virus (RSV) replication in vitro. Mechanism of action studies have demonstrated that BMS-433771 halts virus entry through inhibition of F protein-mediated membrane fusion. BMS-433771 also exhibited in vivo efficacy following oral administration in a mouse model of RSV infection (C. Cianci, K. Y. Yu, K. Combrink, N. Sin, B. Pearce, A. Wang, R. Civiello, S. Voss, G. Luo, K. Kadow, E. Genovesi, B. Venables, H. Gulgeze, A. Trehan, J. James, L. Lamb, I. Medina, J. Roach, Z. Yang, L. Zadjura, R. Colonno, J. Clark, N. Meanwell, and M. Krystal, Antimicrob. Agents Chemother. 48:413-422, 2004). In this report, the in vivo efficacy of BMS-433771 against RSV was further examined in the BALB/c mouse and cotton rat host models of infection. By using the Long strain of RSV, prophylactic efficacy via oral dosing was observed in both animal models. A single oral dose, administered 1 h prior to intranasal RSV inoculation, was as effective against infection as a 4-day b.i.d. dosing regimen in which the first oral dose was given 1 h prior to virus inoculation. Results of dose titration experiments suggested that RSV infection was more sensitive to inhibition by BMS-433771 treatment in the BALB/c mouse host than in the cotton rat. This was reflected by the pharmacokinetic and pharmacodynamic analysis of the efficacy data, where the area under the concentration-time curve required to achieve 50% of the maximum response was approximately 7.5-fold less for mice than for cotton rats. Inhibition of RSV by BMS-433771 in the mouse is the result of F1-mediated inhibition, as shown by the fact that a virus selected for resistance to BMS-433771 in vitro and containing a single amino acid change in the F1 region was also refractory to treatment in the mouse host. BMS-433771 efficacy against RSV infection was also demonstrated for mice that were chemically immunosuppressed by cyclophosphamide treatment, indicating that compound inhibition

  20. Ozone effect on respiratory syncytial virus infectivity and cytokine production by human alveolar macrophages

    SciTech Connect

    Soukup, J.; Koren, H.S.; Becker, S. )

    1993-02-01

    This study was performed to evaluate the effect of ozone (O3) exposure at 1 ppm for 2 hr on the susceptibility/resistance of adult human alveolar macrophages (AM) to infection with respiratory syncytial virus (RSV) in vitro and on RSV-induced cytokine production by the AM. AM were first exposed to O3 or to filtered air and then infected with RSV at multiplicities of infection (m.o.i.) of 0.1, 1.0, and 10. The percentage RSV-infected AM and the amount of infectious virus released by the cells were determined at Days 2 and 4 after infection. Interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF) levels in the supernatants were determined on Day 2. No difference in the percentage infected AM or in the amount of infectious RSV produced was found between control and O3-exposed cultures. However, O3-exposed AM infected with RSV at m.o.i. 1 produced less IL-1 in response to RSV infection than control AM: 63.6 pg/ml compared with 98.5 pg/ml. No difference in IL-1 was seen with m.o.i. 10. IL-6 levels were also decreased, but only after infection with m.o.i. 0.1. At this level of infection 830 pg/ml was produced by control AM as compared to 468.2 pg/ml by O3-exposed AM. TNF production was unaffected by O3 at all multiplicities of infection. Statistical analysis of the O3 effect on AM cytokine production induced by the different multiplicities, however, revealed no significant effect of O3. Based on these observations it appears unlikely that O3 alters susceptibility of AM to infection with RSV, nor does O3 dramatically alter cytokine production in response to RSV since effects on IL-1 and IL-6 secretion were only found with the lowest levels of infection which induced cytokine release.

  1. [Respiratory syncytial virus (RSV) infections in the pediatric teaching hospital Charles de Gaulle of Ouagadougou, Burkina Faso].

    PubMed

    Ouédraogo Yugbaré, S O; Ouédraogo, R; Nenebi, A; Traoré, B; Congo, L; Yonli, F; Kima, D; Bonané, P; Yé, D; Plantier, J-C; Vabret, A; Marguet, C; Gueudin, M

    2016-02-01

    Human respiratory syncytial virus (RSV) infections are little known in Burkina Faso. The objective of our work is to study the epidemiological and clinical aspects of RSV infections in infants in the Pediatric Teaching Hospital Charles de Gaulle of Ouagadougou. Between July 1(st) 2010 and June 30(th) 2011, we analyzed by direct immunofluorescence and PCR nasopharyngeal swabs from children from 0 to 36 months old. All in all, 210 patients among whom 74 from the external consultation (35.2%) and 136 hospitalized (64.7%) benefited from a nasopharyngeal aspiration. The motives for consultation were cough (91.7%), rhinitis (79.2%), fever (79.2%) and respiratory distress syndrome (66.7%). The evoked diagnoses were predominantly the acute bronchiolitis in 14 cases (58.3%) followed by the acute pulmonary disease in 7 patients (26.2%) then flue in 1 patient (16.7%). We detected by direct immunofluorescence the antigens of the respiratory viruses in 21 nasopharyngeal aspirations with 10 cases of respiratory syncytial virus (RSV) infections (47.6%). The PCR realized on 208 samples allowed to identify 153 positive samples (73.2%) with 24 RSV, i.e. a global prevalence of 16.1% with a peak of 18 cases (75%). In October, all the patients benefited from an often multiple antibiotic treatment of at least 10 days which was not still necessary. The evolution was favorable for all patients. This study confirms the important place of the viruses which are detected in 70% of the cases. The PCR multiplex, certainly expensive but effective and successful, deserves to be used in our developing countries to avoid the irrational prescription of antibiotic.

  2. Recombinant Subgroup B Human Respiratory Syncytial Virus Expressing Enhanced Green Fluorescent Protein Efficiently Replicates in Primary Human Cells and Is Virulent in Cotton Rats

    PubMed Central

    Lemon, Ken; Nguyen, D. Tien; Ludlow, Martin; Rennick, Linda J.; Yüksel, Selma; van Amerongen, Geert; McQuaid, Stephen; Rima, Bert K.; de Swart, Rik L.

    2014-01-01

    ABSTRACT Human respiratory syncytial virus (HRSV) is the most important viral cause of severe respiratory tract disease in infants. Two subgroups (A and B) have been identified, which cocirculate during, or alternate between, yearly epidemics and cause indistinguishable disease. Existing in vitro and in vivo models of HRSV focus almost exclusively on subgroup A viruses. Here, a recombinant (r) subgroup B virus (rHRSVB05) was generated based on a consensus genome sequence obtained directly from an unpassaged clinical specimen from a hospitalized infant. An additional transcription unit containing the gene encoding enhanced green fluorescent protein (EGFP) was introduced between the phosphoprotein and matrix genes (position 5) of the genome to generate rHRSVB05EGFP(5). The recombinant viruses replicated efficiently in both HEp-2 cells and in well-differentiated normal human bronchial cells grown at air-liquid interface. Intranasal infection of cotton rats (Sigmodon hispidus) resulted in high numbers of EGFP+ cells in epithelia of the nasal septum and conchae. When administered in a relatively large inoculum volume, the virus also replicated efficiently in bronchiolar epithelial cells and spread extensively in both the upper and lower respiratory tracts. Virus replication was not observed in ciliated epithelial cells of the trachea. This is the first virulent rHRSV strain with the genetic composition of a currently circulating wild-type virus. In vivo tracking of infected cells by means of EGFP fluorescence in the absence of cytopathic changes increases the sensitivity of virus detection in HRSV pathogenesis studies. IMPORTANCE Virology as a discipline has depended on monitoring cytopathic effects following virus culture in vitro. However, wild-type viruses isolated from patients often do not cause significant changes to infected cells, necessitating blind passage. This can lead to genetic and phenotypic changes and the generation of high-titer, laboratory

  3. Potential siRNA Molecules for Nucleoprotein and M2/L Overlapping Region of Respiratory Syncytial Virus: In Silico Design

    PubMed Central

    Shatizadeh Malekshahi, Somayeh; Arefian, Ehsan; Salimi, Vahid; Mokhtari Azad, Talat; Yavarian, Jila

    2016-01-01

    Background Human respiratory syncytial virus (RSV) is a leading cause of severe lower respiratory tract disease in the pediatric population, elderly and in immunosuppressed individuals. Respiratory syncytial virus is also responsible for bronchiolitis, pneumonia, and chronic obstructive pulmonary infections in all age groups. With this high disease burden and the lack of an effective RSV treatment and vaccine, there is a clear need for discovery and development of novel, effective and safe drugs to prevent and treat RSV disease. The most innovative approach is the use of small interfering RNAs (siRNAs) which represent a revolutionary new concept in human therapeutics. The nucleoprotein gene of RSV which is known as the most conserved gene and the M2/L mRNA, which encompass sixty-eight overlapping nucleotides, were selected as suitable targets for siRNA design. Objectives The present study is aimed to design potential siRNAs for silencing nucleoprotein and an overlapping region of M2-L coding mRNAs by computational analysis. Materials and Methods Various computational methods (target alignment, similarity search, secondary structure prediction, and RNA interaction calculation) have been used for siRNA designing against different strains of RSV. Results In this study, seven siRNA molecules were rationally designed against the nucleoprotein gene and validated using various computational methods for silencing different strains of RSV. Additionally, three effective siRNA molecules targeting the overlapping region of M2/L mRNA were designed. Conclusions This approach provides insight and a validated strategy for chemical synthesis of an antiviral RNA molecule which meets many sequence features for efficient silencing and treatment at the genomic level. PMID:27303618

  4. Immunostimulatory Defective Viral Genomes from Respiratory Syncytial Virus Promote a Strong Innate Antiviral Response during Infection in Mice and Humans

    PubMed Central

    Sun, Yan; Jain, Deepika; Koziol-White, Cynthia J.; Genoyer, Emmanuelle; Gilbert, Micah; Tapia, Karla; Panettieri, Reynold A.; Hodinka, Richard L.; López, Carolina B.

    2015-01-01

    Human respiratory syncytial virus (RSV) is a major cause of severe respiratory illness in children and susceptible adults. RSV blocks the development of the innate antiviral immune response and can grow to high titers in the respiratory tract. Here we demonstrate that immunostimulatory defective viral genomes (iDVGs) that are naturally generated during RSV replication are strong inducers of the innate antiviral response to RSV in mice and humans. In mice, RSV iDVGs stimulated the expression of antiviral genes, restricted viral replication, and prevented weight loss and lung inflammation. In human cells, the antiviral response to RSV iDVGs was dominated by the expression of IFN-λ1 over IFN-β and was driven by rapid intranuclear accumulation of the transcription factor IRF1. RSV iDVGs were detected in respiratory secretions of hospitalized patients, and their amount positively correlated with the level of expression of antiviral genes in the samples. Infection of explanted human lung tissue from different donors revealed that most humans can respond to RSV iDVGs and that the rate of accumulation of iDVGs during infection directly correlates with the quality of the antiviral response. Taken together, our data establish iDVGs as primary triggers of robust antiviral responses to RSV and provide the first evidence for an important biological role for naturally occurring iDVGs during a paramyxovirus infection in humans. PMID:26336095

  5. Clinical utility of a near patient care microarray based diagnostic test for influenza and respiratory syncytial virus infections

    PubMed Central

    Chen, Xiu-Hong; Wang, Ji-Hua; Yao, Xiao-Hong

    2015-01-01

    In primary care medicine, establishing a diagnosis of influenza and respiratory syncytial virus (RSV) infections is usually based on clinical history and physical examination as well as a consideration of time of the year and circulating respiratory viruses in the community. Methods: We tested the potential clinical samples using the automated molecular assay which included rapid influenza diagnostic test, Rapid Immunochromatographic Antigen Test, Verigene Respiratory Virus Plus Nucleic Acid Test, BD VeritorTM System for Rapid Detection of RSV in the paediatric setting for diagnosis of influenza and respiratory syntactical virus infections when testing was done by the paediatrician seeing the patient. Results: Principally, with respect influenza virus specificity and sensitivity for RIAT were 100% and 68.8%; compared to 100% and 100%, respectively for RV+. The specificity and sensitivity for 92.23% and 98% for BD VeritorTM System for Rapid Detection of RSV as compared to 96.6% and 98.42% for RIDT. Conclusion: Therefore, this study confirms the clinical utility of RV+ in the pediatric setting. PMID:26629177

  6. Ferrets as a Novel Animal Model for Studying Human Respiratory Syncytial Virus Infections in Immunocompetent and Immunocompromised Hosts

    PubMed Central

    Stittelaar, Koert J.; de Waal, Leon; van Amerongen, Geert; Veldhuis Kroeze, Edwin J.B.; Fraaij, Pieter L.A.; van Baalen, Carel A.; van Kampen, Jeroen J.A.; van der Vries, Erhard; Osterhaus, Albert D.M.E.; de Swart, Rik L.

    2016-01-01

    Human respiratory syncytial virus (HRSV) is an important cause of severe respiratory tract disease in immunocompromised patients. Animal models are indispensable for evaluating novel intervention strategies in this complex patient population. To complement existing models in rodents and non-human primates, we have evaluated the potential benefits of an HRSV infection model in ferrets (Mustela putorius furo). Nine- to 12-month-old HRSV-seronegative immunocompetent or immunocompromised ferrets were infected with a low-passage wild-type strain of HRSV subgroup A (105 TCID50) administered by intra-tracheal or intra-nasal inoculation. Immune suppression was achieved by bi-daily oral administration of tacrolimus, mycophenolate mofetil, and prednisolone. Throat and nose swabs were collected daily and animals were euthanized four, seven, or 21 days post-infection (DPI). Virus loads were determined by quantitative virus culture and qPCR. We observed efficient HRSV replication in both the upper and lower respiratory tract. In immunocompromised ferrets, virus loads reached higher levels and showed delayed clearance as compared to those in immunocompetent animals. Histopathological evaluation of animals euthanized 4 DPI demonstrated that the virus replicated in the respiratory epithelial cells of the trachea, bronchi, and bronchioles. These animal models can contribute to an assessment of the efficacy and safety of novel HRSV intervention strategies. PMID:27314379

  7. Investigating the Influence of Ribavirin on Human Respiratory Syncytial Virus RNA Synthesis by Using a High-Resolution Transcriptome Sequencing Approach

    PubMed Central

    Aljabr, Waleed; Touzelet, Olivier; Pollakis, Georgios; Wu, Weining; Munday, Diane C.; Hughes, Margaret; Hertz-Fowler, Christiane; Kenny, John; Fearns, Rachel; Barr, John N.

    2015-01-01

    ABSTRACT Human respiratory syncytial virus (HRSV) is a major cause of serious respiratory tract infection. Treatment options include administration of ribavirin, a purine analog, although the mechanism of its anti-HRSV activity is unknown. We used transcriptome sequencing (RNA-seq) to investigate the genome mutation frequency and viral mRNA accumulation in HRSV-infected cells that were left untreated or treated with ribavirin. In the absence of ribavirin, HRSV-specific transcripts accounted for up to one-third of total RNA reads from the infected-cell RNA population. Ribavirin treatment resulted in a >90% reduction in abundance of viral mRNA reads, while at the same time no such reduction was detected for the abundance of cellular transcripts. The presented data reveal that ribavirin significantly increases the frequency of HRSV-specific RNA mutations, suggesting a direct influence on the fidelity of the HRSV polymerase. The presented data show that transitions and transversions occur during HRSV replication and that these changes occur in hot spots along the HRSV genome. Examination of nucleotide substitution rates in the viral genome indicated an increase in the frequency of transition but not transversion mutations in the presence of ribavirin. In addition, our data indicate that in the continuous cell types used and at the time points analyzed, the abundances of some HRSV mRNAs do not reflect the order in which the mRNAs are transcribed. IMPORTANCE Human respiratory syncytial virus (HRSV) is a major pediatric pathogen. Ribavirin can be used in children who are extremely ill to reduce the amount of virus and to lower the burden of disease. Ribavirin is used as an experimental therapy with other viruses. The mechanism of action of ribavirin against HRSV is not well understood, although it is thought to increase the mutation rate of the viral polymerase during replication. To investigate this hypothesis, we used a high-resolution approach that allowed us to

  8. Interactome Analysis of the Human Respiratory Syncytial Virus RNA Polymerase Complex Identifies Protein Chaperones as Important Cofactors That Promote L-Protein Stability and RNA Synthesis

    PubMed Central

    Munday, Diane C.; Wu, Weining; Smith, Nikki; Fix, Jenna; Noton, Sarah Louise; Galloux, Marie; Touzelet, Olivier; Armstrong, Stuart D.; Dawson, Jenna M.; Aljabr, Waleed; Easton, Andrew J.; Rameix-Welti, Marie-Anne; de Oliveira, Andressa Peres; Simabuco, Fernando M.; Ventura, Armando M.; Hughes, David J.; Barr, John N.; Fearns, Rachel; Digard, Paul

    2014-01-01

    ABSTRACT The human respiratory syncytial virus (HRSV) core viral RNA polymerase comprises the large polymerase protein (L) and its cofactor, the phosphoprotein (P), which associate with the viral ribonucleoprotein complex to replicate the genome and, together with the M2-1 protein, transcribe viral mRNAs. While cellular proteins have long been proposed to be involved in the synthesis of HRSV RNA by associating with the polymerase complex, their characterization has been hindered by the difficulty of purifying the viral polymerase from mammalian cell culture. In this study, enhanced green fluorescent protein (EGFP)-tagged L- and P-protein expression was coupled with high-affinity anti-GFP antibody-based immunoprecipitation and quantitative proteomics to identify cellular proteins that interacted with either the L- or the P-proteins when expressed as part of a biologically active viral RNP. Several core groups of cellular proteins were identified that interacted with each viral protein including, in both cases, protein chaperones. Ablation of chaperone activity by using small-molecule inhibitors confirmed previously reported studies which suggested that this class of proteins acted as positive viral factors. Inhibition of HSP90 chaperone function in the current study showed that HSP90 is critical for L-protein function and stability, whether in the presence or absence of the P-protein. Inhibition studies suggested that HSP70 also disrupts virus biology and might help the polymerase remodel the nucleocapsid to allow RNA synthesis to occur efficiently. This indicated a proviral role for protein chaperones in HRSV replication and demonstrates that the function of cellular proteins can be targeted as potential therapeutics to disrupt virus replication. IMPORTANCE Human respiratory syncytial virus (HRSV) represents a major health care and economic burden, being the main cause of severe respiratory infections in infants worldwide. No vaccine or effective therapy is

  9. Respiratory Syncytial Virus Infection Upregulates NLRC5 and Major Histocompatibility Complex Class I Expression through RIG-I Induction in Airway Epithelial Cells

    PubMed Central

    Guo, Xuancheng; Liu, Taixiang; Shi, Hengfei; Wang, Jingjing; Ji, Ping; Wang, Hongwei; Hou, Yayi; Tan, Ren Xiang

    2015-01-01

    ABSTRACT Respiratory syncytial virus (RSV) is the leading cause of acute respiratory tract viral infection in infants, causing bronchiolitis and pneumonia. The host antiviral response to RSV acts via retinoic acid-inducible gene I (RIG-I). We show here that RSV infection upregulates major histocompatibility complex class I (MHC-I) expression through the induction of NLRC5, a NOD-like, CARD domain-containing intracellular protein that has recently been identified as a class I MHC transactivator (CITA). RSV infection of A549 cells promotes upregulation of NLRC5 via beta interferon (IFN-β) production, since the NLRC5-inducing activity in a conditioned medium from RSV-infected A549 cells was removed by antibody to IFN-β, but not by antibody to IFN-γ. RSV infection resulted in RIG-I upregulation and induction of NLRC5 and MHC-I. Suppression of RIG-I induction significantly blocked NLRC5, as well as MHC-I, upregulation and diminished IRF3 activation. Importantly, Vero cells deficient in interferon production still upregulated MHC-I following introduction of the RSV genome by infection or transfection, further supporting a key role for RIG-I. A model is therefore proposed in which the host upregulates MHC-I expression during RSV infection directly via the induction of RIG-I and NLRC5 expression. Since elevated expression of MHC-I molecules can sensitize host cells to T lymphocyte-mediated cytotoxicity or immunopathologic damage, the results have significant implications for the modification of immunity in RSV disease. IMPORTANCE Human respiratory syncytial virus (RSV) is the leading cause of bronchiolitis and pneumonia in infants and young children worldwide. Infection early in life is linked to persistent wheezing and allergic asthma in later life, possibly related to upregulation of major histocompatibility class I (MHC-I) on the cell surface, which facilitates cytotoxic T cell activation and antiviral immunity. Here, we show that RSV infection of lung epithelial

  10. Double-stranded RNA-activated protein kinase regulates early innate immune responses during respiratory syncytial virus infection.

    PubMed

    Minor, Radiah A Corn; Limmon, Gino V; Miller-DeGraff, Laura; Dixon, Darlene; Andrews, Danica M K; Kaufman, Randal J; Imani, Farhad

    2010-04-01

    Respiratory syncytial virus (RSV) is the most common cause of childhood viral bronchiolitis and lung injury. Inflammatory responses significantly contribute to lung pathologies during RSV infections and bronchiolitis but the exact mechanisms have not been completely defined. The double-stranded RNA-activated protein kinase (PKR) functions to inhibit viral replication and participates in several signaling pathways associated with innate inflammatory immune responses. Using a functionally defective PKR (PKR(-/-)) mouse model, we investigated the role of this kinase in early events of RSV-induced inflammation. Our data showed that bronchoalveolar lavage (BAL) fluid from infected PKR(-/-) mice had significantly lower levels of several innate inflammatory cytokines and chemokines. Histological examinations revealed that there was less lung injury in infected PKR(-/-) mice as compared to the wild type. A genome-wide analysis showed that several early antiviral and immune regulatory genes were affected by PKR activation. These data suggest that PKR is a signaling molecule for immune responses during RSV infections.

  11. [The standardization of an ultramicro ELISA assay for the detection of IgG antibodies to the respiratory syncytial virus].

    PubMed

    Savón, C; Laferté, J; Goyenechea, A; Valdivia, A; Morier, L; Tejeiro, Y

    1996-01-01

    An ultramicro ELISA assay of double antibody for the detection of IgG antibodies to the respiratory syncytial virus (RSV) was standardized. It was used a RVS antiprotein F monoclonal antibody produced by the Genetic Engineering and Biotechnology Center (GEBC) in Havana. The use of this antibody allowed to include crude antigenic preparations instead of purified fractions, which caused a significant reduction of the reactivity obtained with the antigen control. The assay conditions were determined by crossed titration. It was obtained a sensitivity of 97.2%, a coincidence of 91%, and a specificity of 83.3% of the UMELISA as regards the complement fixation. The results may be qualitatively expressed or by antibody titres using only one serum dilution (1:40) and a pattern curve.

  12. Innate and adaptive cellular phenotypes contributing to pulmonary disease in mice after respiratory syncytial virus immunization and infection.

    PubMed

    Lee, Young-Tae; Kim, Ki-Hye; Hwang, Hye Suk; Lee, Youri; Kwon, Young-Man; Ko, Eun-Ju; Jung, Yu-Jin; Lee, Yu-Na; Kim, Min-Chul; Kang, Sang-Moo

    2015-11-01

    Respiratory syncytial virus (RSV) is the major leading cause of infantile viral bronchiolitis. However, cellular phenotypes contributing to the RSV protection and vaccine-enhanced disease remain largely unknown. Upon RSV challenge, we analyzed phenotypes and cellularity in the lung of mice that were naïve, immunized with formalin inactivated RSV (FI-RSV), or re-infected with RSV. In comparison with naïve and live RSV re-infected mice, the high levels of eosinophils, neutrophils, plasmacytoid and CD11b(+) dendritic cells, and IL-4(+) CD4(+) T cells were found to be contributing to pulmonary inflammation in FI-RSV immune mice despite lung viral clearance. Alveolar macrophages appeared to play differential roles in protection and inflammation upon RSV infection of different RSV immune mice. These results suggest that multiple innate and adaptive immune components differentially contribute to RSV disease and inflammation.

  13. Characterization of oligosaccharide structures on a chimeric respiratory syncytial virus protein expressed in insect cell line Sf9

    SciTech Connect

    Wathen, M.W.; Aeed, P.A.; Elhammer, A.P. )

    1991-03-19

    The oligosaccharide structures added to a chimeric protein (FG) composed of the extracellular domains of respiratory syncytial virus F and G proteins, expressed in the insect cell line Sf9, were investigated. Cells were labeled in vivo with ({sup 3}H)glucosamine and infected wit a recombinant baculovirus containing the FG gene. The secreted chimeric protein was isolated by immunoprecipitation and subjected to oligosaccharide analysis. The FG protein contains two types of O-linked oligosaccharides: GalNAc and Gal{beta}1-3GalNAc constituting 17 and 66% of the total number of structures respectively. Only one type of N-linked oligosaccharide, constituting the remaining 17% of the structures on FG, was detected: a trimannosyl core structure with a fucose residue linked {alpha}1-6 to the asparagine-linked N-acetylglucosamine.

  14. Analysis of the interaction between respiratory syncytial virus and lipid-rafts in Hep2 cells during infection

    SciTech Connect

    Brown, Gaie; Jeffree, Chris E.; McDonald, Terence; McL Rixon, Helen W.; Aitken, James D.; Sugrue, Richard J. . E-mail: r.sugrue@vir.gla.ac.uk

    2004-10-01

    The assembly of respiratory syncytial virus (RSV) in lipid-rafts was examined in Hep2 cells. Confocal and electron microscopy showed that during RSV assembly, the cellular distribution of the complement regulatory proteins, decay accelerating factor (CD55) and CD59, changes and high levels of these cellular proteins are incorporated into mature virus filaments. The detergent-solubility properties of CD55, CD59, and the RSV fusion (F) protein were found to be consistent with each protein being located predominantly within lipid-raft structures. The levels of these proteins in cell-released virus were examined by immunoelectronmicroscopy and found to account for between 5% and 15% of the virus attachment (G) glycoprotein levels. Collectively, our findings suggest that an intimate association exists between RSV and lipid-raft membranes and that significant levels of these host-derived raft proteins, such as those regulating complement activation, are subsequently incorporated into the envelope of mature virus particles.

  15. A Proteomic-Based Workflow Using Purified Respiratory Syncytial Virus Particles to Identify Cellular Factors as Drug Targets.

    PubMed

    Huong, Tra Nguyen; Tan, Boon Huan; Sugrue, Richard J

    2016-01-01

    The identification of cellular factors that play a role in respiratory syncytial virus (RSV) replication is an alternative strategy in the identification of druggable cellular protein that are essential for RSV replication. In this regard experimental strategies that are able to screen relevant proteins from the vast array of proteins in the cellular milieu will facilitate the identification of potential drug targets. In this chapter we describe a procedure where RSV particles are purified from cells that are permissive for RSV infection, and the protein composition of the purified virus particles characterized using a proteomics-based strategy. This procedure revealed that actin, several actin-binding proteins, and the chaperones HSP70 and HSP90 also co-purified with the virus particles. The relevance of the HSP90 protein to virus replication was then further validated using imaging, gene silencing and by using an established small molecule HSP90 inhibitor. PMID:27464695

  16. Analysis of Immune Epitopes of Respiratory Syncytial Virus for Designing of Vectored Vaccines Based on Influenza Virus Platform.

    PubMed

    Isakova-Sivak, I N; Korenkov, D A; Fedorova, E A; Tretiak, T S; Matyushenko, V A; Smolonogina, T A; Rudenko, L G

    2016-08-01

    The immunoepitope database was used for analysis of experimentally detected epitopes of the respiratory syncytial virus (RSV) proteins and for selection of the epitope combinations for subsequent designing of recombinant vectored anti-RSV vaccines based on attenuated influenza viruses. Three cassettes containing the most promising B- and T-cell RSV epitopes were selected: peptide F (243-294) supporting the formation of humoral immunity in animals; fragment M2-1 (70-101+114-146) containing two MHC I epitopes (82-90 and 127-135); and MHC II-epitope (51-66). The selected constructions contained no neoepitopes causing undesirable effects of vaccination, such as immunotolerance or autoimmunity. PMID:27590768

  17. Real-Time Light Scattering Tracking of Gold Nanoparticles- bioconjugated Respiratory Syncytial Virus Infecting HEp-2 Cells

    NASA Astrophysics Data System (ADS)

    Wan, Xiao-Yan; Zheng, Lin-Ling; Gao, Peng-Fei; Yang, Xiao-Xi; Li, Chun-Mei; Li, Yuan Fang; Huang, Cheng Zhi

    2014-03-01

    Real-time tracking of virus invasion is crucial for understanding viral infection mechanism, which, however, needs simple and efficient labeling chemistry with improved signal-to-noise ratio. For that purpose, herein we investigated the invasion dynamics of respiratory syncytial virus (RSV) through dark-field microscopic imaging (iDFM) technique by using Au nanoparticles (AuNPs) as light scattering labels. RSV, a ubiquitous, non-segmented, pleiomorphic and negative-sense RNA virus, is an important human pathogen in infants, the elderly, and the immunocompromised. In order to label the enveloped virus of paramyxoviridae family, an efficient streptavidin (SA)-biotin binding chemistry was employed, wherein AuNPs and RSV particles modified with SA and biotin, respectively, allowing the AuNP-modified RSVs to maintain their virulence without affecting the native activities of RSV, making the long dynamic visualization successful for the RSV infections into human epidermis larynx carcinoma cells.

  18. Using mathematical transmission modelling to investigate drivers of respiratory syncytial virus seasonality in children in the Philippines.

    PubMed

    Paynter, Stuart; Yakob, Laith; Simões, Eric A F; Lucero, Marilla G; Tallo, Veronica; Nohynek, Hanna; Ware, Robert S; Weinstein, Philip; Williams, Gail; Sly, Peter D

    2014-01-01

    We used a mathematical transmission model to estimate when ecological drivers of respiratory syncytial virus (RSV) transmissibility would need to act in order to produce the observed seasonality of RSV in the Philippines. We estimated that a seasonal peak in transmissibility would need to occur approximately 51 days prior to the observed peak in RSV cases (range 49 to 67 days). We then compared this estimated seasonal pattern of transmissibility to the seasonal patterns of possible ecological drivers of transmissibility: rainfall, humidity and temperature patterns, nutritional status, and school holidays. The timing of the seasonal patterns of nutritional status and rainfall were both consistent with the estimated seasonal pattern of transmissibility and these are both plausible drivers of the seasonality of RSV in this setting. PMID:24587222

  19. Using Mathematical Transmission Modelling to Investigate Drivers of Respiratory Syncytial Virus Seasonality in Children in the Philippines

    PubMed Central

    Paynter, Stuart; Yakob, Laith; Simões, Eric A. F.; Lucero, Marilla G.; Tallo, Veronica; Nohynek, Hanna; Ware, Robert S.; Weinstein, Philip; Williams, Gail; Sly, Peter D.

    2014-01-01

    We used a mathematical transmission model to estimate when ecological drivers of respiratory syncytial virus (RSV) transmissibility would need to act in order to produce the observed seasonality of RSV in the Philippines. We estimated that a seasonal peak in transmissibility would need to occur approximately 51 days prior to the observed peak in RSV cases (range 49 to 67 days). We then compared this estimated seasonal pattern of transmissibility to the seasonal patterns of possible ecological drivers of transmissibility: rainfall, humidity and temperature patterns, nutritional status, and school holidays. The timing of the seasonal patterns of nutritional status and rainfall were both consistent with the estimated seasonal pattern of transmissibility and these are both plausible drivers of the seasonality of RSV in this setting. PMID:24587222

  20. Real-Time Light Scattering Tracking of Gold Nanoparticles- bioconjugated Respiratory Syncytial Virus Infecting HEp-2 Cells

    PubMed Central

    Wan, Xiao-Yan; Zheng, Lin-Ling; Gao, Peng-Fei; Yang, Xiao-Xi; Li, Chun-Mei; Li, Yuan Fang; Huang, Cheng Zhi

    2014-01-01

    Real-time tracking of virus invasion is crucial for understanding viral infection mechanism, which, however, needs simple and efficient labeling chemistry with improved signal-to-noise ratio. For that purpose, herein we investigated the invasion dynamics of respiratory syncytial virus (RSV) through dark-field microscopic imaging (iDFM) technique by using Au nanoparticles (AuNPs) as light scattering labels. RSV, a ubiquitous, non-segmented, pleiomorphic and negative-sense RNA virus, is an important human pathogen in infants, the elderly, and the immunocompromised. In order to label the enveloped virus of paramyxoviridae family, an efficient streptavidin (SA)-biotin binding chemistry was employed, wherein AuNPs and RSV particles modified with SA and biotin, respectively, allowing the AuNP-modified RSVs to maintain their virulence without affecting the native activities of RSV, making the long dynamic visualization successful for the RSV infections into human epidermis larynx carcinoma cells. PMID:24681709

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

    PubMed

    Amann, V L; Lerch, R A; Anderson, K; Wertz, G W

    1992-04-01

    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.

  2. Clinical and epidemiological comparison of human metapneumovirus and respiratory syncytial virus in seoul, Korea, 2003-2008.

    PubMed

    Kim, Chang Keun; Choi, Jungi; Callaway, Zak; Kim, Hyo Bin; Chung, Ju Young; Koh, Young-Yull; Shin, Bo Moon

    2010-03-01

    Human metapneumovirus (HMPV) shares clinical and epidemiological characteristics with well-known respiratory syncytial virus (RSV). The aim of this study was to investigate the clinical and epidemiological differences between HMPV- and RSV-induced wheezing illnesses. A total of 1,008 nasopharyngeal aspirate specimens was collected from 1,008 pediatric patients hospitalized with acute respiratory tract infection at Inje University Sanggye Paik Hospital from December 2003 to April 2008, and tested for seven common respiratory viruses. Conditions classified as wheezing illness were bronchiolitis, reactive airways disease, and bronchial asthma. HMPV caused a significantly lower proportion of wheezing illness when compared to RSV (48.1% vs. 82.2%, P<0.05). HMPV-induced wheezing illness occurred predominantly in older patients when compared to RSV patients (P<0.001). RSV infections peaked in the fall and winter followed by peaks of HMPV infection in winter and spring. Eosinophil counts were significantly higher (P<0.01) in RSV patients when compared to HMPV patients. These results show that human metapneumovirus patients exhibit several different clinical and epidemiological characteristics, such as higher proportion of wheezing illness, age and seasonal incidence, and eosinophil counts, when compared to RSV patients.

  3. Non-typeable Haemophilus influenzae protects human airway epithelial cells from a subsequent respiratory syncytial virus challenge.

    PubMed

    Hartwig, Stacey M; Ketterer, Margaret; Apicella, Michael A; Varga, Steven M

    2016-11-01

    Respiratory syncytial virus (RSV) and the common commensal and opportunistic pathogen, non-typeable Haemophilus influenzae (NTHi) both serve as a frequent cause of respiratory infection in children. Although it is well established that some respiratory viruses can increase host susceptibility to secondary bacterial infections, few studies have examined how commensal bacteria could influence a secondary viral response. Here, we examined the impact of NTHi exposure on a subsequent RSV infection of human bronchial epithelial cells (16HBE14o-). Co-culture of 16HBE14o- cells with NTHi resulted in inhibition of viral gene expression following RSV infection. 16HBE14o- cells co-cultured with heat-killed NTHi failed to protect against an RSV infection, indicating that protection requires live bacteria. However, NTHi did not inhibit influenza A virus replication, indicating that NTHi-mediated protection was RSV-specific. Our data demonstrates that prior exposure to a commensal bacterium such as NTHi can elicit protection against a subsequent RSV infection.

  4. Protective Efficacy and Immunogenicity of a Combinatory DNA Vaccine against Influenza A Virus and the Respiratory Syncytial Virus

    PubMed Central

    Stab, Viktoria; Nitsche, Sandra; Niezold, Thomas; Storcksdieck genannt Bonsmann, Michael; Wiechers, Andrea; Tippler, Bettina; Hannaman, Drew; Ehrhardt, Christina; Überla, Klaus

    2013-01-01

    The Respiratory Syncytial Virus (RSV) and Influenza A Virus (IAV) are both two major causative agents of severe respiratory tract infections in humans leading to hospitalization and thousands of deaths each year. In this study, we evaluated the immunogenicity and efficacy of a combinatory DNA vaccine in comparison to the single component vaccines against both diseases in a mouse model. Intramuscular electroporation with plasmids expressing the hemagglutinin (HA) of IAV and the F protein of RSV induced strong humoral immune responses regardless if they were delivered in combination or alone. In consequence, high neutralizing antibody titers were detected, which conferred protection against a lethal challenge with IAV. Furthermore, the viral load in the lungs after a RSV infection could be dramatically reduced in vaccinated mice. Concurrently, substantial amounts of antigen-specific, polyfunctional CD8+ T-cells were measured after vaccination. Interestingly, the cellular response to the hemagglutinin was significantly reduced in the presence of the RSV-F encoding plasmid, but not vice versa. Although these results indicate a suppressive effect of the RSV-F protein, the protective efficacy of the combinatory vaccine was comparable to the efficacy of both single-component vaccines. In conclusion, the novel combinatory vaccine against RSV and IAV may have great potential to reduce the rate of severe respiratory tract infections in humans without increasing the number of necessary vaccinations. PMID:23967287

  5. Non-typeable Haemophilus influenzae protects human airway epithelial cells from a subsequent respiratory syncytial virus challenge.

    PubMed

    Hartwig, Stacey M; Ketterer, Margaret; Apicella, Michael A; Varga, Steven M

    2016-11-01

    Respiratory syncytial virus (RSV) and the common commensal and opportunistic pathogen, non-typeable Haemophilus influenzae (NTHi) both serve as a frequent cause of respiratory infection in children. Although it is well established that some respiratory viruses can increase host susceptibility to secondary bacterial infections, few studies have examined how commensal bacteria could influence a secondary viral response. Here, we examined the impact of NTHi exposure on a subsequent RSV infection of human bronchial epithelial cells (16HBE14o-). Co-culture of 16HBE14o- cells with NTHi resulted in inhibition of viral gene expression following RSV infection. 16HBE14o- cells co-cultured with heat-killed NTHi failed to protect against an RSV infection, indicating that protection requires live bacteria. However, NTHi did not inhibit influenza A virus replication, indicating that NTHi-mediated protection was RSV-specific. Our data demonstrates that prior exposure to a commensal bacterium such as NTHi can elicit protection against a subsequent RSV infection. PMID:27573069

  6. Inhibition of G1P3 expression found in the differential display study on respiratory syncytial virus infection

    PubMed Central

    Zhao, Dongchi; Peng, Dan; Li, Lei; Zhang, Qiwei; Zhang, Chuyu

    2008-01-01

    Background Respiratory syncytial virus (RSV) is the leading viral pathogen associated with bronchiolitis and lower respiratory tract disease in infants and young children worldwide. The respiratory epithelium is the primary initiator of pulmonary inflammation in RSV infections, which cause significant perturbations of global gene expression controlling multiple cellular processes. In this study, differential display reverse transcription polymerase chain reaction amplification was performed to examine mRNA expression in a human alveolar cell line (SPC-A1) infected with RSV. Results Of the 2,500 interpretable bands on denaturing polyacrylamide gels, 40 (1.6%) cDNA bands were differentially regulated by RSV, in which 28 (70%) appeared to be upregulated and another 12 (30%) appeared to be downregulated. Forty of the expressed sequence tags (EST) were isolated, and 20 matched homologs in GenBank. RSV infection upregulated the mRNA expression of chemokines CC and CXC and interfered with type α/β interferon-inducible gene expression by upregulation of MG11 and downregulation of G1P3. Conclusion RSV replication could induce widespread changes in gene expression including both positive and negative regulation and play a different role in the down-regulation of IFN-α and up-regulation of IFN-γ inducible gene expression, which suggests that RSV interferes with the innate antiviral response of epithelial cells by multiple mechanisms. PMID:18838000

  7. Partial Attenuation of Respiratory Syncytial Virus with a Deletion of a Small Hydrophobic Gene Is Associated with Elevated Interleukin-1β Responses

    PubMed Central

    Russell, Ryan F.; McDonald, Jacqueline U.; Ivanova, Maria; Zhong, Ziyun; Bukreyev, Alexander

    2015-01-01

    ABSTRACT The small hydrophobic (SH) gene of respiratory syncytial virus (RSV), a major cause of infant hospitalization, encodes a viroporin of unknown function. SH gene knockout virus (RSV ΔSH) is partially attenuated in vivo, but not in vitro, suggesting that the SH protein may have an immunomodulatory role. RSV ΔSH has been tested as a live attenuated vaccine in humans and cattle, and here we demonstrate that it protected against viral rechallenge in mice. We compared the immune response to infection with RSV wild type and RSV ΔSH in vivo using BALB/c mice and in vitro using epithelial cells, neutrophils, and macrophages. Strikingly, the interleukin-1β (IL-1β) response to RSV ΔSH infection was greater than to wild-type RSV, in spite of a decreased viral load, and when IL-1β was blocked in vivo, the viral load returned to wild-type levels. A significantly greater IL-1β response to RSV ΔSH was also detected in vitro, with higher-magnitude responses in neutrophils and macrophages than in epithelial cells. Depleting macrophages (with clodronate liposome) and neutrophils (with anti-Ly6G/1A8) demonstrated the contribution of these cells to the IL-1β response in vivo, the first demonstration of neutrophilic IL-1β production in response to viral lung infection. In this study, we describe an increased IL-1β response to RSV ΔSH, which may explain the attenuation in vivo and supports targeting the SH gene in live attenuated vaccines. IMPORTANCE There is a pressing need for a vaccine for respiratory syncytial virus (RSV). A number of live attenuated RSV vaccine strains have been developed in which the small hydrophobic (SH) gene has been deleted, even though the function of the SH protein is unknown. The structure of the SH protein has recently been solved, showing it is a pore-forming protein (viroporin). Here, we demonstrate that the IL-1β response to RSV ΔSH is greater in spite of a lower viral load, which contributes to the attenuation in vivo. This

  8. Detection of influenza vaccine effectiveness among nursery school children: Lesson from a season with cocirculating respiratory syncytial virus

    PubMed Central

    Nakata, Keiko; Fujieda, Megumi; Miki, Hitoshi; Fukushima, Wakaba; Ohfuji, Satoko; Maeda, Akiko; Kase, Tetsuo; Hirota, Yoshio

    2015-01-01

    In the winter influenza epidemic season, patients with respiratory illnesses including respiratory syncytial virus (RSV) infections increase among young children. Therefore, we evaluated the effectiveness of influenza vaccine against influenza-like illness (ILI) using a technique to identify outbreaks of RSV infection and to distinguish those patients from ILI patients. The study subjects were 101 children aged 12 to 84 months attending nursery school. We classified the cases into 6 levels based on the definitions of ILI for outcomes. We established observation periods according to information obtained from regional surveillance and rapid diagnostic tests among children. Multivariate odds ratios (ORs) for each case classification were obtained using a logistic regression model for each observation period. For the entire observation period, ORs for cases with fever plus respiratory symptoms were reduced marginally significantly. For the local influenza epidemic period, only the OR for the most serious cases was significantly decreased (0.20 [95%CI: 0.04-0.94]). During the influenza outbreak among the nursery school children, multivariate ORs for fever plus respiratory symptoms decreased significantly (≥ 38.0°C plus ≥ one symptoms: 0.23 [0.06-0.91), ≥ 38.0°C plus ≥ 2 symptoms: 0.21 [0.05-0.85], ≥ 39.0°C plus ≥ one symptoms: 0.18 [0.04-0.93] and ≥ 39.0°C plus ≥ 2 symptoms: 0.16 [0.03-0.87]). These results suggest that confining observation to the peak influenza epidemic period and adoption of a strict case classification system can minimize outcome misclassification when evaluating the effectiveness of influenza vaccine against ILI, even if influenza and RSV cocirculate in the same season. PMID:25714791

  9. Control of pathogenic effector T-cell activities in situ by PD-L1 expression on respiratory inflammatory dendritic cells during respiratory syncytial virus infection

    PubMed Central

    Yao, S; Jiang, L; Moser, EK; Jewett, LB; Wright, J; Du, J; Zhou, B; Davis, SD; Krupp, NL; Braciale, TJ; Sun, J

    2015-01-01

    Respiratory syncytial virus (RSV) infection is a leading cause of severe lower respiratory tract illness in young infants, the elderly and immunocompromised individuals. We demonstrate here that the co-inhibitory molecule programmed cell death 1 (PD-1) is selectively upregulated on T cells within the respiratory tract during both murine and human RSV infection. Importantly, the interaction of PD-1 with its ligand PD-L1 is vital to restrict the pro-inflammatory activities of lung effector T cells in situ, thereby inhibiting the development of excessive pulmonary inflammation and injury during RSV infection. We further identify that PD-L1 expression on lung inflammatory dendritic cells is critical to suppress inflammatory T-cell activities, and an interferon–STAT1–IRF1 axis is responsible for increased PD-L1 expression on lung inflammatory dendritic cells. Our findings suggest a potentially critical role of PD-L1 and PD-1 interactions in the lung for controlling host inflammatory responses and disease progression in clinical RSV infection. PMID:25465101

  10. Identification of a Human Respiratory Syncytial Virus Cell Entry Inhibitor by Using a Novel Lentiviral Pseudotype System

    PubMed Central

    Haid, Sibylle; Grethe, Christina; Bankwitz, Dorothea; Grunwald, Thomas

    2015-01-01

    ABSTRACT Lentiviral budding is governed by group-specific antigens (Gag proteins) and proceeds in the absence of cognate viral envelope proteins, which has been exploited to create pseudotypes incorporating envelope proteins from nonlentiviral families. Here, we report the generation of infectious lentiviral pseudoparticles incorporating human respiratory syncytial virus (hRSV) F protein alone (hRSV-Fpp) or carrying SH, G, and F proteins (hRSV-SH/G/Fpp). These particles recapitulate key infection steps of authentic hRSV particles, including utilization of glycosaminoglycans and low-pH-independent cell entry. Moreover, hRSV pseudoparticles (hRSVpp) can faithfully reproduce phenotypic resistance to a small-molecule fusion inhibitor in clinical development (BMS-433771) and a licensed therapeutic F protein-targeting antibody (palivizumab). Inoculation of several human cell lines from lung and liver revealed more than 30-fold differences in susceptibility to hRSVpp infection, suggesting differential expression of hRSV entry cofactors and/or restriction factors between these cell types. Moreover, we observed cell-type-dependent functional differences between hRSVpp carrying solely F protein or SH, G, and F proteins with regard to utilization of glycosaminoglycans. Using hRSVpp, we identified penta-O-galloyl-β-d-glucose (PGG) as a novel hRSV cell entry inhibitor. Moreover, we show that PGG also inhibits cell entry of hRSVpp carrying F proteins resistant to BMS-433771 or palivizumab. This work sheds new light on the mechanisms of hRSV cell entry, including possible strategies for antiviral intervention. Moreover, hRSVpp should prove valuable to dissect hRSV envelope protein functions, including the interaction with cell entry factors. IMPORTANCE Lentiviral pseudotypes are highly useful to specifically dissect the functions of viral and host factors in cell entry, which have been exploited for numerous viruses. Here, we successfully created hRSVpp and show that they

  11. Respiratory syncytial virus matrix protein induces lung epithelial cell cycle arrest through a p53 dependent pathway.

    PubMed

    Bian, Tao; Gibbs, John D; Örvell, Claes; Imani, Farhad

    2012-01-01

    Respiratory syncytial virus (RSV) is the major cause of viral respiratory infections in children. Our previous study showed that the RSV infection induced lung epithelial cell cycle arrest, which enhanced virus replication. To address the mechanism of RSV-induced cell cycle arrest, we examined the contribution of RSV-matrix (RSV-M) protein. In this report, we show that in both the A549 cell line and primary human bronchial epithelial (PHBE) cells, transfection with RSV-M protein caused the cells to proliferate at a slower rate than in control cells. The cell cycle analysis showed that RSV-M protein induced G1 phase arrest in A549 cells, and G1 and G2/M phase arrest in PHBE cells. Interestingly, RSV-M expression induced p53 and p21 accumulation and decreased phosphorylation of retinoblastoma protein (Rb). Further, induction of cell cycle arrest by RSV-M was not observed in a p53-deficient epithelial cell line (H1299). However, cell cycle arrest was restored after transfection of p53 cDNA into H1299 cells. Taken together, these results indicate that RSV-M protein regulates lung epithelial cell cycle through a p53-dependent pathway, which enhances RSV replication.

  12. The effect of dietary bovine colostrum on respiratory syncytial virus infection and immune responses following the infection in the mouse.

    PubMed

    Xu, Mei Ling; Kim, Hyoung Jin; Wi, Ga Ram; Kim, Hong-Jin

    2015-09-01

    Human respiratory syncytial virus (hRSV) is the most common cause of respiratory tract infection among young children because of immature T cell immunity of them against hRSV. CD8 T cells play a pivotal role in clearing hRSV and preventing subsequent infection. We examined the effects of dietary bovine colostrum on virus infection and CD8 T cell responses following hRSV infection in the mouse model. Mice received bovine colostrum for 14 days prior to hRSV challenge, and lung indexes (severity of symptom) and lung virus titers were analyzed. In addition, the activation of CD8 T cells in the bronchoalveolar lavage fluids (BALFs) of mice receiving bovine colostrum were compared with those in the BALFs of mice receiving phosphate-buffered saline (PBS) or ribavirin, post virus challenge. The severity of infection and lung virus titers were reduced in the mice receiving bovine colostrum, compared to those receiving PBS. Moreover CD8 T cell responses were selectively enhanced in the former. Our results suggest that dietary bovine colostrum exerts the effects to inhibit hRSV and ameliorate the symptom by hRSV infection, and enhances the CD8 T cell response during the hRSV infection. PMID:26310306

  13. Respiratory Syncytial Virus Utilizes a tRNA Fragment to Suppress Antiviral Responses Through a Novel Targeting Mechanism

    PubMed Central

    Deng, Junfang; Ptashkin, Ryan N; Chen, Yu; Cheng, Zhi; Liu, Guangliang; Phan, Thien; Deng, Xiaoling; Zhou, Jiehua; Lee, Inhan; Lee, Yong Sun; Bao, Xiaoyong

    2015-01-01

    Target identification is highly instructive in defining the biological roles of microRNAs. However, little is known about other small noncoding RNAs; for example, tRNA-derived RNA Fragments (tRFs). Some tRFs exhibit a gene-silencing mechanism distinctly different from that of typical microRNAs. We recently demonstrated that a respiratory syncytial virus (RSV)-induced tRF, called tRF5-GluCTC, promotes RSV replication. RSV is the single most important cause of lower respiratory tract infection in children. By using biochemical screening and bioinformatics analyses, we have identified apolipoprotein E receptor 2 (APOER2) as a target of tRF5-GluCTC. The 3′-portion of tRF5-GluCTC recognizes a target site in the 3′-untranslated region of APOER2 and suppresses its expression. We have also discovered that APOER2 is an anti-RSV protein whose suppression by tRF5-GluCTC promotes RSV replication. Our report represents the first identification of a natural target of a tRF and illustrates how a virus utilizes a host tRF to control a host gene to favor its replication. PMID:26156244

  14. Respiratory Syncytial Virus Utilizes a tRNA Fragment to Suppress Antiviral Responses Through a Novel Targeting Mechanism.

    PubMed

    Deng, Junfang; Ptashkin, Ryan N; Chen, Yu; Cheng, Zhi; Liu, Guangliang; Phan, Thien; Deng, Xiaoling; Zhou, Jiehua; Lee, Inhan; Lee, Yong Sun; Bao, Xiaoyong

    2015-10-01

    Target identification is highly instructive in defining the biological roles of microRNAs. However, little is known about other small noncoding RNAs; for example, tRNA-derived RNA Fragments (tRFs). Some tRFs exhibit a gene-silencing mechanism distinctly different from that of typical microRNAs. We recently demonstrated that a respiratory syncytial virus (RSV)-induced tRF, called tRF5-GluCTC, promotes RSV replication. RSV is the single most important cause of lower respiratory tract infection in children. By using biochemical screening and bioinformatics analyses, we have identified apolipoprotein E receptor 2 (APOER2) as a target of tRF5-GluCTC. The 3'-portion of tRF5-GluCTC recognizes a target site in the 3'-untranslated region of APOER2 and suppresses its expression. We have also discovered that APOER2 is an anti-RSV protein whose suppression by tRF5-GluCTC promotes RSV replication. Our report represents the first identification of a natural target of a tRF and illustrates how a virus utilizes a host tRF to control a host gene to favor its replication.

  15. IgG and IgM antibodies to viral glycoproteins in respiratory syncytial virus infections of graded severity.

    PubMed Central

    Toms, G L; Webb, M S; Milner, P D; Milner, A D; Routledge, E G; Scott, R; Stokes, G M; Swarbrick, A; Taylor, C E

    1989-01-01

    Serum antibodies to the fusion (F) and large glycoprotein (G) of respiratory syncytial virus in the serum of 57 infected infants were measured by enzyme linked immunosorbent assay (ELISA). Most serum samples taken at the time of admission to hospital contained antibodies to both glycoproteins, and overall there was no significant evidence of a selective deficiency of antibody to either viral antigen. Less than a quarter of the infants showed rising IgG antibody titres to either glycoprotein after infection, whereas over threequarters produced an IgM response. There was a significant correlation between IgG response to viral glycoproteins and the age of the infant. The correlation of age with the IgM response was less pronounced, and there was no correlation between serum IgG antibody derived transplancentally in the acute phase of infection and IgM response to either glycoprotein. Neither IgG or IgM responses correlated with a clinical assessment of the severity of infection in the infants. IgM responses, however, were weakly correlated with reduced secretion of infectious virus in the upper respiratory tract. PMID:2624472

  16. Generation and Characterization of ALX-0171, a Potent Novel Therapeutic Nanobody for the Treatment of Respiratory Syncytial Virus Infection

    PubMed Central

    Stohr, Thomas; Palomo, Concepción; Piedra, Pedro A.; Gilbert, Brian E.; Mas, Vicente; Millar, Andrena; Power, Ultan F.; Stortelers, Catelijne; Allosery, Koen; Melero, José A.; Depla, Erik

    2015-01-01

    Respiratory syncytial virus (RSV) is an important causative agent of lower respiratory tract infections in infants and elderly individuals. Its fusion (F) protein is critical for virus infection. It is targeted by several investigational antivirals and by palivizumab, a humanized monoclonal antibody used prophylactically in infants considered at high risk of severe RSV disease. ALX-0171 is a trimeric Nanobody that binds the antigenic site II of RSV F protein with subnanomolar affinity. ALX-0171 demonstrated in vitro neutralization superior to that of palivizumab against prototypic RSV subtype A and B strains. Moreover, ALX-0171 completely blocked replication to below the limit of detection for 87% of the viruses tested, whereas palivizumab did so for 18% of the viruses tested at a fixed concentration. Importantly, ALX-0171 was highly effective in reducing both nasal and lung RSV titers when delivered prophylactically or therapeutically directly to the lungs of cotton rats. ALX-0171 represents a potent novel antiviral compound with significant potential to treat RSV-mediated disease. PMID:26438495

  17. A single amino acid in the F2 subunit of respiratory syncytial virus fusion protein alters growth and fusogenicity

    PubMed Central

    Schickli, Jeanne H.; Tang, Roderick S.

    2013-01-01

    Respiratory syncytial virus (RSV) causes severe lower respiratory tract infection in children, especially in infants less than 1 year of age. There are currently no licensed vaccines against RSV. rA2ΔM2-2 is a promising live-attenuated vaccine candidate that is currently being evaluated in the clinic. Attenuation of rA2ΔM2-2 is achieved by a single deletion of the M2-2 gene, which disrupts the balance between viral transcription and replication. Whilst performing a manufacturing feasibility study in a serum-free adapted Vero cell line, differences in growth kinetics and cytopathic effect (CPE) were identified between two rA2ΔM2-2 vaccine candidates. Comparative sequence analysis identified four amino acid differences between the two vaccine viruses. Recombinant rA2ΔM2-2 viruses carrying each of the four amino acid differences identified a K66E mutation in the F2 fragment of the fusion (F) protein as the cause of the growth and CPE differences. Syncytium-formation experiments with RSV F protein carrying mutations at aa 66 suggested that a change in charge at this residue within the F2 fragment can have a significant impact on fusion. PMID:24092758

  18. Inhibition of priming for bovine respiratory syncytial virus-specific protective immune responses following parenteral vaccination of passively immune calves.

    PubMed

    Ellis, John; Gow, Sheryl; Bolton, Michael; Burdett, William; Nordstrom, Scott

    2014-12-01

    The effect of maternal antibodies (MatAb) on immunological priming by neonatal parenteral vaccination for bovine respiratory syncytial virus (BRSV) was addressed for the first time in experimental infection in 34 Holstein calves. Both vaccinated and control calves developed moderate to severe respiratory disease characteristic of acute BRSV infection. There were no differences in clinical signs, BRSV shed, arterial oxygen concentrations, or mortality between vaccinated and control calves after BRSV challenge approximately 11 wk after vaccination. There were no anamnestic antibody or cytokine responses in the vaccinates after challenge. Lung lesions were extensive in both groups, and although there was a statistically significant (P = 0.05) difference between groups, this difference was considered not biologically significant. These data indicate that stimulation of protective immune responses was inhibited by maternal antibodies when a combination modified-live BRSV vaccine was administered parenterally to young passively immune calves. Alternate routes of administration or different vaccine formulations should be used to successfully immunize young calves with good passive antibody transfer.

  19. Evolution and Transmission of Respiratory Syncytial Group A (RSV-A) Viruses in Guangdong, China 2008–2015

    PubMed Central

    Zou, Lirong; Yi, Lina; Wu, Jie; Song, Yingchao; Huang, Guofeng; Zhang, Xin; Liang, Lijun; Ni, Hanzhong; Pybus, Oliver G.; Ke, Changwen; Lu, Jing

    2016-01-01

    Respiratory syncytial viruses (RSVs) including subgroups A (RSV-A) and B (RSV-B) are an important cause of acute respiratory tract infections worldwide. RSV-A include major epidemic strains. Fundamental questions concerning the evolution, persistence and transmission of RSV-A are critical for disease control and prevention, yet remain unanswered. In this study, we generated 64 complete G gene sequences of RSV-A strains collected between 2008 and 2015 in Guangdong, China. Phylogenetic analysis was undertaken by incorporating 572 publicly available RSV-A sequences. Current data indicate that genotypes GA1, GA4, and GA5 are endemic with limited epidemic activity. In contrast, the GA2 genotype which likely originated in 1980 has spread rapidly and caused epidemics worldwide. By analyzing GA2 genotype sequences across epidemic seasons within Guangdong, we find that RSV-A epidemics in Guangdong are caused by a combination of virus importation and local persistence, although the magnitude of the latter is likely overestimated due to infrequent sampling in other regions. Our results provide new insights into RSV-A evolution and transmission at global and local scales and highlights the rapid and wide spread of genotype GA2 compared to other genotypes. In order to control RSV transmission and outbreak, both local persistence and external introduction should be taken into account when designing optimal strategies. PMID:27574518

  20. Respiratory Syncytial Virus Matrix Protein Induces Lung Epithelial Cell Cycle Arrest through a p53 Dependent Pathway

    PubMed Central

    Bian, Tao; Gibbs, John D.; Örvell, Claes; Imani, Farhad

    2012-01-01

    Respiratory syncytial virus (RSV) is the major cause of viral respiratory infections in children. Our previous study showed that the RSV infection induced lung epithelial cell cycle arrest, which enhanced virus replication. To address the mechanism of RSV-induced cell cycle arrest, we examined the contribution of RSV-matrix (RSV-M) protein. In this report, we show that in both the A549 cell line and primary human bronchial epithelial (PHBE) cells, transfection with RSV-M protein caused the cells to proliferate at a slower rate than in control cells. The cell cycle analysis showed that RSV-M protein induced G1 phase arrest in A549 cells, and G1 and G2/M phase arrest in PHBE cells. Interestingly, RSV-M expression induced p53 and p21 accumulation and decreased phosphorylation of retinoblastoma protein (Rb). Further, induction of cell cycle arrest by RSV-M was not observed in a p53-deficient epithelial cell line (H1299). However, cell cycle arrest was restored after transfection of p53 cDNA into H1299 cells. Taken together, these results indicate that RSV-M protein regulates lung epithelial cell cycle through a p53-dependent pathway, which enhances RSV replication. PMID:22662266

  1. Sirtuin 1 regulates dendritic cell activation and autophagy during Respiratory Syncytial Virus-induced immune responses1

    PubMed Central

    Owczarczyk, Anna B.; Schaller, Matthew A.; Reed, Michelle; Rasky, Andrew J.; Lombard, David B.; Lukacs, Nicholas W.

    2015-01-01

    Respiratory syncytial virus (RSV) is the major cause of lower respiratory tract infection in children worldwide. Sirtuin 1 (SIRT1), an NAD+ dependent deacetylase, has been associated with the induction of autophagy and the regulation of inflammatory mediators. We found that Sirt1 was upregulated in mouse lung after RSV infection. Infected animals that received EX-527, a selective SIRT1 inhibitor, displayed exacerbated lung pathology, with increased mucus production, elevated viral load, and enhanced Th2 cytokine production. Gene expression analysis of isolated cell populations revealed that Sirt1 was most highly upregulated in RSV-treated dendritic cells (DCs). Upon RSV infection, EX-527-treated DCs, Sirt1 siRNA-treated DCs, or DCs from conditional knockout (Sirt1f/f-CD11c–Cre+) mice showed downregulated inflammatory cytokine gene expression and attenuated autophagy. Finally, RSV infection of Sirt1f/f-CD11c–Cre+ mice resulted in altered lung and lymph node cytokine responses, leading to exacerbated pathology. These data indicate that SIRT1 promotes DC activation associated with autophagy-mediated processes during RSV infection, thereby directing efficient antiviral immune responses. PMID:26157176

  2. Cyclopiazonic acid, an inhibitor of calcium-dependent ATPases with antiviral activity against human respiratory syncytial virus.

    PubMed

    Cui, Rui; Wang, Yizhuo; Wang, Liu; Li, Guiming; Lan, Ke; Altmeyer, Ralf; Zou, Gang

    2016-08-01

    Human respiratory syncytial virus (RSV) is a common cause of lower respiratory tract infections in infants and young children worldwide, yet no vaccine or effective antiviral treatment is available. To search for new anti-RSV agents, we developed a cell-based assay that measures inhibition of RSV-induced cytopathic effect (CPE) and identified cyclopiazonic acid (CPA), an intracellular calcium ATPase inhibitor as a RSV inhibitor (EC50 values 4.13 μM) by screening of natural product library. CPA inhibited the replication of RSV strains belonging to both A and B subgroups and human parainfluenza virus type 3, but not Enterovirus 71. Mechanism of action study by time-of-addition assay and minigenome assay revealed that CPA acts at the step of virus genome replication and/or transcription. Moreover, two other calcium ATPase inhibitors (Thapsigargin and BHQ) and calcium ionophores (A23187 and ionomycin), but not calcium channel blockers (nifedipine, nimodipine, and tetrandrine), also had similar effect. These results indicate that an increase in intracellular calcium concentration is detrimental to RSV replication. Thus, our findings provide a new strategy for anti-RSV therapy via increasing intracellular calcium concentration. PMID:27210812

  3. Evolution and Transmission of Respiratory Syncytial Group A (RSV-A) Viruses in Guangdong, China 2008-2015.

    PubMed

    Zou, Lirong; Yi, Lina; Wu, Jie; Song, Yingchao; Huang, Guofeng; Zhang, Xin; Liang, Lijun; Ni, Hanzhong; Pybus, Oliver G; Ke, Changwen; Lu, Jing

    2016-01-01

    Respiratory syncytial viruses (RSVs) including subgroups A (RSV-A) and B (RSV-B) are an important cause of acute respiratory tract infections worldwide. RSV-A include major epidemic strains. Fundamental questions concerning the evolution, persistence and transmission of RSV-A are critical for disease control and prevention, yet remain unanswered. In this study, we generated 64 complete G gene sequences of RSV-A strains collected between 2008 and 2015 in Guangdong, China. Phylogenetic analysis was undertaken by incorporating 572 publicly available RSV-A sequences. Current data indicate that genotypes GA1, GA4, and GA5 are endemic with limited epidemic activity. In contrast, the GA2 genotype which likely originated in 1980 has spread rapidly and caused epidemics worldwide. By analyzing GA2 genotype sequences across epidemic seasons within Guangdong, we find that RSV-A epidemics in Guangdong are caused by a combination of virus importation and local persistence, although the magnitude of the latter is likely overestimated due to infrequent sampling in other regions. Our results provide new insights into RSV-A evolution and transmission at global and local scales and highlights the rapid and wide spread of genotype GA2 compared to other genotypes. In order to control RSV transmission and outbreak, both local persistence and external introduction should be taken into account when designing optimal strategies. PMID:27574518

  4. Targeting human respiratory syncytial virus transcription anti-termination factor M2-1 to inhibit in vivo viral replication

    PubMed Central

    Bailly, B.; Richard, C.-A.; Sharma, G.; Wang, L.; Johansen, L.; Cao, J.; Pendharkar, V.; Sharma, D.-C.; Galloux, M.; Wang, Y.; Cui, R.; Zou, G.; Guillon, P.; von Itzstein, M.; Eléouët, J.-F.; Altmeyer, R.

    2016-01-01

    Human respiratory syncytial virus (hRSV) is a leading cause of acute lower respiratory tract infection in infants, elderly and immunocompromised individuals. To date, no specific antiviral drug is available to treat or prevent this disease. Here, we report that the Smoothened receptor (Smo) antagonist cyclopamine acts as a potent and selective inhibitor of in vitro and in vivo hRSV replication. Cyclopamine inhibits hRSV through a novel, Smo-independent mechanism. It specifically impairs the function of the hRSV RNA-dependent RNA polymerase complex notably by reducing expression levels of the viral anti-termination factor M2-1. The relevance of these findings is corroborated by the demonstration that a single R151K mutation in M2-1 is sufficient to confer virus resistance to cyclopamine in vitro and that cyclopamine is able to reduce virus titers in a mouse model of hRSV infection. The results of our study open a novel avenue for the development of future therapies against hRSV infection. PMID:27194388

  5. Prospective and retrospective evaluation of the Cepheid Xpert® Flu/RSV XC assay for rapid detection of influenza A, influenza B, and respiratory syncytial virus.

    PubMed

    Salez, Nicolas; Nougairede, Antoine; Ninove, Laetitia; Zandotti, Christine; de Lamballerie, Xavier; Charrel, Remi N

    2015-04-01

    A total of 281 clinical specimens (nasal swabs and nasopharyngeal aspirates) were tested with the Xpert® Flu/RSV XC. The results were compared to those obtained with the real-time retro transcriptase-polymerase chain reaction assays routinely used in our laboratory. The Xpert® Flu/RSV XC showed sensitivity/specificity of 97.8%/100% and 97.9%/100% for flu and respiratory syncytial virus, respectively.

  6. Animal Model of Respiratory Syncytial Virus: CD8+ T Cells Cause a Cytokine Storm That Is Chemically Tractable by Sphingosine-1-Phosphate 1 Receptor Agonist Therapy

    PubMed Central

    Walsh, Kevin B.; Teijaro, John R.; Brock, Linda G.; Fremgen, Daniel M.; Collins, Peter L.

    2014-01-01

    ABSTRACT The cytokine storm is an intensified, dysregulated, tissue-injurious inflammatory response driven by cytokine and immune cell components. The cytokine storm during influenza virus infection, whereby the amplified innate immune response is primarily responsible for pulmonary damage, has been well characterized. Now we describe a novel event where virus-specific T cells induce a cytokine storm. The paramyxovirus pneumonia virus of mice (PVM) is a model of human respiratory syncytial virus (hRSV). Unexpectedly, when C57BL/6 mice were infected with PVM, the innate inflammatory response was undetectable until day 5 postinfection, at which time CD8+ T cells infiltrated into the lung, initiating a cytokine storm by their production of gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α). Administration of an immunomodulatory sphingosine-1-phosphate (S1P) receptor 1 (S1P1R) agonist significantly inhibited PVM-elicited cytokine storm by blunting the PVM-specific CD8+ T cell response, resulting in diminished pulmonary disease and enhanced survival. IMPORTANCE A dysregulated overly exuberant immune response, termed a “cytokine storm,” accompanies virus-induced acute respiratory diseases (VARV), is primarily responsible for the accompanying high morbidity and mortality, and can be controlled therapeutically in influenza virus infection of mice and ferrets by administration of sphingosine-1-phosphate 1 receptor (S1P1R) agonists. Here, two novel findings are recorded. First, in contrast to influenza infection, where the cytokine storm is initiated early by the innate immune system, for pneumonia virus of mice (PVM), a model of RSV, the cytokine storm is initiated late in infection by the adaptive immune response: specifically, by virus-specific CD8 T cells via their release of IFN-γ and TNF-α. Blockading these cytokines with neutralizing antibodies blunts the cytokine storm and protects the host. Second, PVM infection is controlled by administration

  7. Respiratory Syncytial Virus Inhibits Ciliagenesis in Differentiated Normal Human Bronchial Epithelial Cells: Effectiveness of N-Acetylcysteine

    PubMed Central

    Mata, Manuel; Sarrion, Irene; Armengot, Miguel; Carda, Carmen; Martinez, Isidoro; Melero, Jose A.; Cortijo, Julio

    2012-01-01

    Persistent respiratory syncytial virus (RSV) infections have been associated with the exacerbation of chronic inflammatory diseases, including chronic obstructive pulmonary disease (COPD). This virus infects the respiratory epithelium, leading to chronic inflammation, and induces the release of mucins and the loss of cilia activity, two factors that determine mucus clearance and the increase in sputum volume. These alterations involve reactive oxygen species-dependent mechanisms. The antioxidant N-acetylcysteine (NAC) has proven useful in the management of COPD, reducing symptoms, exacerbations, and accelerated lung function decline. NAC inhibits RSV infection and mucin release in human A549 cells. The main objective of this study was to analyze the effects of NAC in modulating ciliary activity, ciliagenesis, and metaplasia in primary normal human bronchial epithelial cell (NHBEC) cultures infected with RSV. Our results indicated that RSV induced ultrastructural abnormalities in axonemal basal bodies and decreased the expression of β-tubulin as well as two genes involved in ciliagenesis, FOXJ1 and DNAI2. These alterations led to a decrease in ciliary activity. Furthermore, RSV induced metaplastic changes to the epithelium and increased the number of goblet cells and the expression of MUC5AC and GOB5. NAC restored the normal functions of the epithelium, inhibiting ICAM1 expression, subsequent RSV infection through mechanisms involving nuclear receptor factor 2, and the expression of heme oxygenase 1, which correlated with the restoration of the antioxidant capacity, the intracellular H2O2 levels and glutathione content of NHBECs. The results presented in this study support the therapeutic use of NAC for the management of chronic respiratory diseases, including COPD. PMID:23118923

  8. Respiratory syncytial virus inhibits ciliagenesis in differentiated normal human bronchial epithelial cells: effectiveness of N-acetylcysteine.

    PubMed

    Mata, Manuel; Sarrion, Irene; Armengot, Miguel; Carda, Carmen; Martinez, Isidoro; Melero, Jose A; Cortijo, Julio

    2012-01-01

    Persistent respiratory syncytial virus (RSV) infections have been associated with the exacerbation of chronic inflammatory diseases, including chronic obstructive pulmonary disease (COPD). This virus infects the respiratory epithelium, leading to chronic inflammation, and induces the release of mucins and the loss of cilia activity, two factors that determine mucus clearance and the increase in sputum volume. These alterations involve reactive oxygen species-dependent mechanisms. The antioxidant N-acetylcysteine (NAC) has proven useful in the management of COPD, reducing symptoms, exacerbations, and accelerated lung function decline. NAC inhibits RSV infection and mucin release in human A549 cells. The main objective of this study was to analyze the effects of NAC in modulating ciliary activity, ciliagenesis, and metaplasia in primary normal human bronchial epithelial cell (NHBEC) cultures infected with RSV. Our results indicated that RSV induced ultrastructural abnormalities in axonemal basal bodies and decreased the expression of β-tubulin as well as two genes involved in ciliagenesis, FOXJ1 and DNAI2. These alterations led to a decrease in ciliary activity. Furthermore, RSV induced metaplastic changes to the epithelium and increased the number of goblet cells and the expression of MUC5AC and GOB5. NAC restored the normal functions of the epithelium, inhibiting ICAM1 expression, subsequent RSV infection through mechanisms involving nuclear receptor factor 2, and the expression of heme oxygenase 1, which correlated with the restoration of the antioxidant capacity, the intracellular H(2)O(2) levels and glutathione content of NHBECs. The results presented in this study support the therapeutic use of NAC for the management of chronic respiratory diseases, including COPD. PMID:23118923

  9. Moderate local and systemic respiratory syncytial virus-specific T-cell responses upon mild or subclinical RSV infection.

    PubMed

    de Waal, L; Koopman, L P; van Benten, I J; Brandenburg, A H; Mulder, P G H; de Swart, R L; Fokkens, W J; Neijens, H J; Osterhaus, A D M E

    2003-06-01

    Respiratory syncytial virus (RSV) infections are a major cause of severe respiratory disease in infants. It has been shown that there is an increased frequency of childhood wheezing in ex-bronchiolitic preteen children. This was postulated to be mediated by a vigorous virus-specific Th2 response influencing the further development of the immune system. Little is known about the possible role of the immune response to clinically mild RSV infections in this respect. We have studied the RSV-specific cellular immune response in infants with a laboratory-confirmed RSV upper respiratory tract infection (URTI; n = 13, mean age 12 months, range 2-22 months) in comparison with infants with non-RSV mediated URTI (n = 9, mean age 9.3 months, range 4-18 months) or infants with severe RSV bronchiolitis (n = 11, mean age 2.3 months, range 1-6 months). RSV-specific cytokine-producing cells were enumerated using the ELISPOT method in peripheral blood mononuclear cells and nasal brush T-cells, collected during the acute and convalescent phase of the infection. Mixed Th1 (IFN-gamma) and Th2 (IL-4 and IL-13) responses were detected in all three groups. Frequencies of RSV-specific T-cells were lower in both URTI groups than in the RSV bronchiolitis group, and not significantly different between the RSV URTI and the non-RSV URTI group. The absence of vigorous virus-specific Th2 responses upon mild RSV infection does not support the hypothesis that these infections influence the development of the immune system and that they predispose for the development of atopic disease. PMID:12696123

  10. Core bead chromatography for preparation of highly pure, infectious respiratory syncytial virus in the negative purification mode.

    PubMed

    Mundle, Sophia T; Kishko, Michael; Groppo, Rachel; DiNapoli, Joshua; Hamberger, John; McNeil, Bryan; Kleanthous, Harry; Parrington, Mark; Zhang, Linong; Anderson, Stephen F

    2016-07-12

    Respiratory syncytial virus (RSV) is an important human pathogen, and is the most frequent viral cause of severe respiratory disease in infants. In addition, it is increasingly being recognized as an important cause of respiratory disease in the elderly and immunocompromised. Although a passive prophylactic treatment does exist for high-risk neonates and children, the overall disease burden warrants the development of a safe and effective prophylactic vaccine for use in otherwise healthy newborns and children. RSV is known to be an extremely labile virus, prone to aggregation and loss of infectious titer during virus handling and preparation procedures. To date infective RSV virions have been prepared by methods which are not readily scalable, such as density gradient ultracentrifugation. In this study we describe a scalable, chromatography-based purification procedure for preparation of highly pure, infectious RSV. The purification scheme is based on core bead technology and hollow fiber tangential flow filtration (TFF) and results in a ∼60% recovery of infectious virus titer. This method can be used to prepare highly purified wild type or live-attenuated vaccine strain viruses with titers as high as 1×10(8) plaque forming units per mL. A live-attenuated RSV vaccine prepared by this method was found to be immunogenic and protective in vivo, and its purity was 50-200-fold greater with respect to host cell dsDNA and Vero host cell proteins, than the raw feed stream. The results presented here can be considered a starting point for downstream process development of a live-attenuated vaccine approach for prevention of disease by RSV. PMID:27238375

  11. IgE anti-respiratory syncytial virus antibodies detected in serum of pediatric patients with asthma.

    PubMed

    Smith-Norowitz, Tamar A; Mandal, Mira; Joks, Rauno; Norowitz, Levana T; Weaver, Diana; Durkin, Helen G; Bluth, Martin H; Kohlhoff, Stephan

    2015-07-01

    Respiratory syncytial virus (RSV) causes lower respiratory tract disease in infants and young children, and is a public health concern, as is the increase in pediatric asthma. Respiratory viral infections may trigger asthma exacerbations. However, it remains unknown whether RSV infection may have a specific association with asthma. Total serum IgE, and IgE- and IgG-anti-RSV Ab responses were studied in older asthmatic compared with non-asthmatic children (M/F, mean age: 14) (N=30, N=43, respectively). We found: (1) total serum IgE was higher in asthmatic compared with non-asthmatics (P<0.001); (2) total serum IgE did correlate with IgE anti-RSV Abs (P<0.001), and with IgG anti-RSV Abs (P=0.008) in all subjects; (3) total serum IgE levels did correlate with IgE anti-RSV in asthmatics (P=0.047), but not in non-asthmatics (P=0.13); (4) IgE anti-RSV Abs did correlate with IgG anti-RSV Abs in all subjects (P=0.001); (5) IgE- and IgG-anti RSV Abs were higher in asthma compared with no asthma (P=0.003; <0.001, respectively); (6) there was a significant association between age and IgE anti-RSV in non-asthma (P=0.008), but not in asthma (P=0.64). Our findings indicate that IgE-anti-RSV Ab responses may play important roles in RSV infection and asthma.

  12. Respiratory syncytial virus inhibits ciliagenesis in differentiated normal human bronchial epithelial cells: effectiveness of N-acetylcysteine.

    PubMed

    Mata, Manuel; Sarrion, Irene; Armengot, Miguel; Carda, Carmen; Martinez, Isidoro; Melero, Jose A; Cortijo, Julio

    2012-01-01

    Persistent respiratory syncytial virus (RSV) infections have been associated with the exacerbation of chronic inflammatory diseases, including chronic obstructive pulmonary disease (COPD). This virus infects the respiratory epithelium, leading to chronic inflammation, and induces the release of mucins and the loss of cilia activity, two factors that determine mucus clearance and the increase in sputum volume. These alterations involve reactive oxygen species-dependent mechanisms. The antioxidant N-acetylcysteine (NAC) has proven useful in the management of COPD, reducing symptoms, exacerbations, and accelerated lung function decline. NAC inhibits RSV infection and mucin release in human A549 cells. The main objective of this study was to analyze the effects of NAC in modulating ciliary activity, ciliagenesis, and metaplasia in primary normal human bronchial epithelial cell (NHBEC) cultures infected with RSV. Our results indicated that RSV induced ultrastructural abnormalities in axonemal basal bodies and decreased the expression of β-tubulin as well as two genes involved in ciliagenesis, FOXJ1 and DNAI2. These alterations led to a decrease in ciliary activity. Furthermore, RSV induced metaplastic changes to the epithelium and increased the number of goblet cells and the expression of MUC5AC and GOB5. NAC restored the normal functions of the epithelium, inhibiting ICAM1 expression, subsequent RSV infection through mechanisms involving nuclear receptor factor 2, and the expression of heme oxygenase 1, which correlated with the restoration of the antioxidant capacity, the intracellular H(2)O(2) levels and glutathione content of NHBECs. The results presented in this study support the therapeutic use of NAC for the management of chronic respiratory diseases, including COPD.

  13. mRNA sequence of three respiratory syncytial virus genes encoding two nonstructural proteins and a 22K structural protein.

    PubMed Central

    Elango, N; Satake, M; Venkatesan, S

    1985-01-01

    An mRNA sequence of two human respiratory syncytial viral nonstructural protein genes and of a gene for a 22,000-molecular-weight (22K) protein was obtained by cDNA cloning and DNA sequencing. Sequences corresponding to the 5' ends of the respective transcripts were deduced directly by primer extension and dideoxy nucleotide sequencing of the mRNAs. The availability of a bicistronic clone (pRSC6) confirmed the gene order for this portion of the genome. Contrary to other unsegmented negative-stranded RNA viruses, a 19-nucleotide intercistronic sequence was present between the NS1 and NS2 genes. The translation of cloned viral sequences in the bicistronic and monocistronic clones (pRSNS1 and pRSNS2) revealed two moderately hydrophobic proteins of 15,568 and 14,703 daltons. Their similarity in molecular size explained our earlier inability to resolve these proteins. A DNA sequence of an additional recombinant plasmid (pRSA2) revealed a long open reading frame encoding a 22,156-dalton protein containing 194 amino acids. It was relatively basic and moderately hydrophobic. A protein of this size was readily translated in vitro from a viral mRNA hybrid selected by this plasmid and corresponded to an unglycosylated 22K protein seen in purified extracellular virus but not associated with detergent- and salt-resistant cores. A second open reading frame of 90 amino acids partially overlapping with the C terminus of the 22K protein was also present within this sequence. This was reminiscent of the viral matrix protein gene which was previously shown by us to contain two overlapping reading frames. The finding of three additional viral transcripts encoding at least three identifiable proteins in human respiratory syncytial virus was a novel departure from the usual genetic organization of paramyxoviruses. The 5' ends of all three transcripts had a 5'NGGGCAAAU sequence that is common to all viral transcripts analyzed so far. Although there was no obvious homology immediately

  14. Genetic variation of human respiratory syncytial virus among children with fever and respiratory symptoms in Shanghai, China, from 2009 to 2012.

    PubMed

    Liu, Jia; Mu, Yonglin; Dong, Wei; Yao, Fujia; Wang, Lili; Yan, Huajie; Lan, Ke; Zhang, Chiyu

    2014-10-01

    Human respiratory syncytial virus (HRSV) of genus Pneumovirus is one of the most common pathogens causing severe acute lower respiratory tract infection in infants and children. No information on the genotype distribution of HRSV is available in East China (e.g. Shanghai). From August 2009 to December 2012, 2407 nasopharyngeal swabs were collected from outpatient children with fever and respiratory symptoms in Shanghai. HRSV infection was determined using a multiplex RT-PCR assay. The second hypervariable region (HVR2) of G protein gene of HRSV was amplified and sequenced from HRSV positive samples. Genotypes were characterized by phylogenetic analyses. Of 2407 nasopharyngeal samples, 184 (7.6%) were tested as HRSV positive. From 160 positive subjects with sufficient nasopharyngeal samples, 69 HVR2 sequences were obtained by RT-PCR and sequencing. Three HRSV epidemic seasons were observed from August 2009 to December 2012, and an extreme outbreak of HRSV occurred in the 2009-2010 epidemic season. A genotype shift of predominant HRSV strains from B group in the 2009-2010 epidemic season to group A in the subsequent epidemic seasons was observed. Ten HRSV genotypes, including four group A genotypes NA1, NA3, NA4, and ON1, and six group B genotypes BA9, BA10, SAB4, CB1, BAc, and BA?, were detected in Shanghai. Seven genotypes (NA1, BA9-10, SAB4, CB1, BAc and BA?) were found in the 2009-2010 epidemic season. The co-circulation of multiple genotypes was associated with the extreme outbreak of HRSV among children with fever and respiratory symptoms in the 2009-2010 epidemic season.

  15. Influenza A virus among the hospitalized young children with acute respiratory infection. Is influenza A co infected with respiratory syncytial virus?

    PubMed Central

    Alavi, Seyed Mohammad; Makvandi, Manoochehr; Najafi-Fard, Saied; Alavi, Leila

    2012-01-01

    Background: Both influenza A virus (IAV) and respiratory syncytial virus (RSV) cause acute respiratory infection (ARI) in infants and young children. This study was conducted to determine Influenza A virus and its co infection with RSV among the hospitalized children with ARI. Methods: A total of 153 throat samples of the hospitalized young children aged between below one year and 5 years with the clinical signs of ARI were collected from the different hospitals in Khuzestan from June 2009 to April 2010. The samples were tested for Influenza A viruses by real time PCR. Positive IAV samples were tested for influenza A sub type H1N1 and for RSV by the nested PCR. Results: In this study, from the total 153 samples, 35 samples (22.9%) including 15 (42.8%) females and 20 (57.2%) males were positive for influenza A viruses. From the 35 positive samples for IAV, 14 were positive for swine H1N1 subtype. All the positive samples for influenza showed negative for RSV infection which revealed no coinfection with RSV. The prevalence of influenza A among age/sex groups was not significant. Conclusion: Influenza A is a prevalent viral agent isolated from young children with ARI. Influenza A subtype H1N1 was accounted for the 40 percent all laboratory-proven diagnoses of influenza in 2009. No evidence of coinfection of influenza A and RSV has been observed in the present study. PMID:24009929

  16. Baicalin from Scutellaria baicalensis blocks respiratory syncytial virus (RSV) infection and reduces inflammatory cell infiltration and lung injury in mice

    PubMed Central

    Shi, Hengfei; Ren, Ke; Lv, Baojie; Zhang, Wei; Zhao, Ying; Tan, Ren Xiang; Li, Erguang

    2016-01-01

    The roots of Scutellaria baicalensis has been used as a remedy for inflammatory and infective diseases for thousands of years. We evaluated the antiviral activity against respiratory syncytial virus (RSV) infection, the leading cause of childhood infection and hospitalization. By fractionation and chromatographic analysis, we determined that baicalin was responsible for the antiviral activity of S. baicalensis against RSV infection. The concentration for 50% inhibition (IC50) of RSV infection was determined at 19.9 ± 1.8 μM, while the 50% cytotoxic concentration (CC50) was measured at 370 ± 10 μM. We then used a mouse model of RSV infection to further demonstrate baicalin antiviral effect. RSV infection caused significant lung injury and proinflammatory response, including CD4 and CD8 T lymphocyte infiltration. Baicalin treatment resulted in reduction of T lymphocyte infiltration and gene expression of proinflammatory factors, while the treatment moderately reduced RSV titers recovered from the lung tissues. T lymphocyte infiltration and cytotoxic T lymphocyte modulated tissue damage has been identified critical factors of RSV disease. The study therefore demonstrates that baicalin subjugates RSV disease through antiviral and anti-inflammatory effect. PMID:27767097

  17. Identification and functional characterization of tRNA-derived RNA fragments (tRFs) in respiratory syncytial virus infection.

    PubMed

    Wang, Qingrong; Lee, Inhan; Ren, Junping; Ajay, Subramanian Shankar; Lee, Yong Sun; Bao, Xiaoyong

    2013-02-01

    The discovery of small noncoding RNAs (sncRNAs) with regulatory functions is a recent breakthrough in biology. Among sncRNAs, microRNA (miRNA), derived from host or virus, has emerged as elements with high importance in control of viral replication and host responses. However, the expression pattern and functional aspects of other types of sncRNAs, following viral infection, are unexplored. In order to define expression patterns of sncRNAs, as well as to discover novel regulatory sncRNAs in response to viral infection, we applied deep sequencing to cells infected with human respiratory syncytial virus (RSV), the most common cause of bronchiolitis and pneumonia in babies. RSV infection leads to abundant production of transfer RNA (tRNA)-derived RNA Fragments (tRFs) that are ~30 nucleotides (nts) long and correspond to the 5'-half of mature tRNAs. At least one tRF, which is derived from tRNA-Glu-CTC, represses target mRNA in the cytoplasm and promotes RSV replication. This demonstrates that this tRF is not a random by-product of tRNA degradation but a functional molecule. The biogenesis of this tRF is also specific, as it is mediated by the endonuclease angiogenin (ANG), not by other nucleases. In summary, our study presents novel information on the induction of a functional tRF by viral infection.

  18. Residues in human respiratory syncytial virus P protein that are essential for its activity on RNA viral synthesis.

    PubMed

    Asenjo, Ana; Mendieta, Jesús; Gómez-Puertas, Paulino; Villanueva, Nieves

    2008-03-01

    Human respiratory syncytial virus (HRSV) P protein, 241 amino acid long, is a structural homotetrameric phosphoprotein. Viral transcription and replication processes are dependent on functional P protein interactions inside viral ribonucleoprotein complexes (RNPs). Binding capacity to RNPs proteins and transcription and replication complementation analyses, using inactive P protein variants, have identified residues essential for functional interactions with itself, L, N and M2-1 proteins. P protein may establish some of these interactions as monomer, but efficient viral transcription and replication requires P protein oligomerization through the central region of the molecule. A structurally stable three-dimensional model has been generated in silico for this region (residues 98-158). Our analysis has indicated that P protein residues L135, D139, E140 and L142 are involved in homotetramerization. Additionally, the residues D136, S156, T160 and E179 appear to be essential for P protein activity on viral RNA synthesis and very high turnover phosphorylation at S143, T160 and T210 could regulate it. Thus, compounds targeted to those of these residues, located in the modeled three-dimensional structure, could have specific anti-HRSV effect.

  19. Type I interferons induce lung protease responses following respiratory syncytial virus infection via RIG-I-like receptors

    PubMed Central

    Foronjy, Robert F.; Taggart, Clifford C.; Dabo, Abdoulaye J.; Weldon, Sinéad; Cummins, Neville; Geraghty, Patrick

    2014-01-01

    The role of proteases in viral infection of the lung is poorly understood. Thus, we examined MMP and cathepsin proteases in respiratory syncytial virus (RSV) infected mouse lungs. RSV induced gene expression for matrix metalloproteinases (MMP) -2, -3, -7, -8, -9, -10, -12, -13, -14, -16, -17, -19, -20, -25, -27, -28 and cathepsins B, C, E, G, H, K, L1, S, W and Z in the airways of FVB/NJ mice. Increased proteases were present in the bronchoalveolar lavage fluid (BALF) and lung tissue during infection. Mitochondrial antiviral-signaling protein (Mavs) and Trif deficient mice were exposed to RSV. Mavs deficient mice had significantly lower expression of airway MMP-2, -3, -7, -8, -9, -10, -12, -13 and -28 and cathepsins C, G, K, S, W and Z. In lung epithelial cells, retinoic acid–inducible gene-1 (RIG-I) was identified as the major RIG-I- like receptor (RLR) required for RSV induced protease expression via MAVS. Overexpression of RIG-I or treatment with IFN-β in these cells induced MMP and cathepsin gene and protein expression. The significance of RIG-1 protease induction was demonstrated by the fact that inhibiting proteases with batimastat, E64 or ribavirin prevented airway hyperresponsiveness and enhanced viral clearance in RSV infected mice. PMID:25005357

  20. IL-13 is associated with reduced illness and replication in primary respiratory syncytial virus infection in the mouse

    PubMed Central

    Zhou, Weisong; Hashimoto, Koichi; Moore, Martin L.; Elias, Jack A.; Zhu, Zhou; Durbin, Joan; Colasurdo, Giuseppe; Rutigliano, John A.; Chiappetta, Constance L.; Goleniewska, Kasia; O’Neal, Jamye F.; Graham, Barney S.; Peebles, R. Stokes

    2007-01-01

    The role of IL-13 in respiratory syncytial virus (RSV) immunopathogenesis is incompletely described. To assess the effect of IL-13 on primary RSV infection, transgenic mice which either overexpress IL-13 in the lung (IL-13 OE) or nontransgenic littermates (IL-13 NT) were challenged intranasally with RSV. IL-13 OE mice had significantly decreased peak viral titers four days after infection compared to non-transgenic littermates. In addition, the IL-13 OE mice had significantly lower RSV-induced weight loss and reduced lung IFN-γ protein expression compared with IL-13 NT mice. In contrast, primary RSV challenge of IL-13 deficient mice resulted in a small, but statistically significant increase in viral titers on day four after infection, no difference in RSV-induced weight loss compared to wild type mice, and augmented IFN-γ production on day 6 after infection. In STAT1-deficient (STAT1 KO) mice, where primary RSV challenge produced high levels of IL-13 production in the lungs, treatment with an IL-13 neutralizing protein resulted in greater peak viral titers both four and six days after RSV and greater RSV-induced weight loss compared to mice treated with a control protein. These results suggest that IL-13 modulates illness from RSV-infection. PMID:17110149

  1. Structure of Respiratory Syncytial Virus Fusion Glycoprotein in the Postfusion Conformation Reveals Preservation of Neutralizing Epitopes▿†

    PubMed Central

    McLellan, Jason S.; Yang, Yongping; Graham, Barney S.; Kwong, Peter D.

    2011-01-01

    Respiratory syncytial virus (RSV) invades host cells via a type I fusion (F) glycoprotein that undergoes dramatic structural rearrangements during the fusion process. Neutralizing monoclonal antibodies, such as 101F, palivizumab, and motavizumab, target two major antigenic sites on the RSV F glycoprotein. The structures of these sites as peptide complexes with motavizumab and 101F have been previously determined, but a structure for the trimeric RSV F glycoprotein ectodomain has remained elusive. To address this issue, we undertook structural and biophysical studies on stable ectodomain constructs. Here, we present the 2.8-Å crystal structure of the trimeric RSV F ectodomain in its postfusion conformation. The structure revealed that the 101F and motavizumab epitopes are present in the postfusion state and that their conformations are similar to those observed in the antibody-bound peptide structures. Both antibodies bound the postfusion F glycoprotein with high affinity in surface plasmon resonance experiments. Modeling of the antibodies bound to the F glycoprotein predicts that the 101F epitope is larger than the linear peptide and restricted to a single protomer in the trimer, whereas motavizumab likely contacts residues on two protomers, indicating a quaternary epitope. Mechanistically, these results suggest that 101F and motavizumab can bind to multiple conformations of the fusion glycoprotein and can neutralize late in the entry process. The structural preservation of neutralizing epitopes in the postfusion state suggests that this conformation can elicit neutralizing antibodies and serve as a useful vaccine antigen. PMID:21613394

  2. A Numerically Subdominant CD8 T Cell Response to Matrix Protein of Respiratory Syncytial Virus Controls Infection with Limited Immunopathology

    PubMed Central

    Liu, Jie; Haddad, Elias K.; Marceau, Joshua; Morabito, Kaitlyn M.; Rao, Srinivas S.; Filali-Mouhim, Ali; Sekaly, Rafick-Pierre; Graham, Barney S.

    2016-01-01

    CD8 T cells are involved in pathogen clearance and infection-induced pathology in respiratory syncytial virus (RSV) infection. Studying bulk responses masks the contribution of individual CD8 T cell subsets to protective immunity and immunopathology. In particular, the roles of subdominant responses that are potentially beneficial to the host are rarely appreciated when the focus is on magnitude instead of quality of response. Here, by evaluating CD8 T cell responses in CB6F1 hybrid mice, in which multiple epitopes are recognized, we found that a numerically subdominant CD8 T cell response against DbM187 epitope of the virus matrix protein expressed high avidity TCR and enhanced signaling pathways associated with CD8 T cell effector functions. Each DbM187 T effector cell lysed more infected targets on a per cell basis than the numerically dominant KdM282 T cells, and controlled virus replication more efficiently with less pulmonary inflammation and illness than the previously well-characterized KdM282 T cell response. Our data suggest that the clinical outcome of viral infections is determined by the integrated functional properties of a variety of responding CD8 T cells, and that the highest magnitude response may not necessarily be the best in terms of benefit to the host. Understanding how to induce highly efficient and functional T cells would inform strategies for designing vaccines intended to provide T cell-mediated immunity. PMID:26943673

  3. A Numerically Subdominant CD8 T Cell Response to Matrix Protein of Respiratory Syncytial Virus Controls Infection with Limited Immunopathology.

    PubMed

    Liu, Jie; Haddad, Elias K; Marceau, Joshua; Morabito, Kaitlyn M; Rao, Srinivas S; Filali-Mouhim, Ali; Sekaly, Rafick-Pierre; Graham, Barney S

    2016-03-01

    CD8 T cells are involved in pathogen clearance and infection-induced pathology in respiratory syncytial virus (RSV) infection. Studying bulk responses masks the contribution of individual CD8 T cell subsets to protective immunity and immunopathology. In particular, the roles of subdominant responses that are potentially beneficial to the host are rarely appreciated when the focus is on magnitude instead of quality of response. Here, by evaluating CD8 T cell responses in CB6F1 hybrid mice, in which multiple epitopes are recognized, we found that a numerically subdominant CD8 T cell response against DbM187 epitope of the virus matrix protein expressed high avidity TCR and enhanced signaling pathways associated with CD8 T cell effector functions. Each DbM187 T effector cell lysed more infected targets on a per cell basis than the numerically dominant KdM282 T cells, and controlled virus replication more efficiently with less pulmonary inflammation and illness than the previously well-characterized KdM282 T cell response. Our data suggest that the clinical outcome of viral infections is determined by the integrated functional properties of a variety of responding CD8 T cells, and that the highest magnitude response may not necessarily be the best in terms of benefit to the host. Understanding how to induce highly efficient and functional T cells would inform strategies for designing vaccines intended to provide T cell-mediated immunity.

  4. Clarithromycin prevents human respiratory syncytial virus-induced airway epithelial responses by modulating activation of interferon regulatory factor-3.

    PubMed

    Yamamoto, Keisuke; Yamamoto, Soh; Ogasawara, Noriko; Takano, Kenichi; Shiraishi, Tsukasa; Sato, Toyotaka; Miyata, Ryo; Kakuki, Takuya; Kamekura, Ryuta; Kojima, Takashi; Tsutsumi, Hiroyuki; Himi, Tetsuo; Yokota, Shin-Ichi

    2016-09-01

    Macrolide antibiotics exert immunomodulatory activity by reducing pro-inflammatory cytokine production by airway epithelial cells, fibroblasts, vascular endothelial cells, and immune cells. However, the underlying mechanism of action remains unclear. Here, we examined the effect of clarithromycin (CAM) on pro-inflammatory cytokine production, including interferons (IFNs), by primary human nasal epithelial cells and lung epithelial cell lines (A549 and BEAS-2B cells) after stimulation by Toll-like receptor (TLR) and RIG-I-like receptor (RLR) agonists and after infection by human respiratory syncytial virus (RSV). CAM treatment led to a significant reduction in poly I:C- and RSV-mediated IL-8, CCL5, IFN-β and -λ production. Furthermore, IFN-β promoter activity (activated by poly I:C and RSV infection) was significantly reduced after treatment with CAM. CAM also inhibited IRF-3 dimerization and subsequent translocation to the nucleus. We conclude that CAM acts a crucial modulator of the innate immune response, particularly IFN production, by modulating IRF-3 dimerization and subsequent translocation to the nucleus of airway epithelial cells. This newly identified immunomodulatory action of CAM will facilitate the discovery of new macrolides with an anti-inflammatory role. PMID:27468646

  5. Vaccination against respiratory syncytial virus in pregnancy: a suitable tool to combat global infant morbidity and mortality?

    PubMed

    Saso, Anja; Kampmann, Beate

    2016-08-01

    Respiratory syncytial virus (RSV) is the most important viral cause of pneumonia in early childhood (ie, younger than 2 years), responsible for high infant morbidity and mortality worldwide. It is widely accepted that an effective vaccine against RSV would have a major impact on child health globally. Despite the setbacks of the clinical trials in the 1960s, there has been a recent and significant revival of interest in vaccines against RSV, with several promising candidates undergoing evaluation. In this Review, we describe the epidemiological and immunological background to RSV infection and subsequently focus on the promising pipeline of RSV vaccine development. We discuss the potential for implementation of a safe and immunogenic RSV vaccine within the context of global health and with regards to a range of strategies, including vaccination of women during pregnancy, which is likely to emerge as a beneficial and feasible public health tool. This approach would provide interim protection to vulnerable, RSV-naive infants and other high risk groups, in which the burden of admission to hospital and death is greatest. Extending research and implementation from resource-rich to resource-poor settings is required to enhance our understanding of RSV immunity and inform vaccine development and delivery strategies for all settings. We summarise key outstanding issues for researchers and policy makers to understand the interplay of biological and non-biological factors affecting design and distribution of a successful RSV vaccine globally. PMID:27317449

  6. Evidence that the respiratory syncytial virus polymerase complex associates with lipid rafts in virus-infected cells: a proteomic analysis

    SciTech Connect

    McDonald, Terence P.; Pitt, Andrew R.; Brown, Gaie; Rixon, Helen W. McL.; Sugrue, Richard J. . E-mail: r.sugrue@vir.gla.ac.uk

    2004-12-05

    The interaction between the respiratory syncytial virus (RSV) polymerase complex and lipid rafts was examined in HEp2 cells. Lipid-raft membranes were prepared from virus-infected cells and their protein content was analysed by Western blotting and mass spectrometry. This analysis revealed the presence of the N, P, L, M2-1 and M proteins. However, these proteins appeared to differ from one another in their association with these structures, with the M2-1 protein showing a greater partitioning into raft membranes compared to that of the N, P or M proteins. Determination of the polymerase activity profile of the gradient fractions revealed that 95% of the detectable viral enzyme activity was associated with lipid-raft membranes. Furthermore, analysis of virus-infected cells by confocal microscopy suggested an association between these proteins and the raft-lipid, GM1. Together, these results provide evidence that the RSV polymerase complex is able to associate with lipid rafts in virus-infected cells.

  7. Temporal association between the influenza virus and respiratory syncytial virus (RSV): RSV as a predictor of seasonal influenza.

    PubMed

    Míguez, A; Iftimi, A; Montes, F

    2016-09-01

    Epidemiologists agree that there is a prevailing seasonality in the presentation of epidemic waves of respiratory syncytial virus (RSV) infections and influenza. The aim of this study is to quantify the potential relationship between the activity of RSV, with respect to the influenza virus, in order to use the RSV seasonal curve as a predictor of the evolution of an influenza virus epidemic wave. Two statistical tools, logistic regression and time series, are used for predicting the evolution of influenza. Both logistic models and time series of influenza consider RSV information from previous weeks. Data consist of influenza and confirmed RSV cases reported in Comunitat Valenciana (Spain) during the period from week 40 (2010) to week 8 (2014). Binomial logistic regression models used to predict the two states of influenza wave, basal or peak, result in a rate of correct classification higher than 92% with the validation set. When a finer three-states categorization is established, basal, increasing peak and decreasing peak, the multinomial logistic model performs well in 88% of cases of the validation set. The ARMAX model fits well for influenza waves and shows good performance for short-term forecasts up to 3 weeks. The seasonal evolution of influenza virus can be predicted a minimum of 4 weeks in advance using logistic models based on RSV. It would be necessary to study more inter-pandemic seasons to establish a stronger relationship between the epidemic waves of both viruses. PMID:27165946

  8. A Preliminary Assessment of the Role of Ambient Nitric Oxide Exposure in Hospitalization with Respiratory Syncytial Virus Bronchiolitis.

    PubMed

    Mohammed, Nuredin I; Everard, Mark L; Ayres, Jon G; Barker, Nicola J; Litchfield, Ian J

    2016-01-01

    Some in vitro studies have indicated a possible link between respiratory syncytial virus (RSV) infection and exposure to Nitric Oxide (NO). However, these studies used much higher NO concentrations than normally found in the ambient environment. This preliminary study explored whether an association was present with short-term exposure to NO in the environment. RSV-related admission data between November 2011 and February 2012 were obtained from Sheffield Children's Hospital. The dates of admission were linked to contemporaneous ambient NO derived from sentinel air monitors. The case-crossover design was used to study the relationship between daily RSV admissions and NO, controlling for temperature and relative humidity. We found little evidence of association between daily RSV admission rates and exposure to ambient NO at different lags or average exposure across several lags. The findings should, however, be viewed with caution due to the low number of events observed during the time frame. It is possible that the apparent lack of association may be accounted for by the timing of the seasonal RSV epidemic in relation to peaks in NO concentrations. A larger study incorporating a wider range of RSV and NO peaks would determine whether said peaks enhanced the number of RSV hospitalizations in children. PMID:27294948

  9. Respiratory Syncytial Virus Nonstructural Proteins Upregulate SOCS1 and SOCS3 in the Different Manner from Endogenous IFN Signaling

    PubMed Central

    Zheng, Junwen; Yang, Pu; Tang, Yan; Pan, Zishu; Zhao, Dongchi

    2015-01-01

    Respiratory syncytial virus (RSV) infection upregulates genes of the suppressor of cytokine signaling (SOCS) family, which utilize a feedback loop to inhibit type I interferon dependent antiviral signaling pathway. Here, we reconstituted RSV nonstructural (NS) protein expression plasmids (pNS1, pNS2, and pNS1/2) and tested whether NS1 or NS2 would trigger SOCS1 and SOCS3 protein expression. These NS proteins inhibited interferon- (IFN-) α signaling through a mechanism involving the induction of SOCS1 and SOCS3, which appeared to be different from autocrine IFN dependent. NS1 induced both SOCS1 and SOCS3 upregulation, while NS2 only induced SOCS1 expression. The induced expression of SOCS1 and SOCS3 preceded endogenous IFN-signaling activation and inhibited the IFN-inducible antiviral response as well as chemokine induction. Treatments with INF-α and NS proteins both induced SOCS1 expression; however, they had opposing effects on IFN-α-dependent antiviral gene expression. Our results indicate that NS1 and NS2, which induce the expression of SOCS1 or SOCS3, might represent an independent pathway of stimulating endogenous IFN signaling. PMID:26557722

  10. Vaccination against respiratory syncytial virus in pregnancy: a suitable tool to combat global infant morbidity and mortality?

    PubMed

    Saso, Anja; Kampmann, Beate

    2016-08-01

    Respiratory syncytial virus (RSV) is the most important viral cause of pneumonia in early childhood (ie, younger than 2 years), responsible for high infant morbidity and mortality worldwide. It is widely accepted that an effective vaccine against RSV would have a major impact on child health globally. Despite the setbacks of the clinical trials in the 1960s, there has been a recent and significant revival of interest in vaccines against RSV, with several promising candidates undergoing evaluation. In this Review, we describe the epidemiological and immunological background to RSV infection and subsequently focus on the promising pipeline of RSV vaccine development. We discuss the potential for implementation of a safe and immunogenic RSV vaccine within the context of global health and with regards to a range of strategies, including vaccination of women during pregnancy, which is likely to emerge as a beneficial and feasible public health tool. This approach would provide interim protection to vulnerable, RSV-naive infants and other high risk groups, in which the burden of admission to hospital and death is greatest. Extending research and implementation from resource-rich to resource-poor settings is required to enhance our understanding of RSV immunity and inform vaccine development and delivery strategies for all settings. We summarise key outstanding issues for researchers and policy makers to understand the interplay of biological and non-biological factors affecting design and distribution of a successful RSV vaccine globally.

  11. A comparison of diagnostic methods for the detection of bovine respiratory syncytial virus in experimental clinical specimens.

    PubMed Central

    West, K; Bogdan, J; Hamel, A; Nayar, G; Morley, P S; Haines, D M; Ellis, J A

    1998-01-01

    Virus shedding was monitored in nasal secretions of 12 calves experimentally infected with bovine respiratory syncytial virus (BRSV) using an antigen capture enzyme-linked immunosorbent assay (ELISA) detecting the nucleoprotein (NP) antigen of BRSV, by a polymerase chain reaction (PCR) amplifying the fusion protein of BRSV, and by a microisolation assay combined with immunoperoxidase staining for the F protein of BRSV. Under the conditions of this study, similar limits of detection and quantitative results were obtained from all three assays. BRSV was detected in nasal secretions of all calves for a minimum of 4 d. Virus shedding began on Day 2 after infection, peaked on Days 3-5, and was cleared in most calves by Day 8. The PCR, and to a lesser extent the ELISA, may detect virus shedding for a longer period after infection than virus isolation, possibly due to neutralization of the virus by rising mucosal antibody. Simulated environmental conditions likely to be experienced during transport of clinical field specimens markedly reduced the sensitivity of virus isolation but had a minimal effect on the results of the NP ELISA. Actual field transport conditions (overnight on ice) had minimal apparent effect on the results of the PCR assay. The less stringent specimen handling requirements, combined with low limits of detection, of both the nucleoprotein ELISA and PCR, indicate either of these assays are more suitable for diagnostic applications than virus isolation. PMID:9798088

  12. Live-cell characterization and analysis of a clinical isolate of bovine respiratory syncytial virus, using molecular beacons.

    PubMed

    Santangelo, Philip; Nitin, Nitin; LaConte, Leslie; Woolums, Amelia; Bao, Gang

    2006-01-01

    Understanding viral pathogenesis is critical for prevention of outbreaks, development of antiviral drugs, and biodefense. Here, we utilize molecular beacons to directly detect the viral genome and characterize a clinical isolate of bovine respiratory syncytial virus (bRSV) in living cells. Molecular beacons are dual-labeled, hairpin oligonucleotide probes with a reporter fluorophore at one end and a quencher at the other; they are designed to fluoresce only when hybridizing to a complementary target. By imaging the fluorescence signal of molecular beacons, the spread of bRSV was monitored for 7 days with a signal-to-noise ratio of 50 to 200, and the measured time course of infection was quantified with a mathematical model for viral growth. We found that molecular beacon signal could be detected in single living cells infected with a viral titer of 2 x 10(3.6) 50% tissue culture infective doses/ml diluted 1,000 fold, demonstrating high detection sensitivity. Low background in uninfected cells and simultaneous staining of fixed cells with molecular beacons and antibodies showed high detection specificity. Furthermore, using confocal microscopy to image the viral genome in live, infected cells, we observed a connected, highly three-dimensional, amorphous inclusion body structure not seen in fixed cells. Taken together, the use of molecular beacons for active virus imaging provides a powerful tool for rapid viral infection detection, the characterization of RNA viruses, and the design of new antiviral drugs.

  13. Structural basis for immunization with postfusion respiratory syncytial virus fusion F glycoprotein (RSV F) to elicit high neutralizing antibody titers

    SciTech Connect

    Swanson, Kurt A.; Settembre, Ethan C.; Shaw, Christine A.; Dey, Antu K.; Rappuoli, Rino; Mandl, Christian W.; Dormitzer, Philip R.; Carfi, Andrea

    2012-02-07

    Respiratory syncytial virus (RSV), the main cause of infant bronchiolitis, remains a major unmet vaccine need despite more than 40 years of vaccine research. Vaccine candidates based on a chief RSV neutralization antigen, the fusion (F) glycoprotein, have foundered due to problems with stability, purity, reproducibility, and potency. Crystal structures of related parainfluenza F glycoproteins have revealed a large conformational change between the prefusion and postfusion states, suggesting that postfusion F antigens might not efficiently elicit neutralizing antibodies. We have generated a homogeneous, stable, and reproducible postfusion RSV F immunogen that elicits high titers of neutralizing antibodies in immunized animals. The 3.2-{angstrom} X-ray crystal structure of this substantially complete RSV F reveals important differences from homology-based structural models. Specifically, the RSV F crystal structure demonstrates the exposure of key neutralizing antibody binding sites on the surface of the postfusion RSV F trimer. This unanticipated structural feature explains the engineered RSV F antigen's efficiency as an immunogen. This work illustrates how structural-based antigen design can guide the rational optimization of candidate vaccine antigens.

  14. Structure of a Major Antigenic Site on the Respiratory Syncytial Virus Fusion Glycoprotein in Complex with Neutralizing Antibody 101F

    SciTech Connect

    McLellan, Jason S.; Chen, Man; Chang, Jung-San; Yang, Yongping; Kim, Albert; Graham, Barney S.; Kwong, Peter D.

    2010-11-19

    Respiratory syncytial virus (RSV) is a major cause of pneumonia and bronchiolitis in infants and elderly people. Currently there is no effective vaccine against RSV, but passive prophylaxis with neutralizing antibodies reduces hospitalizations. To investigate the mechanism of antibody-mediated RSV neutralization, we undertook structure-function studies of monoclonal antibody 101F, which binds a linear epitope in the RSV fusion glycoprotein. Crystal structures of the 101F antigen-binding fragment in complex with peptides from the fusion glycoprotein defined both the extent of the linear epitope and the interactions of residues that are mutated in antibody escape variants. The structure allowed for modeling of 101F in complex with trimers of the fusion glycoprotein, and the resulting models suggested that 101F may contact additional surfaces located outside the linear epitope. This hypothesis was supported by surface plasmon resonance experiments that demonstrated 101F bound the peptide epitope {approx}16,000-fold more weakly than the fusion glycoprotein. The modeling also showed no substantial clashes between 101F and the fusion glycoprotein in either the pre- or postfusion state, and cell-based assays indicated that 101F neutralization was not associated with blocking virus attachment. Collectively, these results provide a structural basis for RSV neutralization by antibodies that target a major antigenic site on the fusion glycoprotein.

  15. Reduced innate immune response, apoptosis, and virus release in cells cured of respiratory syncytial virus persistent infection.

    PubMed

    Herranz, Cristina; Melero, José A; Martínez, Isidoro

    2011-02-01

    It has been reported that cell clones isolated at different passages from a culture of HEp-2 cells infected persistently with human respiratory syncytial virus (HRSV) were cured of the virus. Further studies on one of these clones (31C1) are reported here, showing that 31C1 cells can still be infected by HRSV but release low amounts of virus to the culture supernatant, develop smaller and less numerous syncytia than the original HEp-2 cells, and display only a weak innate immune response to the infection. Accordingly, uninfected 31C1 cells, but not clones derived from uninfected HEp-2 cells, express low levels of TLR3 and RIG-I. In addition, 31C1 cells are partly resistant to apoptosis. These results indicate that persistent infection of HEp-2 cells by HRSV has selected cell variants, with changes affecting cell survival, virus growth and the innate immune response that may be valuable for studies of virus-cell interaction. PMID:21093006

  16. Molecular Basis for the Selective Inhibition of Respiratory Syncytial Virus RNA Polymerase by 2'-Fluoro-4'-Chloromethyl-Cytidine Triphosphate

    PubMed Central

    Deval, Jerome; Hong, Jin; Wang, Guangyi; Taylor, Josh; Smith, Lucas K.; Fung, Amy; Stevens, Sarah K.; Liu, Hong; Jin, Zhinan; Dyatkina, Natalia; Prhavc, Marija; Stoycheva, Antitsa D.; Serebryany, Vladimir; Liu, Jyanwei; Smith, David B.; Tam, Yuen; Zhang, Qingling; Moore, Martin L.; Fearns, Rachel; Chanda, Sushmita M.; Blatt, Lawrence M.; Symons, Julian A.; Beigelman, Leo

    2015-01-01

    Respiratory syncytial virus (RSV) causes severe lower respiratory tract infections, yet no vaccines or effective therapeutics are available. ALS-8176 is a first-in-class nucleoside analog prodrug effective in RSV-infected adult volunteers, and currently under evaluation in hospitalized infants. Here, we report the mechanism of inhibition and selectivity of ALS-8176 and its parent ALS-8112. ALS-8176 inhibited RSV replication in non-human primates, while ALS-8112 inhibited all strains of RSV in vitro and was specific for paramyxoviruses and rhabdoviruses. The antiviral effect of ALS-8112 was mediated by the intracellular formation of its 5'-triphosphate metabolite (ALS-8112-TP) inhibiting the viral RNA polymerase. ALS-8112 selected for resistance-associated mutations within the region of the L gene of RSV encoding the RNA polymerase. In biochemical assays, ALS-8112-TP was efficiently recognized by the recombinant RSV polymerase complex, causing chain termination of RNA synthesis. ALS-8112-TP did not inhibit polymerases from host or viruses unrelated to RSV such as hepatitis C virus (HCV), whereas structurally related molecules displayed dual RSV/HCV inhibition. The combination of molecular modeling and enzymatic analysis showed that both the 2'F and the 4'ClCH2 groups contributed to the selectivity of ALS-8112-TP. The lack of antiviral effect of ALS-8112-TP against HCV polymerase was caused by Asn291 that is well-conserved within positive-strand RNA viruses. This represents the first comparative study employing recombinant RSV and HCV polymerases to define the selectivity of clinically relevant nucleotide analogs. Understanding nucleotide selectivity towards distant viral RNA polymerases could not only be used to repurpose existing drugs against new viral infections, but also to design novel molecules. PMID:26098424

  17. Recombinant measles viruses expressing respiratory syncytial virus proteins induced virus-specific CTL responses in cotton rats.

    PubMed

    Yamaji, Yoshiaki; Nakayama, Tetsuo

    2014-07-31

    Respiratory syncytial virus (RSV) is a common cause of serious lower respiratory tract illnesses in infants. Natural infections with RSV provide limited protection against reinfection because of inefficient immunological responses that do not induce long-term memory. RSV natural infection has been shown to induce unbalanced immune response. The effective clearance of RSV is known to require the induction of a balanced Th1/Th2 immune response, which involves the induction of cytotoxic T lymphocytes (CTL). In our previous study, recombinant AIK-C measles vaccine strains MVAIK/RSV/F and MVAIK/RSV/G were developed, which expressed the RSV fusion (F) protein or glycoprotein (G). These recombinant viruses elicited antibody responses against RSV in cotton rats, and no infectious virus was recovered, but small amounts of infiltration of inflammatory cells were observed in the lungs following RSV challenge. In the present study, recombinant AIK-C measles vaccine strains MVAIK/RSV/M2-1 and MVAIK/RSV/NP were developed, expressing RSV M2-1 or Nucleoprotein (NP), respectively. These viruses exhibited temperature-sensitivity (ts), which was derived from AIK-C, and expressed respective RSV antigens. The intramuscular inoculation of cotton rats with the recombinant measles virus led to the induction of CD8(+) IFN-γ(+) cells. No infectious virus was recovered from a lung homogenate following the challenge. A Histological examination of the lungs revealed a significant reduction in inflammatory reactions without alveolar damage. These results support the recombinant measles viruses being effective vaccine candidates against RSV that induce RSV-specific CTL responses with or without the development of an antibody response.

  18. Palivizumab prophylaxis of respiratory syncytial virus disease in 2000-2001: results from The Palivizumab Outcomes Registry.

    PubMed

    Parnes, Curt; Guillermin, Judith; Habersang, Rolf; Nicholes, Peggy; Chawla, Vijay; Kelly, Tammy; Fishbein, Judith; McRae, Patty; Goessler, Mary; Gatti, Antoinette; Calcagno, John A; Eki, Cheryl; Harris, Kristen A; Joyave, Joseph; McFarland, Kathy; Protter, Paul; Sullivan, Mary; Stanford, Allan; Lovett, Nancy; Ortiz, Marisol; Rojas, Sharon; Cyrus, Scott; Cyrus, Janell; Cohen, Stuart; Buchin, Debbie; Riordan, Linda; Zuniga, Monica; Shah, Rupa; Minard, Carmen; Quintin, Arden; Douglas, Glenda; van Houten, John; Freutner, Sharyn; Chartrand, Stephen; Nowatzke, Patsy; Romero, Jose; Rhodes, Torunn; Benoit, Michelle; Walter, Emmanuel; Walker, Leslie; DeBonnett, Laurie; Cross, Mia; Free, Teresa; Martin, Sharman; Shank, Karen; Guedes, Ben; Atkinson, Lee Ann; Halpin, George J; Rouse, Kathy; Hand, Ivan; Geiss, Donna; Marshall, James R; Burleson, Lois; Boland, Jim; Seybold, Kelsey; Hunter, Vicki; Unfer, Susan; Schmucker, Jackie; Gley, Margaret; Marcus, Michael; Thompson, Patricia; Milla, Paulino; Young, Connie; Zanni, Robert; Zinno, Virginia; Fetter-Zarzeka, Alexandra; Busey, Amanda; Sokunbi, Modupe A; Airington, Sherrie; Richard, Nancy; Muraligopal, Vellore; Lewis, Stephanie; Weber, F Thomas; Giordano, Beverly P; Linehan, Denise; Roach, Jane; Davis, Randle; Rzepka, Andrew A; Booth, Teri; Smeltzer, David; Walsh, Jeanne; Arispe, Emilio; Rowley, Rhonda; Bolling, Christopher; Botts, Tanya; Haskett, Kateri; Raby, Deana; Batiz, Evelyn; Gelfand, Andrew; Farrell, Lynn; Butler, Stephen; Colby, Linda; Schochet, Peter; Bentler, Julie; Hirsch, David; Wilkinson, Lisa; Aaronson, Allen; Bennett, Eleanora; Wingate, Julie; Quinn, Dawn; Komendowski, Katherine; Deckard, Marcia; Frogel, Michael; Nerwen, Cliff; Copenhaver, Steven; Prater, Michele; Wolsztein, Jacob; Mackey, Kristine; Benbow, Marshall; Naranjo, Marisela; Hensley, Sandra; Hayes, Cindy; Sadeghi, Hossein; Lawson, Sally May; McCall, Mark; Combs, Karla; Ledbetter, Joel; Sarnosky, Karen; Swafford, Cathy; Speer, Michael; Barton, Wendy J; Mink, J W; Lemm, Dianne; Hudak, Mark; Case, Elizabeth; Rowen, Judith; Fuentes, Sandra; Pane, Carly; Richardson, Leslie; Chavarria, Cesar; Cassino, Deanne; Ghaffari, Kourosh; Carroll, Carol; Lee, Haesoon; Guclu, Lydia; Johnson, Christopher; Blum, Valerie; Boron, Marnie L; Sorrentino, Mark; Hirsch, Robert L; Van Veldhuisen, Paul C; Smith, Carol

    2003-06-01

    The objective of the Registry was to characterize the population of infants receiving prophylaxis for respiratory syncytial virus (RSV) disease by describing the patterns and scope of usage of palivizumab in a cross section of US infants. RSV hospitalization outcomes were also described. The Palivizumab (Synagis, MedImmune, Inc., 25 West Watkins Mill Road, Gaithersburg, MD 20878) Outcomes Registry was a prospective multicenter survey conducted at 63 sites. Demographics, injection history, and RSV hospitalization outcomes were collected on 2,116 infants receiving palivizumab. Infants were enrolled in the Registry between September 1, 2000-March 1, 2001, at the time of their first injection. Infants born at less than 32 weeks of gestation accounted for 47% of infants enrolled, and those between 32-35 weeks accounted for 45%; approximately 8% were greater than 35 weeks of gestation. Lower RSV hospitalization rates were observed in infants who had greater adherence to regularly scheduled injections. Nearly one-half of all hospitalizations occurred within the first and second injection intervals, suggesting the importance of early RSV protection. The confirmed RSV hospitalization rate of all infants in the Registry was 2.9%; the rate was 5.8% in infants with chronic lung disease of infancy, and 2.1% in premature infants without chronic lung disease. In conclusion, these data support the continued effectiveness of palivizumab prophylaxis for severe RSV lower respiratory tract disease in a large cohort of high-risk infants from geographically diverse pediatric offices and clinics. The Palivizumab Outcomes Registry provides an opportunity to assess palivizumab utilization and clinical effectiveness in the US. PMID:12746948

  19. Antiviral Efficacy of a Respiratory Syncytial Virus (RSV) Fusion Inhibitor in a Bovine Model of RSV Infection

    PubMed Central

    Jordan, Robert; Shao, Matt; Mackman, Richard L.; Perron, Michel; Cihlar, Tomas; Lewis, Sandy A.; Eisenberg, Eugene J.; Carey, Anne; Strickley, Robert G.; Chien, Jason W.; Anderson, Mark L.; McEligot, Heather A.; Behrens, Nicole E.

    2015-01-01

    Respiratory syncytial virus (RSV) is the leading cause of bronchiolitis and pneumonia in infants. Effective treatment for RSV infection is a significant unmet medical need. While new RSV therapeutics are now in development, there are very few animal models that mimic the pathogenesis of human RSV, making it difficult to evaluate new disease interventions. Experimental infection of Holstein calves with bovine RSV (bRSV) causes a severe respiratory infection that is similar to human RSV infection, providing a relevant model for testing novel therapeutic agents. In this model, viral load is readily detected in nasal secretions by quantitative real-time PCR (qRT-PCR), and cumulative symptom scoring together with histopathology evaluations of infected tissue allow for the assessment of disease severity. The bovine RSV model was used to evaluate the antiviral activity of an RSV fusion inhibitor, GS1, which blocks virus entry by inhibiting the fusion of the viral envelope with the host cell membrane. The efficacy of GS1, a close structural analog of GS-5806 that is being developed to treat RSV infection in humans was evaluated in two randomized, blind, placebo-controlled studies in bRSV-infected calves. Intravenous administration of GS1 at 4 mg/kg of body weight/day for 7 days starting 24 h or 72 h postinoculation provided clear therapeutic benefit by reducing the viral load, disease symptom score, respiration rate, and lung pathology associated with bRSV infection. These data support the use of the bovine RSV model for evaluation of experimental therapeutics for treatment of RSV. PMID:26055364

  20. Whole Genome Sequencing and Evolutionary Analysis of Human Respiratory Syncytial Virus A and B from Milwaukee, WI 1998-2010

    PubMed Central

    Rebuffo-Scheer, Cecilia; Bose, Michael; He, Jie; Khaja, Shamim; Ulatowski, Michael; Beck, Eric T.; Fan, Jiang; Kumar, Swati; Nelson, Martha I.; Henrickson, Kelly J.

    2011-01-01

    Background Respiratory Syncytial Virus (RSV) is the leading cause of lower respiratory-tract infections in infants and young children worldwide. Despite this, only six complete genome sequences of original strains have been previously published, the most recent of which dates back 35 and 26 years for RSV group A and group B respectively. Methodology/Principal Findings We present a semi-automated sequencing method allowing for the sequencing of four RSV whole genomes simultaneously. We were able to sequence the complete coding sequences of 13 RSV A and 4 RSV B strains from Milwaukee collected from 1998–2010. Another 12 RSV A and 5 RSV B strains sequenced in this study cover the majority of the genome. All RSV A and RSV B sequences were analyzed by neighbor-joining, maximum parsimony and Bayesian phylogeny methods. Genetic diversity was high among RSV A viruses in Milwaukee including the circulation of multiple genotypes (GA1, GA2, GA5, GA7) with GA2 persisting throughout the 13 years of the study. However, RSV B genomes showed little variation with all belonging to the BA genotype. For RSV A, the same evolutionary patterns and clades were seen consistently across the whole genome including all intergenic, coding, and non-coding regions sequences. Conclusions/Significance The sequencing strategy presented in this work allows for RSV A and B genomes to be sequenced simultaneously in two working days and with a low cost. We have significantly increased the amount of genomic data that is available for both RSV A and B, providing the basic molecular characteristics of RSV strains circulating in Milwaukee over the last 13 years. This information can be used for comparative analysis with strains circulating in other communities around the world which should also help with the development of new strategies for control of RSV, specifically vaccine development and improvement of RSV diagnostics. PMID:21998661

  1. GS-5806 Inhibits a Broad Range of Respiratory Syncytial Virus Clinical Isolates by Blocking the Virus-Cell Fusion Process

    PubMed Central

    Stray, Kirsten; Kinkade, April; Theodore, Dorothy; Lee, Gary; Eisenberg, Eugene; Sangi, Michael; Gilbert, Brian E.; Jordan, Robert; Piedra, Pedro A.; Toms, Geoffery L.; Mackman, Richard; Cihlar, Tomas

    2015-01-01

    Respiratory syncytial virus (RSV) is a leading cause of lower respiratory tract infections in infants and young children. In addition, RSV causes significant morbidity and mortality in hospitalized elderly and immunocompromised patients. Currently, only palivizumab, a monoclonal antibody against the RSV fusion (F) protein, and inhaled ribavirin are approved for the prophylactic and therapeutic treatment of RSV, respectively. Therefore, there is a clinical need for safe and effective therapeutic agents for RSV infections. GS-5806, discovered via chemical optimization of a hit from a high-throughput antiviral-screening campaign, selectively inhibits a diverse set of 75 RSV subtype A and B clinical isolates (mean 50% effective concentration [EC50] = 0.43 nM). The compound maintained potency in primary human airway epithelial cells and exhibited low cytotoxicity in human cell lines and primary cell cultures (selectivity > 23,000-fold). Time-of-addition and temperature shift studies demonstrated that GS-5806 does not block RSV attachment to cells but interferes with virus entry. Follow-up experiments showed potent inhibition of RSV F-mediated cell-to-cell fusion. RSV A and B variants resistant to GS-5806, due to mutations in F protein (RSV A, L138F or F140L/N517I, and RSV B, F488L or F488S), were isolated and showed cross-resistance to other RSV fusion inhibitors, such as VP-14637, but remained fully sensitive to palivizumab and ribavirin. In summary, GS-5806 is a potent and selective RSV fusion inhibitor with antiviral activity against a diverse set of RSV clinical isolates. The compound is currently under clinical investigation for the treatment of RSV infection in pediatric, immunocompromised, and elderly patients. PMID:26666922

  2. Design and characterization of epitope-scaffold immunogens that present the motavizumab epitope from respiratory syncytial virus.

    PubMed

    McLellan, Jason S; Correia, Bruno E; Chen, Man; Yang, Yongping; Graham, Barney S; Schief, William R; Kwong, Peter D

    2011-06-24

    Respiratory syncytial virus (RSV) is a major cause of respiratory tract infections in infants, but an effective vaccine has not yet been developed. An ideal vaccine would elicit protective antibodies while avoiding virus-specific T-cell responses, which have been implicated in vaccine-enhanced disease with previous RSV vaccines. We propose that heterologous proteins designed to present RSV-neutralizing antibody epitopes and to elicit cognate antibodies have the potential to fulfill these vaccine requirements, as they can be fashioned to be free of viral T-cell epitopes. Here we present the design and characterization of three epitope-scaffolds that present the epitope of motavizumab, a potent neutralizing antibody that binds to a helix-loop-helix motif in the RSV fusion glycoprotein. Two of the epitope-scaffolds could be purified, and one epitope-scaffold based on a Staphylococcus aureus protein A domain bound motavizumab with kinetic and thermodynamic properties consistent with the free epitope-scaffold being stabilized in a conformation that closely resembled the motavizumab-bound state. This epitope-scaffold was well folded as assessed by circular dichroism and isothermal titration calorimetry, and its crystal structure (determined in complex with motavizumab to 1.9 Å resolution) was similar to the computationally designed model, with all hydrogen-bond interactions critical for binding to motavizumab preserved. Immunization of mice with this epitope-scaffold failed to elicit neutralizing antibodies but did elicit sera with F binding activity. The elicitation of F binding antibodies suggests that some of the design criteria for eliciting protective antibodies without virus-specific T-cell responses are being met, but additional optimization of these novel immunogens is required.

  3. Novel Respiratory Syncytial Virus-Like Particle Vaccine Composed of the Postfusion and Prefusion Conformations of the F Glycoprotein

    PubMed Central

    Boigard, Hélène; Bhatia, Bipin; Fallon, John T.; Alimova, Alexandra; Gottlieb, Paul

    2016-01-01

    Respiratory syncytial virus (RSV) is the leading cause of severe respiratory disease in infants and children and represents an important global health burden for the elderly and the immunocompromised. Despite decades of research efforts, no licensed vaccine for RSV is available. We have developed virus-like particle (VLP)-based RSV vaccines assembled with the human metapneumovirus (hMPV) matrix protein (M) as the structural scaffold and the RSV fusion glycoprotein (F) in either the postfusion or prefusion conformation as its prime surface immunogen. Vaccines were composed of postfusion F, prefusion F, or a combination of the two conformations and formulated with a squalene-based oil emulsion as adjuvant. Immunization with these VLP vaccines afforded full protection against RSV infection and prevented detectable viral replication in the mouse lung after challenge. Analyses of lung cytokines and chemokines showed that VLP vaccination mostly induced the production of gamma interferon (IFN-γ), a marker of the Th1-mediated immune response, which is predominantly required for viral protection. Conversely, immunization with a formalin-inactivated RSV (FI-RSV) vaccine induced high levels of inflammatory chemokines and cytokines of the Th2- and Th17-mediated types of immune responses, as well as severe lung inflammation and histopathology. The VLP vaccines showed restricted production of these immune mediators and did not induce severe bronchiolitis or perivascular infiltration as seen with the FI-RSV vaccine. Remarkably, analysis of the serum from immunized mice showed that the VLP vaccine formulated using a combination of postfusion and prefusion F elicited the highest level of neutralizing antibody and enhanced the Th1-mediated immune response. PMID:27030590

  4. Epidemiology and Clinical Presentations of Respiratory Syncytial Virus Subgroups A and B Detected with Multiplex Real-Time PCR

    PubMed Central

    Liu, Wenkuan; Chen, Dehui; Tan, Weiping; Xu, Duo; Qiu, Shuyan; Zeng, Zhiqi; Li, Xiao; Zhou, Rong

    2016-01-01

    Respiratory syncytial virus (RSV) is one of the most important pathogenic infections of children and requires in-depth research worldwide, and especially in developing countries. We used a novel multiplex real-time PCR to test 5483 patients (≤ 14 years old) hospitalized with respiratory illness in Guangzhou, China, over a 3-year period. Of these patients, 729 were positive for RSV-A (51.2%, 373/729) or RSV-B (48.8%, 356/729), but none was infected with both viruses. Two seasonal peaks in total RSV were detected at the changes from winter to spring and from summer to autumn. RSV-B was dominant in 2013 and RSV-A in 2015, whereas RSV-A and RSV-B cocirculated in 2014. The clinical presentations of 645 RSV-positive patients were analyzed. Bronchiolitis, dyspnea, coryza, vomiting, poor appetite, and diarrhea occurred more frequently in RSV-A-positive than RSV-B-positive patients, whereas chill, headache, myalgia, debility, and rash etc. were more frequent in RSV-B-positive than RSV-A-positive patients, suggesting specific clinical characteristics for RSV-A and RSV-B. Coinfectons with other pathogens were common and diverse. Bronchiolitis, fever (≥ 38°C), and poor appetite were more frequent in patients with single RSV infections than in coinfected patients, suggesting the key pathogenic activity of RSV. Analysis of the relationships between the comparative viral load and clinical presentations showed significant differences in bronchiolitis, fever (≥ 38°C), and rash etc. among patients with different viral loads. This study provides a novel rapid method for detecting RSV subgroups, and provides new insights into the epidemiology and clinical implications of RSV. PMID:27764220

  5. Design and Characterization of Epitope-Scaffold Immunogens That Present the Motavizumab Epitope from Respiratory Syncytial Virus

    SciTech Connect

    McLellan, Jason S.; Correia, Bruno E.; Chen, Man; Yang, Yongping; Graham, Barney S.; Schief, William R.; Kwong, Peter D.

    2012-06-28

    Respiratory syncytial virus (RSV) is a major cause of respiratory tract infections in infants, but an effective vaccine has not yet been developed. An ideal vaccine would elicit protective antibodies while avoiding virus-specific T-cell responses, which have been implicated in vaccine-enhanced disease with previous RSV vaccines. We propose that heterologous proteins designed to present RSV-neutralizing antibody epitopes and to elicit cognate antibodies have the potential to fulfill these vaccine requirements, as they can be fashioned to be free of viral T-cell epitopes. Here we present the design and characterization of three epitope-scaffolds that present the epitope of motavizumab, a potent neutralizing antibody that binds to a helix-loop-helix motif in the RSV fusion glycoprotein. Two of the epitope-scaffolds could be purified, and one epitope-scaffold based on a Staphylococcus aureus protein A domain bound motavizumab with kinetic and thermodynamic properties consistent with the free epitope-scaffold being stabilized in a conformation that closely resembled the motavizumab-bound state. This epitope-scaffold was well folded as assessed by circular dichroism and isothermal titration calorimetry, and its crystal structure (determined in complex with motavizumab to 1.9 {angstrom} resolution) was similar to the computationally designed model, with all hydrogen-bond interactions critical for binding to motavizumab preserved. Immunization of mice with this epitope-scaffold failed to elicit neutralizing antibodies but did elicit sera with F binding activity. The elicitation of F binding antibodies suggests that some of the design criteria for eliciting protective antibodies without virus-specific T-cell responses are being met, but additional optimization of these novel immunogens is required.

  6. EGFR Interacts with the Fusion Protein of Respiratory Syncytial Virus Strain 2-20 and Mediates Infection and Mucin Expression

    PubMed Central

    Stobart, Christopher C.; Hotard, Anne L.; Villenave, Remi; Meng, Jia; Pretto, Carla D.; Shields, Michael D.; Nguyen, Minh Trang; Todd, Sean O.; Chi, Michael H.; Hammonds, Jason; Krumm, Stefanie A.; Spearman, Paul; Plemper, Richard K.; Sakamoto, Kaori; Peebles, R. Stokes; Power, Ultan F.; Moore, Martin L.

    2016-01-01

    Respiratory syncytial virus (RSV) is the major cause of viral lower respiratory tract illness in children. In contrast to the RSV prototypic strain A2, clinical isolate RSV 2–20 induces airway mucin expression in mice, a clinically relevant phenotype dependent on the fusion (F) protein of the RSV strain. Epidermal growth factor receptor (EGFR) plays a role in airway mucin expression in other systems; therefore, we hypothesized that the RSV 2–20 F protein stimulates EGFR signaling. Infection of cells with chimeric strains RSV A2-2-20F and A2-2-20GF or over-expression of 2–20 F protein resulted in greater phosphorylation of EGFR than infection with RSV A2 or over-expression of A2 F, respectively. Chemical inhibition of EGFR signaling or knockdown of EGFR resulted in diminished infectivity of RSV A2-2-20F but not RSV A2. Over-expression of EGFR enhanced the fusion activity of 2–20 F protein in trans. EGFR co-immunoprecipitated most efficiently with RSV F proteins derived from “mucogenic” strains. RSV 2–20 F and EGFR co-localized in H292 cells, and A2-2-20GF-induced MUC5AC expression was ablated by EGFR inhibitors in these cells. Treatment of BALB/c mice with the EGFR inhibitor erlotinib significantly reduced the amount of RSV A2-2-20F-induced airway mucin expression. Our results demonstrate that RSV F interacts with EGFR in a strain-specific manner, EGFR is a co-factor for infection, and EGFR plays a role in RSV-induced mucin expression, suggesting EGFR is a potential target for RSV disease. PMID:27152417

  7. Respiratory Syncytial Virus Uses CX3CR1 as a Receptor on Primary Human Airway Epithelial Cultures

    PubMed Central

    Johnson, Sara M.; McNally, Beth A.; Ioannidis, Ioannis; Flano, Emilio; Teng, Michael N.; Oomens, Antonius G.; Walsh, Edward E.; Peeples, Mark E.

    2015-01-01

    Respiratory syncytial virus (RSV) is the most frequent cause of lower respiratory disease in infants, but no vaccine or effective therapy is available. The initiation of RSV infection of immortalized cells is largely dependent on cell surface heparan sulfate (HS), a receptor for the RSV attachment (G) glycoprotein in immortalized cells. However, RSV infects the ciliated cells in primary well differentiated human airway epithelial (HAE) cultures via the apical surface, but HS is not detectable on this surface. Here we show that soluble HS inhibits infection of immortalized cells, but not HAE cultures, confirming that HS is not the receptor on HAE cultures. Conversely, a “non-neutralizing” monoclonal antibody against the G protein that does not block RSV infection of immortalized cells, does inhibit infection of HAE cultures. This antibody was previously shown to block the interaction between the G protein and the chemokine receptor CX3CR1 and we have mapped the binding site for this antibody to the CX3C motif and its surrounding region in the G protein. We show that CX3CR1 is present on the apical surface of ciliated cells in HAE cultures and especially on the cilia. RSV infection of HAE cultures is reduced by an antibody against CX3CR1 and by mutations in the G protein CX3C motif. Additionally, mice lacking CX3CR1 are less susceptible to RSV infection. These findings demonstrate that RSV uses CX3CR1 as a cellular receptor on HAE cultures and highlight the importance of using a physiologically relevant model to study virus entry and antibody neutralization. PMID:26658574

  8. A duplex recombinant viral nucleoprotein microbead immunoassay for simultaneous detection of seroresponses to human respiratory syncytial virus and metapneumovirus infections.

    PubMed

    Zhang, Yange; Brooks, W Abdullah; Goswami, Doli; Rahman, Mustafizur; Luby, Stephen P; Erdman, Dean D

    2014-09-01

    Serologic diagnosis of human respiratory syncytial virus (hRSV) and human metapneumovirus (hMPV) infections has been shown to complement virus detection methods in epidemiologic studies. Enzyme immunoassays (EIAs) using cultured virus lysate antigens are often used to diagnose infection by demonstration of a ≥4-fold rises in antibody titer between acute and convalescent serum pairs. In this study, hRSV and hMPV nucleocapsid (recN) proteins were expressed in a baculovirus system and their performance compared with virus culture lysate antigen in EIAs using paired serum specimens collected from symptomatic children. The recN proteins were also used to develop a duplex assay based on the Luminex microbead-based suspension array technology, where diagnostic rises in antibody levels could be determined simultaneously at a single serum dilution. Antibody levels measured by the recN and viral lysate EIAs correlated moderately (hRSV, r(2)=0.72; hMPV, r(2)=0.76); the recN EIAs identified correctly 35 of 37 (94.6%) and 48 of 50 (96%) serum pairs showing diagnostic antibody rises by viral lysate EIAs. Purified recN proteins were then coupled to microbeads and serum pairs were tested at a single dilution on a Luminex MAGPIX(®) analyzer. The duplex recN assay identified correctly 33 of 39 (85%) and 41 of 47 (86.7%) serum pairs showing diagnostic rises to hRSV and hMPV, respectively. The recN assay permits simultaneous testing for acute hRSV and hMPV infections and offers a platform for expanded multiplexing of other respiratory virus assays.

  9. EGFR Interacts with the Fusion Protein of Respiratory Syncytial Virus Strain 2-20 and Mediates Infection and Mucin Expression.

    PubMed

    Currier, Michael G; Lee, Sujin; Stobart, Christopher C; Hotard, Anne L; Villenave, Remi; Meng, Jia; Pretto, Carla D; Shields, Michael D; Nguyen, Minh Trang; Todd, Sean O; Chi, Michael H; Hammonds, Jason; Krumm, Stefanie A; Spearman, Paul; Plemper, Richard K; Sakamoto, Kaori; Peebles, R Stokes; Power, Ultan F; Moore, Martin L

    2016-05-01

    Respiratory syncytial virus (RSV) is the major cause of viral lower respiratory tract illness in children. In contrast to the RSV prototypic strain A2, clinical isolate RSV 2-20 induces airway mucin expression in mice, a clinically relevant phenotype dependent on the fusion (F) protein of the RSV strain. Epidermal growth factor receptor (EGFR) plays a role in airway mucin expression in other systems; therefore, we hypothesized that the RSV 2-20 F protein stimulates EGFR signaling. Infection of cells with chimeric strains RSV A2-2-20F and A2-2-20GF or over-expression of 2-20 F protein resulted in greater phosphorylation of EGFR than infection with RSV A2 or over-expression of A2 F, respectively. Chemical inhibition of EGFR signaling or knockdown of EGFR resulted in diminished infectivity of RSV A2-2-20F but not RSV A2. Over-expression of EGFR enhanced the fusion activity of 2-20 F protein in trans. EGFR co-immunoprecipitated most efficiently with RSV F proteins derived from "mucogenic" strains. RSV 2-20 F and EGFR co-localized in H292 cells, and A2-2-20GF-induced MUC5AC expression was ablated by EGFR inhibitors in these cells. Treatment of BALB/c mice with the EGFR inhibitor erlotinib significantly reduced the amount of RSV A2-2-20F-induced airway mucin expression. Our results demonstrate that RSV F interacts with EGFR in a strain-specific manner, EGFR is a co-factor for infection, and EGFR plays a role in RSV-induced mucin expression, suggesting EGFR is a potential target for RSV disease. PMID:27152417

  10. Inducible Expression of Inflammatory Chemokines in Respiratory Syncytial Virus-Infected Mice: Role of MIP-1α in Lung Pathology

    PubMed Central

    Haeberle, Helene A.; Kuziel, William A.; Dieterich, Hans-Juergen; Casola, Antonella; Gatalica, Zoran; Garofalo, Roberto P.

    2001-01-01

    Lower respiratory tract disease caused by respiratory syncytial virus (RSV) is characterized by profound airway mucosa inflammation, both in infants with naturally acquired infection and in experimentally inoculated animal models. Chemokines are central regulatory molecules in inflammatory, immune, and infectious processes of the lung. In this study, we demonstrate that intranasal infection of BALB/c mice with RSV A results in inducible expression of lung chemokines belonging to the CXC (MIP-2 and IP-10), CC (RANTES, eotaxin, MIP-1β, MIP-1α, MCP-1, TCA-3) and C (lymphotactin) families. Chemokine mRNA expression occurred as early as 24 h following inoculation and persisted for at least 5 days in mice inoculated with the highest dose of virus (107 PFU). In general, levels of chemokine mRNA and protein were dependent on the dose of RSV inoculum and paralleled the intensity of lung cellular inflammation. Immunohisthochemical studies indicated that RSV-induced expression of MIP-1α, one of the most abundantly expressed chemokines, was primarily localized in epithelial cells of the alveoli and bronchioles, as well as in adjoining capillary endothelium. Genetically altered mice with a selective deletion of the MIP-1α gene (−/− mice) demonstrated a significant reduction in lung inflammation following RSV infection, compared to control littermates (+/+ mice). Despite the paucity of infiltrating cells, the peak RSV titer in the lung of −/− mice was not significantly different from that observed in +/+ mice. These results provide the first direct evidence that RSV infection may induce lung inflammation via the early production of inflammatory chemokines. PMID:11134301

  11. Viral-mediated Inhibition of Antioxidant Enzymes Contributes to the Pathogenesis of Severe Respiratory Syncytial Virus Bronchiolitis

    PubMed Central

    Hosakote, Yashoda M.; Jantzi, Paul D.; Esham, Dana L.; Spratt, Heidi; Kurosky, Alexander; Casola, Antonella; Garofalo, Roberto P.

    2011-01-01

    Rationale: Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infections in children, for which no specific treatment or vaccine is currently available. We have previously shown that RSV induces reactive oxygen species in cultured cells and oxidative injury in the lungs of experimentally infected mice. The mechanism(s) of RSV-induced oxidative stress in vivo is not known. Objectives: To measure changes of lung antioxidant enzymes expression/activity and activation of NF-E2-related factor 2 (Nrf2), a transcription factor that regulates detoxifying and antioxidant enzyme gene expression, in mice and in infants with naturally acquired RSV infection. Methods: Superoxide dismutase 1 (SOD 1), SOD 2, SOD 3, catalase, glutathione peroxidase, and glutathione S-transferase, as well as Nrf2 expression, were measured in murine bronchoalveolar lavage, cell extracts of conductive airways, and/or in human nasopharyngeal secretions by Western blot and two-dimensional gel electrophoresis. Antioxidant enzyme activity and markers of oxidative cell injury were measured in either murine bronchoalveolar lavage or nasopharyngeal secretions by colorimetric/immunoassays. Measurements and Main Results: RSV infection induced a significant decrease in the expression and/or activity of SOD, catalase, glutathione S-transferase, and glutathione peroxidase in murine lungs and in the airways of children with severe bronchiolitis. Markers of oxidative damage correlated with severity of clinical illness in RSV-infected infants. Nrf2 expression was also significantly reduced in the lungs of viral-infected mice. Conclusions: RSV infection induces significant down-regulation of the airway antioxidant system in vivo, likely resulting in lung oxidative damage. Modulation of oxidative stress may pave the way toward important advances in the therapeutic approach of RSV-induced acute lung disease. PMID:21471094

  12. Chimpanzee adenovirus and MVA-vectored respiratory syncytial virus vaccine is safe and expands humoral and cellular immunity in adults

    PubMed Central

    Green, CA; Scarselli, E; Sande, CJ; Thompson, AJ; de Lara, CM; Taylor, K; Haworth, K; Del Sorbo, M; Angus, B; Siani, L; Di Marco, S; Traboni, C; Folgori, A; Colloca, S; Capone, S; Vitelli, A; Cortese, R; Klenerman, P; Nicosia, A; Pollard, AJ

    2015-01-01

    Respiratory syncytial virus (RSV) causes respiratory infection in annual epidemics, with infants and the elderly at particular risk of developing severe disease and death. However, despite its importance, no vaccine exists. The chimpanzee adenovirus, PanAd3-RSV, and modified vaccinia virus Ankara, MVA-RSV, are replication defective viral vectors encoding the RSV proteins F, N and M2-1 for the induction of humoral and cellular responses. We performed an open-label, dose-escalation, phase 1 clinical trial in 42 healthy adults in which four different combinations of prime/boost vaccinations were investigated for safety and immunogenicity, including both intra-muscular and intra-nasal administration of the adenoviral vectored vaccine. The vaccines were safe and well tolerated, with the most common reported adverse events being mild injection site reactions. No vaccine-related serious adverse events occurred. RSV neutralising antibody titres rose in response to intramuscular (IM) prime with PanAd3-RSV, and after IM boost for individuals primed by the intra-nasal (IN) route. Circulating anti-F IgG and IgA antibody secreting cells (ASCs) were observed after IM prime and IM boost. RSV-specific T-cell responses were increased after IM PanAd3-RSV prime and were most efficiently boosted by IM MVA-RSV. IFNγ secretion after boost was from both CD4+ and CD8+ T-cells, without detectable Th2 cytokines that have been previously associated with immune pathogenesis following exposure to RSV after formalin inactivated RSV vaccine. In conclusion, PanAd3-RSV and MVA-RSV are safe and immunogenic in healthy adults. These vaccine candidates warrant further clinical evaluation of efficacy to assess their potential to reduce the burden of RSV disease. PMID:26268313

  13. Vaccine Induced Herd Immunity for Control of Respiratory Syncytial Virus Disease in a Low-Income Country Setting

    PubMed Central

    Kinyanjui, Timothy M.; House, Thomas A.; Kiti, Moses C.; Cane, Patricia A.; Nokes, David J.; Medley, Graham F.

    2015-01-01

    Background Respiratory syncytial virus (RSV) is globally ubiquitous, and infection during the first six months of life is a major risk for severe disease and hospital admission; consequently RSV is the most important viral cause of respiratory morbidity and mortality in young children. Development of vaccines for young infants is complicated by the presence of maternal antibodies and immunological immaturity, but vaccines targeted at older children avoid these problems. Vaccine development for young infants has been unsuccessful, but this is not the case for older children (> 6m). Would vaccinating older children have a significant public health impact? We developed a mathematical model to explore the benefits of a vaccine against RSV. Methods and Findings We have used a deterministic age structured model capturing the key epidemiological characteristics of RSV and performed a statistical maximum-likelihood fit to age-specific hospitalization data from a developing country setting. To explore the effects of vaccination under different mixing assumptions, we included two versions of contact matrices: one from a social contact diary study, and the second a synthesised construction based on demographic data. Vaccination is assumed to elicit an immune response equivalent to primary infection. Our results show that immunisation of young children (5–10m) is likely to be a highly effective method of protection of infants (<6m) against hospitalisation. The majority benefit is derived from indirect protection (herd immunity). A full sensitivity and uncertainty analysis using Latin Hypercube Sampling of the parameter space shows that our results are robust to model structure and model parameters. Conclusions This result suggests that vaccinating older infants and children against RSV can have a major public health benefit. PMID:26390032

  14. Detection of respiratory syncytial virus by reverse transcription-PCR and hybridization with a DNA enzyme immunoassay.

    PubMed Central

    Freymuth, F; Eugene, G; Vabret, A; Petitjean, J; Gennetay, E; Brouard, J; Duhamel, J F; Guillois, B

    1995-01-01

    Nasal aspirates from 238 infants hospitalized with acute respiratory infections during the winter of 1994 and 1995 were tested for respiratory syncytial virus (RSV) by immunofluorescence assay (IFA) and the viral isolation technique (VIT) and by two PCR and hybridization methods: reverse transcription PCR 1 (RT-PCR1), which amplifies the RNAs of all RSV strains, and RT-PCR-2, which allows subgroup classification of RSV. RT-PCR-1 and RT-PCR-2 detected viral sequences in 56.7% (135 of 238) and 48.3% (115 of 238) of the samples, respectively, while only 80 (33.6%) samples were found to be positive by IFA and VIT. Of the PCR-positive specimens, 57 were missed by these routine techniques in RT-PCR-1 and 45 were missed in RT-PCR-2. Although the RSV-PCR-1 and RSV-PCR-2 techniques amplified two different sequences of the RSV genome, they gave similar results for 218 (91.6%) nasal aspirates. Compared with conventional methods, the sensitivity, specificity, and agreement were 97.5, 63.9, and 75.2%, respectively, for RT-PCR-1 and 89.7, 71.9, and 77.7%, respectively, for RT-PCR-2, and for these two RT-PCR assays, the positive predictive value (PPV) and the index of agreement (kappa) were comparable and moderate, respectively: PPV was 57.8% and kappa was 0.52 in RT-PCR-1, and PPV was 60.9% and kappa was 0.54 in RT-PCR-2. However, there was a perfect correlation between the two RT-PCRs, with a PPV of 100% and an excellent index of agreement (kappa = 0.88). Therefore, most RT-PCR results were really true positive, and VIT and IFA, which missed some of them, appeared to be less sensitive. PMID:8586738

  15. The Respiratory Syncytial Virus G Protein Conserved Domain Induces a Persistent and Protective Antibody Response in Rodents

    PubMed Central

    Nguyen, Thien N.; Power, Ultan F.; Robert, Alain; Haeuw, Jean-François; Helffer, Katia; Perez, Amadeo; Asin, Miguel-Angel; Corvaia, Nathalie; Libon, Christine

    2012-01-01

    Respiratory syncytial virus (RSV) is an important cause of severe upper and lower respiratory disease in infants and in the elderly. There are 2 main RSV subtypes A and B. A recombinant vaccine was designed based on the central domain of the RSV-A attachment G protein which we had previously named G2Na (aa130–230). Here we evaluated immunogenicity, persistence of antibody (Ab) response and protective efficacy induced in rodents by: (i) G2Na fused to DT (Diphtheria toxin) fragments in cotton rats. DT fusion did not potentiate neutralizing Ab responses against RSV-A or cross-reactivity to RSV-B. (ii) G2Nb (aa130–230 of the RSV-B G protein) either fused to, or admixed with G2Na. G2Nb did not induce RSV-B-reactive Ab responses. (iii) G2Na at low doses. Two injections of 3 µg G2Na in Alum were sufficient to induce protective immune responses in mouse lungs, preventing RSV-A and greatly reducing RSV-B infections. In cotton rats, G2Na-induced RSV-reactive Ab and protective immunity against RSV-A challenge that persisted for at least 24 weeks. (iv) injecting RSV primed mice with a single dose of G2Na/Alum or G2Na/PLGA [poly(D,L-lactide-co-glycolide]. Despite the presence of pre-existing RSV-specific Abs, these formulations effectively boosted anti-RSV Ab titres and increased Ab titres persisted for at least 21 weeks. Affinity maturation of these Abs increased from day 28 to day 148. These data indicate that G2Na has potential as a component of an RSV vaccine formulation. PMID:22479601

  16. Characterization of hospital and community-acquired respiratory syncytial virus in children with severe lower respiratory tract infections in Ho Chi Minh City, Vietnam, 2010

    PubMed Central

    Tuan, Tran Anh; Thanh, Tran Tan; Hai, Nguyen thi Thanh; Tinh, Le Binh Bao; Kim, Le thi Ngoc; Do, Lien Anh Ha; Chinh B'Krong, Nguyen thi Thuy; Tham, Nguyen thi; Hang, Vu thi Ty; Merson, Laura; Farrar, Jeremy; Thuong, Tang Chi; de Jong, Menno D; Schultsz, Constance; van Doorn, H Rogier

    2015-01-01

    Background Human respiratory syncytial virus (RSV) is an important community and nosocomial pathogen in developed countries but data regarding the importance of RSV in developing countries are relatively scarce. Methods During a 1-year surveillance study in 2010, we took serial samples from children admitted to the Emergency Unit of the Respiratory Ward of Children's Hospital 1 in Ho Chi Minh City, Vietnam. RSV was detected within 72 hours of admission to the ward in 26% (376/1439; RSV A: n = 320; RSV B: n = 54; and RSV A and B: n = 2). Among those negative in the first 72 hours after admission, 6·6% (25/377) acquired nosocomial RSV infection during hospitalization (RSV A: n = 22; and RSV B: n = 3). Results Children with nosocomial RSV infection were younger (P = 0·001) and had a longer duration of hospitalization (P < 0·001). The rate of incomplete recovery among children with nosocomial RSV infection was significantly higher than among those without (P < 0·001). Phylogenetic analysis of partial G gene sequences obtained from 79% (316/401) of positive specimens revealed the co-circulation of multiple genotypes with RSV A NA1 being predominant (A NA1: n = 275; A GA5: n = 5; B BA3: n = 3; B BA9: n = 26; and B BA10: n = 7). The RSV A GA5 and RSV B BA3 genotypes have not been reported from Vietnam, previously. Conclusion Besides emphasizing the importance of RSV as a cause of respiratory infection leading to hospitalization in young children and as a nosocomial pathogen, data from this study extend our knowledge on the genetic diversity of RSV circulating in Vietnam. PMID:25702707

  17. Incidence and Risk Factors for Respiratory Syncytial Virus and Human Metapneumovirus Infections among Children in the Remote Highlands of Peru

    PubMed Central

    Wu, Andrew; Budge, Philip J.; Williams, John; Griffin, Marie R.; Edwards, Kathryn M.; Johnson, Monika; Zhu, Yuwei; Hartinger, Stella; Verastegui, Hector; Gil, Ana I.; Lanata, Claudio F.; Grijalva, Carlos G.

    2015-01-01

    Introduction The disease burden and risk factors for respiratory syncytial virus (RSV) and human metapneumovirus (MPV) infections among children living in remote, rural areas remain unclear. Materials and Methods We conducted a prospective, household-based cohort study of children aged <3 years living in remote rural highland communities in San Marcos, Cajamarca, Peru. Acute respiratory illnesses (ARI), including lower respiratory tract infection (LRTI), were monitored through weekly household visits from March 2009 through September 2011. Nasal swabs collected during ARI/LRTI were tested for RSV, MPV, and other respiratory viruses using real-time RT-PCR. Incidence rates and rate ratios were calculated using mixed effects Poisson regression. Results Among 892 enrolled children, incidence rates of RSV and MPV ARI were 30 and 17 episodes per 100 child-years, respectively. The proportions of RSV and MPV ARI that presented as LRTI were 12.5% and 8.9%, respectively. Clinic visits for ARI and hospitalizations were significantly more frequent (all p values <0.05) among children with RSV (clinic 41% and hospital 5.3%) and MPV ARI (38% and 3.5%) when compared with other viral infections (23% and 0.7%) and infections without virus detected (24% and 0.6%). In multivariable analysis, risk factors for RSV detection included younger age (RR 1.02, 95% CI: 1.00-1.03), the presence of a smoker in the house (RR 1.63, 95% CI: 1.12-2.38), residing at higher altitudes (RR 1.93, 95% CI: 1.25-3.00 for 2nd compared to 1st quartile residents; RR 1.98, 95% CI: 1.26-3.13 for 3rd compared to 1st quartile residents). Having an unemployed household head was significantly associated with MPV risk (RR 2.11, 95% CI: 1.12-4.01). Conclusion In rural high altitude communities in Peru, childhood ARI due to RSV or MPV were common and associated with higher morbidity than ARI due to other viruses or with no viral detections. The risk factors identified in this study may be considered for interventional

  18. Large-scale seroprevalence analysis of human metapneumovirus and human respiratory syncytial virus infections in Beijing, China

    PubMed Central

    2011-01-01

    Background Human metapneumovirus (hMPV), a recently identified virus, causes acute respiratory tract infections (ARTIs) in infants and children. However, studies on the seroepidemeology of hMPV are very limited in China. To assess the seroprevalence of hMPV infection in China, we tested a total of 1,156 serum specimens for the presence of anti-hMPV IgG antibody in children and adults free of acute respiratory illness in Beijing, China by using hMPV nucleocapsid (N) protein as an antigen. As a control, we used the human serum antibody against the N protein of human respiratory syncytial virus (hRSV), the most important viral agent responsible for ARIs in children. Results The seropositive rate for hMPV increased steadily with age from 67% at 1-6 mo to 100% at age 20. However, the rate dropped slightly between 6 mo and 1 yr of age. The seropositive rate for hRSV also increased steadily with age from 71% at 1-6 mo to 100% at age 20. In children aged six months to six years, the seropositive rates for the anti-hRSV IgG antibody were significantly higher than those for hMPV. Additionally, IgG antibody titers to hMPV and hRSV were significantly higher in adults than in young children. Consistent with the seropositive rates, the geometric mean titer of anti-hMPV IgG antibody was lower than that of anti-hRSV IgG antibody in children aged six months to six years. Conclusions Our results indicate that similar to hRSV, exposure to hMPV is ubiquitous in the Beijing population. However, the seroprevalence of anti-hMPV IgG antibody is lower than that of hRSV in children between six months and six years old, which suggests a different number of repeat infections or a different response to infections. PMID:21310026

  19. Relating plaque morphology to respiratory syncytial virus subgroup, viral load, and disease severity in children

    PubMed Central

    Kim, Young-In; Murphy, Ryan; Majumdar, Sirshendu; Harrison, Lisa G.; Aitken, Jody; DeVincenzo, John P.

    2015-01-01

    Background Viral culture plaque morphology in human cell lines are markers for growth capability and cytopathic effect, and have been used to assess viral fitness and select pre-attenuation candidates for live viral vaccines. We classified RSV plaque morphology and analyzed the relationship between plaque morphology as compared to subgroup, viral load and clinical severity of infection in infants and children. Methods We obtained respiratory secretions from 149 RSV-infected children. Plaque morphology and viral load was assessed within the first culture passage in HEp-2 cells. Viral load was measured by PCR, as was RSV subgroup. Disease severity was determined by hospitalization, length of stay, intensive care requirement, and respiratory failure. Results Plaque morphology varied between individual subjects; however, similar results were observed among viruses collected from upper and lower respiratory tracts of the same subject. Significant differences in plaque morphology were observed between RSV subgroups. No correlations were found among plaque morphology and viral load. Plaque morphology did not correlate with disease severity. Conclusions Plaque morphology measures parameters that are viral-specific and independent of the human host. Morphologies vary between patients and are related to RSV subgroup. In HEp-2 cells, RSV plaque morphology appears unrelated to disease severity in RSV-infected children. PMID:26107392

  20. Relative frequency, Possible Risk Factors, Viral Codetection Rates, and Seasonality of Respiratory Syncytial Virus Among Children With Lower Respiratory Tract Infection in Northeastern Brazil.

    PubMed

    Gurgel, Ricardo Queiroz; Bezerra, Patrícia Gomes de Matos; Duarte, Maria do Carmo Menezes Bezerra; Moura, Adriana Ávila; Souza, Edna Lucia; Silva, Luciana Sobral da Silveira; Suzuki, Claudia Eiko; Peixoto, Rodrigo Buzzatti

    2016-04-01

    Few studies, each limited to a single major city, have investigated the prevalence and seasonal patterns of different viruses among children with low respiratory tract infections (LRTI) in Northeastern Brazil. The aim of this study was to determine the frequency of respiratory syncytial virus (RSV) and of 7 other viruses in children for LRTI in 4 capitals from this region, and investigate their association with several risk factors, including meteorological data. From April 2012 to March 2013, 507 children, aged up to 24 months and hospitalized with LRTI in one of the participating centers at Aracajú, Salvador, Recife, and Maceió, had a sample of nasopharyngeal aspirate collected and analyzed for the following viruses by reverse-transcription polymerase chain reaction followed by hybridization on low-density microarrays: RSV, influenza, parainfluenza, adenovirus, rhinovirus, metapneumovirus, bocavirus, and coronavirus. The result was positive in 66.5% of cases, RSV was the most common virus (40.2%). Except for rhinovirus (17%), all other virus had frequency rates lower than 6%. Viral coinfections were detected in 13.8% of samples. Possible related risk factors for RSV infection were low age upon entry, attendance of daycare, low gestational age, and low educational level of the father. The relative frequency of viral infections was associated with increasing temperature and decreasing humidity separately, but the results also suggested both associated with increased frequency of RSV. Some of these findings differ from those reported for other regions in Brazil and may be used to guide policies that address LRTI. PMID:27082548

  1. Genetic diversity and evolutionary insights of respiratory syncytial virus A ON1 genotype: global and local transmission dynamics.

    PubMed

    Duvvuri, Venkata R; Granados, Andrea; Rosenfeld, Paul; Bahl, Justin; Eshaghi, Alireza; Gubbay, Jonathan B

    2015-09-30

    Human respiratory syncytial virus (RSV) A ON1 genotype, first detected in 2010 in Ontario, Canada, has been documented in 21 countries to date. This study investigated persistence and transmission dynamics of ON1 by grouping 406 randomly selected RSV-positive specimens submitted to Public Health Ontario from August 2011 to August 2012; RSV-A-positive specimens were genotyped. We identified 370 RSV-A (181 NA1, 135 NA2, 51 ON1 3 GA5) and 36 RSV-B positive specimens. We aligned time-stamped second hypervariable region (330 bp) of G-gene sequence data (global, n = 483; and Ontario, n = 60) to evaluate transmission dynamics. Global data suggests that the most recent common ancestor of ON1 emerged during the 2008-2009 season. Mean evolutionary rate of the global ON1 was 4.10 × 10(-3) substitutions/site/year (95% BCI 3.1-5.0 × 10(-3)), not significantly different to that of Ontario ON1. The estimated mean reproductive number (R0 = ∼ 1.01) from global and Ontario sequences showed no significant difference and implies stability among global RSV-A ON1. This study suggests that local epidemics exhibit similar underlying evolutionary and epidemiological dynamics to that of the persistent global RSV-A ON1 population. These findings underscore the importance of continual molecular surveillance of RSV in order to gain a better understanding of epidemics.

  2. Cytokine expression in respiratory syncytial virus-infected mice as measured by quantitative reverse-transcriptase PCR.

    PubMed

    Deng, Xinqing; Li, Haijing; Tang, Yi Wei

    2003-02-01

    In the murine model for respiratory syncytial virus (RSV) infection, cytokine patterns induced by vaccinations with either killed (i.e. formalin-inactivated, alum-precipitated) virus (KV) or live virus (LV) have been shown to influence disease expression. To determine the mRNA expression of the cytokines IL-4 and IFN-gamma in BALB/c mice challenged with RSV, a real-time quantitative reverse-transcriptase PCR assay was developed. This assay uses 5'-exonuclease fluorogenic probes and is performed on the ABI PRISM 7700 Sequence Detector System (TaqMan). The relative quantitative levels of mRNA for IL-4 and IFN-gamma were compared with those measured by an RNase protection assay (RPA) and an enzyme immunoassay (EIA), which are methods used to measure the levels of mRNA and protein, respectively. Results obtained by the TaqMan assay showed that mice primed with KV induces increased IL-4 mRNA production while LV induces increased IFN-gamma mRNA, which is in agreement with conventional methods. IL-4 and IFN-gamma relative quantities obtained from TaqMan were highly correlated to those determined by RPA (r=0.96 for IFN-gamma, P<0.01) and EIA (r=0.90 for IL-4 and r=0.75 for IFN-gamma, P<0.01). Assay reproducibility was examined by testing a same sample in triplicate at three experiments. Minimal deviation values were observed in both intra- and inter-assays. TaqMan, which is rapid, sensitive and reproducible, provides an alternative tool for the quantitative analysis of cytokine mRNA expression in the murine model of RSV immunopathogenesis. PMID:12505627

  3. The interpretation of rapid antigen testing for respiratory syncytial virus is more complicated than positive or negative

    PubMed Central

    Walsh, Paul; Cabangangan, Juanito; Overmeyer, Christina; Pusavat, James; Hancock, Christine; Shanholtzer, Lucas; Nguyen, Thienphuc; Heffner, Jacquelyn; Mordechai, Eli; Adelson, Martin E.; Iacono, Kathryn T.

    2014-01-01

    Objective To measure the performance characteristics of an immunochromatographic rapid antigen test for respiratory syncytial virus (RSV) and determine how its interpretation should be contextualized in patients presenting to the emergency department (ED) with bronchiolitis. Design Diagnostic accuracy study of a rapid RSV test. Setting County hospital emergency department. Intervention We took paired nasal samples from consecutively enrolled infants with bronchiolitis and tested them with a rapid immunochromatographic antigen test and reverse transcriptase polymerase chain reaction gold standard. Outcome measures Sensitivity, specificity, likelihood ratios, predictive values, evidence of spectrum bias and clinical characteristics of the patients. Using these we constructed a graphical contextual model to show how the results of RSV antigen tests from infants presenting within 24 hours should influence interpretation of subsequent antigen tests. Results We analyzed 607 patients. The sensitivity and specificity for immunochromatographic testing was 79.4% (95% CI 73.9%, 84.2%) and 67.1% (95% CI 61.9%, 72%) respectively. We found little evidence of spectrum bias. In our contextual model the best predictor of a positive RT-PCR test was a positive antigen test OR 5.47 (95%CI 3.65, 8.18) and the number of other infants having positive tests within 24 hours OR 1.48 (95%CI 1.26, 1.72) per infant. Increasing numbers presenting to the ED with bronchiolitis in a given day increases the probability of RSV infection. Conclusion The RSV antigen test we examined had modest performance characteristics. The results of the antigen test should be interpreted in the context of the results of previous tests. PMID:23964062

  4. Respiratory Syncytial Virus-Infected Mesenchymal Stem Cells Regulate Immunity via Interferon Beta and Indoleamine-2,3-Dioxygenase

    PubMed Central

    Cheung, Michael B.; Sampayo-Escobar, Viviana; Green, Ryan; Moore, Martin L.; Mohapatra, Subhra; Mohapatra, Shyam S.

    2016-01-01

    Respiratory syncytial virus (RSV) has been reported to infect human mesenchymal stem cells (MSCs) but the consequences are poorly understood. MSCs are present in nearly every organ including the nasal mucosa and the lung and play a role in regulating immune responses and mediating tissue repair. We sought to determine whether RSV infection of MSCs enhances their immune regulatory functions and contributes to RSV-associated lung disease. RSV was shown to replicate in human MSCs by fluorescence microscopy, plaque assay, and expression of RSV transcripts. RSV-infected MSCs showed differentially altered expression of cytokines and chemokines such as IL-1β, IL6, IL-8 and SDF-1 compared to epithelial cells. Notably, RSV-infected MSCs exhibited significantly increased expression of IFN-β (~100-fold) and indoleamine-2,3-dioxygenase (IDO) (~70-fold) than in mock-infected MSCs. IDO was identified in cytosolic protein of infected cells by Western blots and enzymatic activity was detected by tryptophan catabolism assay. Treatment of PBMCs with culture supernatants from RSV-infected MSCs reduced their proliferation in a dose dependent manner. This effect on PBMC activation was reversed by treatment of MSCs with the IDO inhibitors 1-methyltryptophan and vitamin K3 during RSV infection, a result we confirmed by CRISPR/Cas9-mediated knockout of IDO in MSCs. Neutralizing IFN-β prevented IDO expression and activity. Treatment of MSCs with an endosomal TLR inhibitor, as well as a specific inhibitor of the TLR3/dsRNA complex, prevented IFN-β and IDO expression. Together, these results suggest that RSV infection of MSCs alters their immune regulatory function by upregulating IFN-β and IDO, affecting immune cell proliferation, which may account for the lack of protective RSV immunity and for chronicity of RSV-associated lung diseases such as asthma and COPD. PMID:27695127

  5. Risk of nosocomial respiratory syncytial virus infection and effectiveness of control measures to prevent transmission events: a systematic review.

    PubMed

    French, Clare E; McKenzie, Bruce C; Coope, Caroline; Rajanaidu, Subhadra; Paranthaman, Karthik; Pebody, Richard; Nguyen-Van-Tam, Jonathan S; Higgins, Julian P T; Beck, Charles R

    2016-07-01

    Respiratory syncytial virus (RSV) causes a significant public health burden, and outbreaks among vulnerable patients in hospital settings are of particular concern. We reviewed published and unpublished literature from hospital settings to assess: (i) nosocomial RSV transmission risk (attack rate) during outbreaks, (ii) effectiveness of infection control measures. We searched the following databases: MEDLINE, EMBASE, CINAHL, Cochrane Library, together with key websites, journals and grey literature, to end of 2012. Risk of bias was assessed using the Cochrane risk of bias tool or Newcastle-Ottawa scale. A narrative synthesis was conducted. Forty studies were included (19 addressing research question one, 21 addressing question two). RSV transmission risk varied by hospital setting; 6-56% (median: 28·5%) in neonatal/paediatric settings (n = 14), 6-12% (median: 7%) in adult haematology and transplant units (n = 3), and 30-32% in other adult settings (n = 2). For question two, most studies (n = 13) employed multi-component interventions (e.g. cohort nursing, personal protective equipment (PPE), isolation), and these were largely reported to be effective in reducing nosocomial transmission. Four studies examined staff PPE; eye protection appeared more effective than gowns and masks. One study reported on RSV prophylaxis for patients (RSV-Ig/palivizumab); there was no statistical evidence of effectiveness although the sample size was small. Overall, risk of bias for included studies tended to be high. We conclude that RSV transmission risk varies widely during hospital outbreaks. Although multi-component control strategies appear broadly successful, further research is required to disaggregate the effectiveness of individual components including the potential role of palivizumab prophylaxis.

  6. Attenuated Bordetella pertussis Vaccine Protects against Respiratory Syncytial Virus Disease via an IL-17–Dependent Mechanism

    PubMed Central

    Schnoeller, Corinna; Roux, Xavier; Sawant, Devika; Raze, Dominique; Olszewska, Wieslawa; Locht, Camille

    2014-01-01

    Rationale: We attenuated virulent Bordetella pertussis by genetically eliminating or detoxifying three major toxins. This strain, named BPZE1, is being developed as a possible live nasal vaccine for the prevention of whooping cough. It is immunogenic and safe when given intranasally in adult volunteers. Objectives: Before testing in human infants, we wished to examine the potential effect of BPZE1 on a common pediatric infection (respiratory syncytial virus [RSV]) in a preclinical model. Methods: BPZE1 was administered before or after RSV administration in adult or neonatal mice. Pathogen replication, inflammation, immune cell recruitment, and cytokine responses were measured. Measurements and Main Results: BPZE1 alone did not cause overt disease, but induced efflux of neutrophils into the airway lumen and production of IL-10 and IL-17 by mucosal CD4+ T cells. Given intranasally before RSV infection, BPZE1 markedly attenuated RSV, preventing weight loss, reducing viral load, and attenuating lung cell recruitment. Given neonatally, BPZE1 also protected against RSV-induced weight loss even through to adulthood. Furthermore, it markedly increased IL-17 production by CD4+ T cells and natural killer cells and recruited regulatory cells and neutrophils after virus challenge. Administration of anti–IL-17 antibodies ablated the protective effect of BPZE1 on RSV disease. Conclusions: Rather than enhancing RSV disease, BPZE1 protected against viral infection, modified viral responses, and enhanced natural mucosal resistance. Prevention of RSV infection by BPZE1 seems in part to be caused by induction of IL-17. Clinical trial registered with www.clinicaltrials.gov (NCT 01188512). PMID:24261996

  7. Development of a Luciferase Immunoprecipitation System Assay To Detect IgG Antibodies against Human Respiratory Syncytial Virus Nucleoprotein

    PubMed Central

    Kumari, Sangeeta; Crim, Roberta Lynne; Kulkarni, Ashwin; Audet, Susette A.; Mdluli, Thembi; Murata, Haruhiko

    2014-01-01

    The nucleoprotein of respiratory syncytial virus (RSV-N) is immunogenic and elicits an IgG response following infection. The RSV-N gene was cloned into a mammalian expression vector, pREN2, and the expressed luciferase-tagged protein (Ruc-N) detected anti-RSV-N-specific IgG antibodies using a high-throughput immunoprecipitation method (the luciferase immunoprecipitation system [LIPS]-NRSV assay). The specificity of the assay was evaluated using monoclonal antibodies (MAbs) and monospecific pre- and postimmunization rabbit antisera. Blood serum samples from chimpanzees and humans with proven/probable RSV infection were also tested. The pre- and postimmunization serum samples from rabbits given human metapneumovirus (HMPV) or measles virus were negative when tested by the LIPS-NRSV assay, while antisera obtained after immunization with either the RSV-A or RSV-B strain gave positive signals in a dose-dependent manner. RSV-N MAb 858-3 gave a positive signal in the LIPS-NRSV assay, while MAbs against other paramyxovirus nucleoproteins or RSV-F or RSV-G did not. Serum samples from chimpanzees simultaneously immunized with vaccinia-RSV-F and vaccinia-RSV-G recombinant viruses were negative in the LIPS-NRSV assay; however, anti-RSV-N IgG responses were detected following subsequent RSV challenge. Seven of the 12 infants who were seronegative at 9 months of age had detectable anti-RSV-N antibodies when they were retested at 15 to 18 months of age. The LIPS-NRSV assay detects specific anti-RSV-N IgG responses that may be used as a biomarker of RSV infection. PMID:24403526

  8. Genetic diversity and evolutionary insights of respiratory syncytial virus A ON1 genotype: global and local transmission dynamics

    PubMed Central

    Duvvuri, Venkata R.; Granados, Andrea; Rosenfeld, Paul; Bahl, Justin; Eshaghi, Alireza; Gubbay, Jonathan B.

    2015-01-01

    Human respiratory syncytial virus (RSV) A ON1 genotype, first detected in 2010 in Ontario, Canada, has been documented in 21 countries to date. This study investigated persistence and transmission dynamics of ON1 by grouping 406 randomly selected RSV-positive specimens submitted to Public Health Ontario from August 2011 to August 2012; RSV-A-positive specimens were genotyped. We identified 370 RSV-A (181 NA1, 135 NA2, 51 ON1 3 GA5) and 36 RSV-B positive specimens. We aligned time-stamped second hypervariable region (330 bp) of G-gene sequence data (global, n = 483; and Ontario, n = 60) to evaluate transmission dynamics. Global data suggests that the most recent common ancestor of ON1 emerged during the 2008–2009 season. Mean evolutionary rate of the global ON1 was 4.10 × 10−3 substitutions/site/year (95% BCI 3.1–5.0 × 10−3), not significantly different to that of Ontario ON1. The estimated mean reproductive number (R0 = ∼ 1.01) from global and Ontario sequences showed no significant difference and implies stability among global RSV-A ON1. This study suggests that local epidemics exhibit similar underlying evolutionary and epidemiological dynamics to that of the persistent global RSV-A ON1 population. These findings underscore the importance of continual molecular surveillance of RSV in order to gain a better understanding of epidemics. PMID:26420660

  9. A randomized, double-blind, placebo-controlled study of an RNAi-based therapy directed against respiratory syncytial virus

    PubMed Central

    DeVincenzo, John; Lambkin-Williams, Robert; Wilkinson, Tom; Cehelsky, Jeffrey; Nochur, Sara; Walsh, Edward; Meyers, Rachel; Gollob, Jared; Vaishnaw, Akshay

    2010-01-01

    RNA interference (RNAi) is a natural mechanism regulating protein expression that is mediated by small interfering RNAs (siRNA). Harnessing RNAi has potential to treat human disease; however, clinical evidence for the effectiveness of this therapeutic approach is lacking. ALN-RSV01 is an siRNA directed against the mRNA of the respiratory syncytial virus (RSV) nucleocapsid (N) protein and has substantial antiviral activity in a murine model of RSV infection. We tested the antiviral activity of ALN-RSV01 in adults experimentally infected with wild-type RSV. Eighty-eight healthy subjects were enrolled into a randomized, double-blind, placebo-controlled trial. A nasal spray of ALN-RSV01 or saline placebo was administered daily for 2 days before and for 3 days after RSV inoculation. RSV was measured serially in nasal washes using several different viral assays. Intranasal ALN-RSV01 was well tolerated, exhibiting a safety profile similar to saline placebo. The proportion of culture-defined RSV infections was 71.4 and 44.2% in placebo and ALN-RSV01 recipients, respectively (P = 0.009), representing a 38% decrease in the number of infected and a 95% increase in the number of uninfected subjects. The acquisition of infection over time was significantly lower in ALN-RSV01 recipients (P = 0.007 and P = 0.03, viral culture and PCR, respectively). Multiple logistic regression analysis showed that the ALN-RSV01 antiviral effect was independent of other factors, including preexisting RSV antibody and intranasal proinflammatory cytokine concentrations. ALN-RSV01 has significant antiviral activity against human RSV infection, thus establishing a unique proof-of-concept for an RNAi therapeutic in humans and providing the basis for further evaluation in naturally infected children and adults. PMID:20421463

  10. Evaluation of recent New Vaccine Surveillance Network data regarding respiratory syncytial virus hospitalization rates in US preterm infants.

    PubMed

    DeVincenzo, John P; Ambrose, Christopher S; Makari, Doris; Weiner, Leonard B

    2016-04-01

    In July 2014, the Committee on Infectious Diseases (COID) updated their guidance on the use of palivizumab, recommending against use in preterm infants 29 to 35 weeks' gestational age (wGA). A primary data source cited to support this significant change was the low respiratory syncytial virus (RSV) hospitalization rate observed in the subpopulation of preterm (<37 wGA) infants evaluated from 2000 to 2005 through the New Vaccine Surveillance Network (NVSN). Here we critically appraise the preterm infant data from the NVSN in the context of data regarding the use of palivizumab in this same time period. Data from the NVSN, an analysis of Florida Medicaid data, and a national survey of US in-hospital palivizumab administration demonstrated that during 2001 to 2007, palivizumab was administered to 59% to 83% of preterm infants born at <32 wGA and 21% to 27% of all preterm infants (<37 wGA). When the NVSN data regarding incidence of RSV hospitalization in preterm infant subgroups were evaluated as a function of chronologic age, preterm infants <32 wGA showed a paradoxical increase in RSV hospitalization with older age, with the highest risk of RSV hospitalization occurring at 18 to 23 months of age. This pattern is most consistent with a reduction in RSV hospitalizations in <32 wGA infants in the first 12 to 18 months of life due to high palivizumab use at these young ages. The NVSN data were not designed to and cannot accurately describe RSV disease burden in preterm infants given the small size of the analyzed subpopulation and the high use of palivizumab during the study period.

  11. Functional analysis of antibody responses of feedlot cattle to bovine respiratory syncytial virus following vaccination with mixed vaccines.

    PubMed Central

    West, K; Ellis, J

    1997-01-01

    The antibody response of cattle to bovine respiratory syncytial virus (BRSV) immunization was investigated using 4 different commercially available mixed vaccines. Forty, 5-6 month old, beef calves, randomly assigned to groups of 10, were vaccinated on day 0 and 21 with 1 of 3 inactivated vaccines, (3 groups), or a modified live virus (MLV) vaccine. BRSV-specific antibody responses were measured prior to vaccination and on day 35 by using an enzyme linked immunosorbent assay (ELISA), virus neutralization assay (VN), a fusion inhibition assay (FI); and responses were also measured for their ability to facilitate antibody dependent, complement mediated cytotoxicity (ADCMC) of BRSV infected cells. Sera from day 35 were, in addition, analyzed by use of an IgG1, IgG2 isotype specific ELISA. All vaccines induced significant increases in BRSV specific IgG antibody as measured by ELISA, but only one inactivated and the MLV vaccine induced significant increases in VN titers. Fusion inhibiting antibody titers were low or undetected in calves vaccinated with the inactivated vaccines. Vaccination with modified live virus induced significantly higher titers of fusion inhibiting antibodies, which are considered to be most highly correlated with protection. The VN to ELISA and FI to ELISA ratio of the calves that received MLV vaccine were significantly greater than the calves receiving the 3 inactivated vaccines. Vaccination with MLV induced the highest IgG2/IgG1 ratio. This difference was small, and only significant relative to 2 of the inactivated vaccine groups, which were not significantly different from each other. The higher proportion of IgG2 isotype in the MLV sera was not associated with lower ADCMC, a function not attributed to this isotype. The VN and FI titers, but not the ELISA value of the sera, were most predictive of ADCMC. The inactivation processes apparently alter epitopes and affect the induction of functional antibodies. PMID:9008797

  12. Immunogenetic responses in calves to intranasal delivery of bovine respiratory syncytial virus (BRSV) epitopes encapsulated in poly (DL-lactide-co-glycolide) microparticles.

    PubMed

    Kavanagh, Owen V; Adair, Brian M; Welsh, Michael; Earley, Bernadette

    2013-10-01

    Bovine respiratory syncytial virus (BRSV) is the principal aetiological agent of the bovine respiratory disease complex. A BRSV subunit vaccine candidate consisting of two synthetic peptides representing putative protective epitopes on BRSV surface glycoproteins in soluble form or encapsulated in poly(lactide-co-glycolide) (PLG) microparticles were prepared. Calves (10 weeks old) with diminishing levels of BRSV-specific maternal antibody were intranasally administered a single dose of the different peptide formulations. Peptide-specific local immune responses (nasal secretion IgA), but not systemic humoral (serum IgG) or cellular responses (serum IFN-γ), were generated by all forms of peptide. There was a significant reduction in occurrence of respiratory disease in the animals inoculated with all peptide formulations compared to animals given PBS alone. Furthermore no adverse effects were observed in any of the animals post vaccination. These results suggest that intranasal immunisation with the peptide subunit vaccine does induce an as yet unidentified protective immune response.

  13. [Atypical interstitial pneumonia (AIP) in calves and young cattle in Schleswig-Holstein in conjunction with an infection by the bovine respiratory syncytial virus (BRSV)].

    PubMed

    Appel, G; Heckert, H P

    1989-04-01

    It is reported on atypical interstitial pneumonia (AIP) in 16 mostly Holstein-Frisian calves and feeders from 13 different farms in Schleswig-Holstein in association with an infection by the respiratory syncytial virus (BRSV). All animals were submitted with identical clinical histories. Macroscopically the lung lesions were characterized by alveolar and interstitial edema and emphysema. Microscopically there was a diffuse interstitial pneumonia with formation of hyaline membranes and multinucleated giant cells. From the investigation material of the 16 animals BRSV was confirmed by direct immunofluorescence with monoclonal antibodies in 4 animals from 4 different farms.

  14. Prevalence and clinical features of respiratory syncytial virus in children hospitalized for community-acquired pneumonia in northern Brazil

    PubMed Central

    2012-01-01

    Background Childhood pneumonia and bronchiolitis is a leading cause of illness and death in young children worldwide with Respiratory Syncytial Virus (RSV) as the main viral cause. RSV has been associated with annual respiratory disease outbreaks and bacterial co-infection has also been reported. This study is the first RSV epidemiological study in young children hospitalized with community-acquired pneumonia (CAP) in Belém city, Pará (Northern Brazil). Methods With the objective of determining the prevalence of RSV infection and evaluating the patients’ clinical and epidemiological features, we conducted a prospective study across eight hospitals from November 2006 to October 2007. In this study, 1,050 nasopharyngeal aspirate samples were obtained from hospitalized children up to the age of three years with CAP, and tested for RSV antigen by direct immunofluorescence assay and by Reverse Transcription Polymerase Chain Reaction (RT-PCR) for RSV Group identification. Results RSV infection was detected in 243 (23.1%) children. The mean age of the RSV-positive group was lower than the RSV-negative group (12.1 months vs 15.5 months, p<0.001) whereas gender distribution was similar. The RSV-positive group showed lower means of C-reactive protein (CRP) in comparison to the RSV-negative group (15.3 vs 24.0 mg/dL, p<0.05). Radiological findings showed that 54.2% of RSV-positive group and 50.3% of RSV-negative group had interstitial infiltrate. Bacterial infection was identified predominantly in the RSV-positive group (10% vs 4.5%, p<0.05). Rhinorrhea and nasal obstruction were predominantly observed in the RSV-positive group. A co-circulation of RSV Groups A and B was identified, with a predominance of Group B (209/227). Multivariate analysis revealed that age under 1 year (p<0.015), CRP levels under 48 mg/dL (p<0.001) and bacterial co-infection (p<0.032) were independently associated with the presence of RSV and, in the analyze of symptoms, nasal obstruction

  15. Long-Term Burden and Respiratory Effects of Respiratory Syncytial Virus Hospitalization in Preterm Infants-The SPRING Study.

    PubMed

    Carbonell-Estrany, Xavier; Pérez-Yarza, Eduardo G; García, Laura Sanchez; Guzmán Cabañas, Juana M; Bòria, Elena Villarrubia; Atienza, Belén Bernardo

    2015-01-01

    The health status of premature infants born 321-350 weeks' gestational age (wGA) hospitalized for RSV infection in the first year of life (cases; n = 125) was compared to that of premature infants not hospitalized for RSV (controls; n = 362) through 6 years. The primary endpoints were the percentage of children with wheezing between 2-6 years and lung function at 6 years of age. Secondary endpoints included quality of life, healthcare resource use, and allergic sensitization. A significantly higher proportion of cases than controls experienced recurrent wheezing through 6 years of age (46.7% vs. 27.4%; p = 0.001). The vast majority of lung function tests appeared normal at 6 years of age in both cohorts. In children with pulmonary function in the lower limit of normality (FEV1 Z-score [-2; -1]), wheezing was increased, particularly for cases vs. controls (72.7% vs. 18.9%, p = 0.002). Multivariate analysis revealed the most important factor for wheezing was RSV hospitalization. Quality of life on the respiratory subscale of the TAPQOL was significantly lower (p = 0.001) and healthcare resource utilization was significantly higher (p<0.001) in cases than controls. This study confirms RSV disease is associated with wheezing in 32-35 wGA infants through 6 years of age.

  16. Long-Term Burden and Respiratory Effects of Respiratory Syncytial Virus Hospitalization in Preterm Infants—The SPRING Study

    PubMed Central

    Carbonell-Estrany, Xavier; Pérez-Yarza, Eduardo G.; García, Laura Sanchez; Guzmán Cabañas, Juana M.; Bòria, Elena Villarrubia; Atienza, Belén Bernardo

    2015-01-01

    The health status of premature infants born 321-350 weeks’ gestational age (wGA) hospitalized for RSV infection in the first year of life (cases; n = 125) was compared to that of premature infants not hospitalized for RSV (controls; n = 362) through 6 years. The primary endpoints were the percentage of children with wheezing between 2-6 years and lung function at 6 years of age. Secondary endpoints included quality of life, healthcare resource use, and allergic sensitization. A significantly higher proportion of cases than controls experienced recurrent wheezing through 6 years of age (46.7% vs. 27.4%; p = 0.001). The vast majority of lung function tests appeared normal at 6 years of age in both cohorts. In children with pulmonary function in the lower limit of normality (FEV1 Z-score [-2; -1]), wheezing was increased, particularly for cases vs. controls (72.7% vs. 18.9%, p = 0.002). Multivariate analysis revealed the most important factor for wheezing was RSV hospitalization. Quality of life on the respiratory subscale of the TAPQOL was significantly lower (p = 0.001) and healthcare resource utilization was significantly higher (p<0.001) in cases than controls. This study confirms RSV disease is associated with wheezing in 32-35 wGA infants through 6 years of age. PMID:25955487

  17. Kinetics of Respiratory Syncytial Virus (RSV) Memphis Strain 37 (M37) Infection in the Respiratory Tract of Newborn Lambs as an RSV Infection Model for Human Infants

    PubMed Central

    Larios Mora, Alejandro; Detalle, Laurent; Van Geelen, Albert; Davis, Michael S.; Stohr, Thomas; Gallup, Jack M.; Ackermann, Mark R.

    2015-01-01

    Rationale Respiratory syncytial virus (RSV) infection in preterm and newborn infants can result in severe bronchiolitis and hospitalization. The lamb lung has several key features conducive to modeling RSV infection in human infants, including susceptibility to human strains of RSV such as the A2, Long, and Memphis Strain 37 (M37). In this study, the kinetics of M37 infection was investigated in newborn lambs in order to better define clinical, viral, physiological, and immunological parameters as well as the pathology and lesions. Methods Newborn lambs were nebulized with M37 hRSV (6 mL of 1.27 x 107 FFU/mL), monitored daily for clinical responses, and respiratory tissues were collected from groups of lambs at days 1, 3, 4, 6, and 8 post-inoculation for the assessment of viral replication parameters, lesions and also cellular, immunologic and inflammatory responses. Results Lambs had increased expiratory effort (forced expiration) at days 4, 6, and 8 post-inoculation. Nasal wash lacked RSV titers at day 1, but titers were present at low levels at days 3 (peak), 4, and 8. Viral titers in bronchoalveolar lavage fluid (BALF) reached a plateau at day 3 (4.6 Log10 FFU/mL), which was maintained until day 6 (4.83 Log10 FFU/mL), and were markedly reduced or absent at day 8. Viral RNA levels (detected by RT-qPCR) in BALF were indistinguishable at days 3 (6.22 ± 0.08 Log10 M37 RNA copies/mL; mean ± se) and 4 (6.20 ± 0.16 Log10 M37 RNA copies/mL; mean ± se) and increased slightly on day 6 (7.15 ± 0.2 Log10 M37 RNA copies/mL; mean ± se). Viral antigen in lung tissue as detected by immunohistochemistry was not seen at day 1, was present at days 3 and 4 before reaching a peak by day 6, and was markedly reduced by day 8. Viral antigen was mainly present in airways (bronchi, bronchioles) at day 3 and was increasingly present in alveolar cells at days 4 and 6, with reduction at day 8. Histopathologic lesions such as bronchitis/bronchiolitis, epithelial necrosis and

  18. Defining the timing of respiratory syncytial virus (RSV) outbreaks: an epidemiological study

    PubMed Central

    Terletskaia-Ladwig, Elena; Enders, Gisela; Schalasta, Gunnar; Enders, Martin

    2005-01-01

    Background Seasonal RSV infections occur every year and affect particularly children under six months of age. Passive immunoprophylaxis with monoclonal antibody Palivizumab is recommended in the period with high risk of RSV infection. This study aims to define the period for the southern part of Germany (Stuttgart area). Methods Epidemiological analysis of the RSV situation in southern Germany from 1996 to 2004 and comparison of results with literature was made. The respiratory tract specimens were sent in for the detection of RSV mainly by paediatric clinics. Detection of RSV was carried out mainly by real-time RT-PCR or by ELISA "Pathfinder". RSV outbreaks were depicted as an absolute number and as a percentage of RSV diagnoses in a month. Onsets, offsets, peaks, duration and severity of RSV seasons were defined and analysed. Results An early season with strong RSV activity (early-high phase) was followed by a weaker late season (late-low phase) in a regular biennial rhythm. However, onsets, offsets and durations of outbreaks varied significantly from year to year. RSV epidemics in southern Germany were found to oscillate in an antiphase with RSV epidemics in Finland and Sweden. Conclusion The long-term regular biennial rhythm allows predicting whether the next outbreak will be late or early and whether RSV activity will be strong or weak. Not foreseeable, however, is the precise time of increase and decrease of RSV activity. Moreover, the regular seasonal pattern may be disrupted by irregular outbreaks. Thus, activity of RSV has to be monitored every year to define the period with high risk of infection. PMID:15801975

  19. Phenotype and Hierarchy of Two Transgenic T Cell Lines Targeting the Respiratory Syncytial Virus KdM282-90 Epitope Is Transfer Dose-Dependent.

    PubMed

    Morabito, Kaitlyn M; Erez, Noam; Graham, Barney S; Ruckwardt, Tracy J

    2016-01-01

    In this study, we compared two lines of transgenic CD8+ T cells specific for the same KdM282-90 epitope of respiratory syncytial virus in the CB6F1 hybrid mouse model. Here we found that these two transgenic lines had similar in vivo abilities to control viral load after respiratory syncytial virus infection using adoptive transfer. Transfer of the TRBV13-2 line resulted in higher levels of IL-6 and MIP1-α in the lung than TRBV13-1 transfer. Interestingly, when large numbers of cells were co-transferred, the lines formed a hierarchy, with TRBV13-2 being immunodominant over TRBV13-1 in the mediastinal lymph node despite no identifiable difference in proliferation or apoptosis between the lines. This hierarchy was not established when lower cell numbers were transferred. The phenotype and frequency of proliferating cells were also cell transfer dose-dependent with higher percentages of CD127loCD62LloKLRG1lo and proliferating cells present when lower numbers of cells were transferred. These results illustrate the importance of cell number in adoptive transfer experiments and its influence on the phenotype and hierarchy of the subsequent T cell response. PMID:26752171

  20. Reversion of somatic mutations of the respiratory syncytial virus-specific human monoclonal antibody Fab19 reveal a direct relationship between association rate and neutralizing potency.

    PubMed

    Bates, John T; Keefer, Christopher J; Utley, Thomas J; Correia, Bruno E; Schief, William R; Crowe, James E

    2013-04-01

    The role of affinity in determining neutralizing potency of mAbs directed against viruses is not well understood. We investigated the kinetic, structural, and functional advantage conferred by individual naturally occurring somatic mutations in the Ab H chain V region of Fab19, a well-described neutralizing human mAb directed to respiratory syncytial virus. Comparison of the affinity-matured Ab Fab19 with recombinant Fab19 Abs that were variants containing reverted amino acids from the inferred unmutated ancestor sequence revealed the molecular basis for affinity maturation of this Ab. Enhanced binding was achieved through mutations in the third H chain CDR (HCDR3) that conferred a markedly faster on-rate and a desirable increase in antiviral neutralizing activity. In contrast, most somatic mutations in the HCDR1 and HCDR2 regions did not significantly enhance Ag binding or antiviral activity. We observed a direct relationship between the measured association rate (Kon) for F protein and antiviral activity. Modeling studies of the structure of the Ag-Ab complex suggested the HCDR3 loop interacts with the antigenic site A surface loop of the respiratory syncytial virus F protein, previously shown to contain the epitope for this Ab by experimentation. These studies define a direct relationship of affinity and neutralizing activity for a viral glycoprotein-specific human mAb.

  1. Polyclonal and monoclonal antibodies specific for the six-helix bundle of the human respiratory syncytial virus fusion glycoprotein as probes of the protein post-fusion conformation

    SciTech Connect

    Palomo, Concepción; Mas, Vicente; Vázquez, Mónica; Cano, Olga; Luque, Daniel; Terrón, María C.; Calder, Lesley J.; Melero, José A.

    2014-07-15

    Human respiratory syncytial virus (hRSV) has two major surface glycoproteins (G and F) anchored in the lipid envelope. Membrane fusion promoted by hRSV{sub F} occurs via refolding from a pre-fusion form to a highly stable post-fusion state involving large conformational changes of the F trimer. One of these changes results in assembly of two heptad repeat sequences (HRA and HRB) into a six-helix bundle (6HB) motif. To assist in distinguishing pre- and post-fusion conformations of hRSV{sub F}, we have prepared polyclonal (α-6HB) and monoclonal (R145) rabbit antibodies specific for the 6HB. Among other applications, these antibodies were used to explore the requirements of 6HB formation by isolated protein segments or peptides and by truncated mutants of the F protein. Site-directed mutagenesis and electron microscopy located the R145 epitope in the post-fusion hRSV{sub F} at a site distantly located from previously mapped epitopes, extending the repertoire of antibodies that can decorate the F molecule. - Highlights: • Antibodies specific for post-fusion respiratory syncytial virus fusion protein are described. • Polyclonal antibodies were obtained in rabbit inoculated with chimeric heptad repeats. • Antibody binding required assembly of a six-helix bundle in the post-fusion protein. • A monoclonal antibody with similar structural requirements is also described. • Binding of this antibody to the post-fusion protein was visualized by electron microscopy.

  2. Morphogenesis of respiratory syncytial virus in human primary nasal ciliated epithelial cells occurs at surface membrane microdomains that are distinct from cilia

    SciTech Connect

    Jumat, Muhammad Raihan; Yan, Yan; Ravi, Laxmi Iyer; Wong, Puisan; Huong, Tra Nguyen; Li, Chunwei; Tan, Boon Huan; Wang, De Yun; Sugrue, Richard J.

    2015-10-15

    The distribution of cilia and the respiratory syncytial virus (RSV) nucleocapsid (N) protein, fusion (F) protein, attachment (G) protein, and M2-1 protein in human ciliated nasal epithelial cells was examined at between 1 and 5 days post-infection (dpi). All virus structural proteins were localized at cell surface projections that were distinct from cilia. The F protein was also trafficked into the cilia, and while its presence increased as the infection proceeded, the N protein was not detected in the cilia at any time of infection. The presence of the F protein in the cilia correlated with cellular changes in the cilia and reduced cilia function. At 5 dpi extensive cilia loss and further reduced cilia function was noted. These data suggested that although RSV morphogenesis occurs at non-cilia locations on ciliated nasal epithelial cells, RSV infection induces changes in the cilia body that leads to extensive cilia loss. - Highlights: • Respiratory syncytial virus (RSV) infects nasal ciliated epithelial cells. • Virus morphogenesis occurs within filamentous projections distinct from cilia. • The RSV N protein was not detected in the cilia at any time during infection. • Trafficking of the F protein into the cilia occurred early in infection. • Presence of the F protein in cilia correlated with impaired cilia function.

  3. Sequence analysis of a functional polymerase (L) gene of bovine respiratory syncytial virus: determination of minimal trans-acting requirements for RNA replication.

    PubMed

    Yunus, A S; Collins, P L; Samal, S K

    1998-09-01

    The complete nucleotide sequence of a functional clone of the large polymerase (L) gene of bovine respiratory syncytial virus (BRSV) strain A51908 was determined by analysis of cloned cDNAs obtained from genomic and mRNAs. The BRSV L gene is 6573 nt in length and the derived polypeptide has 2162 aa. Alignment of the sequences of the BRSV L gene, and its encoded protein, with sequences of the L gene and protein of human respiratory syncytial virus strain A2 showed 77% identity at the nucleotide level and 84% identity at the amino acid level. By comparison, the L gene and protein of avian pneumovirus showed only 50% identity at the nucleotide level and 64% identity at the amino acid level. A minigenome was constructed to encode a BRSV vRNA analogue containing the gene for chloramphenicol acetyltransferase (CAT) under the control of putative BRSV transcription motifs and flanked by the BRSV genomic termini. Transfection of plasmids encoding the BRSV minigenome, nucleocapsid protein (N), phosphoprotein (P) and L protein, each under the control of T7 promoter, into cells infected with a vaccinia virus recombinant expressing the T7 RNA polymerase gave rise to CAT activity and progeny with the minigenome. This result indicates that the N, P and L proteins are necessary and sufficient for transcription and replication of the BRSV minigenome and are functional. Further, inclusion of small amounts of the M2 protein along with the N, P and L proteins greatly augmented minigenome transcription.

  4. Use of anionic polymer, poly(methyl vinyl ether-maleic anhydride)-coated beads for capture of respiratory syncytial virus.

    PubMed

    Sakudo, Akikazu; Baba, Koichi; Tsukamoto, Megumi; Ikuta, Kazuyoshi

    2009-08-01

    Respiratory syncytial virus (RSV) is the single most important cause of severe lower respiratory tract infections in infants and young children, and a major public health concern in pediatrics. However, current diagnostic methods for RSV are not sufficiently sensitive. In addition, there is no simple method for enhancing RSV detection. Here, a method for capturing RSV from nasal fluid has been developed using magnetic beads coated with an anionic polymer, poly(methyl vinyl ether-maleic anhydrate). The beads were incubated with RSV-infected nasal fluid, then separated from the supernatant by applying a magnet field and washed. The adsorption [corrected] of RSV by the beads was confirmed by immunochromatography, reverse transcription-polymerase chain reaction, Western blotting and an enzyme-linked immunosorbent assay, which indicated the presence of nucleocapsid protein, fusion protein, and the viral genome of RSV on the incubated beads. Therefore, this capture method will contribute to the improvement of RSV detection.

  5. Resistance to Human Respiratory Syncytial Virus (RSV) Infection Induced by Immunization of Cotton Rats with a Recombinant Vaccinia Virus Expressing the RSV G Glycoprotein

    NASA Astrophysics Data System (ADS)

    Elango, Narayanasamy; Prince, Gregory A.; Murphy, Brian R.; Venkatesan, Sundararajan; Chanock, Robert M.; Moss, Bernard

    1986-03-01

    A cDNA copy of the G glycoprotein gene of human respiratory syncytial virus (RSV) was placed under control of a vaccinia virus promoter and inserted into the thymidine kinase locus of the vaccinia virus genome. The recombinant vaccinia virus retained infectivity and expressed a 93-kDa protein that migrated with the authentic RSV G glycoprotein upon polyacrylamide gel electrophoresis. Glycosylation of the expressed protein and transport to the cell surface were demonstrated in the absence of other RSV proteins. Cotton rats that were inoculated intradermally with the infectious recombinant virus produced serum antibody to the G glycoprotein that neutralized RSV in vitro. Furthermore, the vaccinated animals were resistant to lower respiratory tract infection upon intranasal inoculation with RSV and had reduced titers of RSV in the nose.

  6. Multicenter evaluation of BD Veritor System and RSV K-SeT for rapid detection of respiratory syncytial virus in a diagnostic laboratory setting.

    PubMed

    Jonckheere, Stijn; Verfaillie, Charlotte; Boel, An; Van Vaerenbergh, Kristien; Vanlaere, Elke; Vankeerberghen, Anne; De Beenhouwer, Hans

    2015-09-01

    The recently introduced BD Veritor System RSV laboratory kit (Becton Dickinson, Sparks, MD, USA) with automatic reading was evaluated and compared with the RSV K-SeT (Coris BioConcept, Gembloux, Belgium) for the detection of respiratory syncytial virus (RSV) using 248 nasopharyngeal aspirates of children younger than 6 years old with respiratory tract infection. Compared to reverse transcriptase polymerase chain reaction as gold standard, both tests had an identical sensitivity of 78.1% and a specificity of 96.8% and 95.8% for the BD Veritor System and RSV K-SeT, respectively. Both antigen tests can be used to reliably confirm RSV in young children. However, a negative result does not definitively exclude the presence of RSV.

  7. Antiviral Activity of TMC353121, a Respiratory Syncytial Virus (RSV) Fusion Inhibitor, in a Non-Human Primate Model

    PubMed Central

    Ispas, Gabriela; Koul, Anil; Verbeeck, Johan; Sheehan, Jennifer; Sanders-Beer, Brigitte; Roymans, Dirk; Andries, Koen; Rouan, Marie-Claude; De Jonghe, Sandra; Bonfanti, Jean-François; Vanstockem, Marc; Simmen, Kenneth; Verloes, Rene

    2015-01-01

    Background The study assessed the antiviral activity of TMC353121, a respiratory syncytial virus (RSV) fusion inhibitor, in a preclinical non-human primate challenge model with a viral shedding pattern similar to that seen in humans, following continuous infusion (CI). Methods African green monkeys were administered TMC353121 through CI, in 2 studies. Study 1 evaluated the prophylactic and therapeutic efficacy of TMC353121 at a target plasma level of 50 ng/mL (n=15; Group 1: prophylactic arm [Px50], 0.033 mg/mL TMC353121, flow rate 2.5 mL/kg/h from 24 hours pre-infection to 10 days; Group 2: therapeutic arm [Tx50], 0.033mg/mL TMC353121 from 24 hours postinfection to 8 days; Group 3: control [Vh1] vehicle, 24 hours post-infection to 8 days). Study 2 evaluated the prophylactic efficacy of TMC353121 at target plasma levels of 5 and 500 ng/mL (n=12; Group 1: prophylactic 5 arm [Px5], 0.0033 mg/mL TMC353121, flow rate 2.5 mL/kg/h from 72 hours pre-infection to 14 days; Group 2: prophylactic 500 arm [Px500], 0.33 mg/mL TMC353121; Group 3:control [Vh2] vehicle, 14 days). Bronchoalveolar lavage fluid and plasma were collected every 2 days from day 1 postinfection for pharmacokinetics and safety analysis. Findings TMC353121 showed a dose-dependent antiviral activity, varying from 1log10 reduction of peak viral load to complete inhibition of the RSV replication. Complete inhibition of RSV shedding was observed for a relatively low plasma exposure (0.39 μg/mL) and was associated with a dose-dependent reduction in INFγ, IL6 and MIP1α. TMC353121 administered as CI for 16 days was generally well-tolerated. Conclusion TMC353121 exerted dose-dependent antiviral effect ranging from full inhibition to absence of antiviral activity, in a preclinical model highly permissive for RSV replication. No new safety findings emerged from the study. PMID:26010881

  8. Appearance of a soluble form of the G protein of respiratory syncytial virus in fluids of infected cells.

    PubMed

    Hendricks, D A; Baradaran, K; McIntosh, K; Patterson, J L

    1987-06-01

    We examined the antigenic reactivities and virion associations of glycoproteins that were released into the culture fluids of cells infected with respiratory syncytial (RS) virus. Culture fluids and cell extracts were obtained from cells 24 to 30 h after they were infected with the Long strain of RS virus. Radioimmune precipitation of [3H]glucosamine-labelled glycoproteins by large glycoprotein (G)-specific or fusion protein (F)-specific monoclonal antibodies (MAbs) revealed that the G, F1 and F2 proteins were present in cell extracts but only the G protein was clearly evident in culture fluids. A glycoprotein (Mr 43K) which may be a precursor or a breakdown product of the G protein was also precipitated by the G-specific MAb from cell extracts and culture fluids. The G protein in culture fluids was slightly smaller (Mr 82K) than the G protein in cell extracts (Mr 88K). An abundant or heavily labelled, 18K glycoprotein in the fluids of virus-infected but not of mock-infected cells was weakly precipitated by the F-specific MAb; this suggested that the 18K protein shares epitopes with the fusion protein of RS virus. The absence of F1 and F2 polypeptides from culture fluids is evidence that the cells, which contained an abundance of these proteins, were intact. To determine whether any of the viral glycoproteins released by infected cells were soluble (non-virion-associated), culture fluid was subjected to rate zonal centrifugation in a 10 to 50% sucrose gradient. An assay of fractions using a MAb-capture ELISA for the nucleocapsid (N) and F proteins revealed a peak of activity, due to virions, in the centre of the gradient, and a strong signal for the N protein at the top of the gradient suggesting that N protein was released from intact cells. Radioimmune precipitation of glycoproteins from the fractions at the top of the gradient using a hyperimmune guinea-pig serum revealed the G protein and a heterogeneous band which had the electrophoretic mobility of the 43K

  9. Epidemiology and Molecular Characterization of Human Respiratory Syncytial Virus in Senegal after Four Consecutive Years of Surveillance, 2012–2015

    PubMed Central

    Cisse, El Hadj Abdel Kader; Kiori, Davy E.; Sarr, Fatoumata Diene; Sy, Sara; Goudiaby, Debora; Richard, Vincent; Niang, Mbayame Ndiaye

    2016-01-01

    Background The burden of respiratory syncytial virus (RSV) infection remains poorly defined in Africa. To address this, we carried out a descriptive and retrospective pilot study, with a focus on the epidemiology of RSV in Senegal after 4 years of surveillance. Methodology and Results From January 2012 to October 2015 swabs were collected from consenting ILI outpatients. Viral detection was performed using RV16 kit enabling direct subtyping of RSV-A and B. For the molecular characterization of HRSV, the second hypervariable region of the Glycoprotein (G) gene was targeted for sequencing. We enrolled 5338 patients with 2803 children younger than five years of age (52.5%). 610 (11.4%) were positive for RSV infection: 276 (45.2%) were group A infections, 334 (54.8%) were group B infections and 21 (3.4%) were A/B co-infections. RSV detection rate is significantly higher (P < 0.0001) in children below 5 years. We noted that the annual distribution of RSV varied substantially by season and for the predominant subtype. Globally, results show a clear circulation pattern in the second half of each year; between June and September and possibly extended into November. The majority of RSV-A strains from Senegal clustered with strains that were previously assigned NA1 and novel ON1 genotype sequences. RSV-B sequences from Senegal clustered with the BA9 genotype. At the amino acid level, RSV-A strains from Senegal show proximity with the genotype ON1 characterized by a 72 nt insertion in G, resulting in 24 extra amino acids of which 23 are duplications of aa 261–283. Conclusion Globally our results show a clear circulation pattern of RSV in the second half of each year, between June and September and possibly extending into November, with children under 5 being more susceptible. Molecular studies identified the novel strains ON1 and BA9 as the major genotypes circulating in Senegal between 2012 and 2015. PMID:27315120

  10. CX3CR1 is an important surface molecule for respiratory syncytial virus infection in human airway epithelial cells

    PubMed Central

    Chirkova, Tatiana; Lin, Songbai; Oomens, Antonius G. P.; Gaston, Kelsey A.; Boyoglu-Barnum, Seyhan; Meng, Jia; Stobart, Christopher C.; Cotton, Calvin U.; Hartert, Tina V.; Moore, Martin L.; Ziady, Assem G.

    2015-01-01

    Respiratory syncytial virus (RSV) is a major cause of severe pneumonia and bronchiolitis in infants and young children, and causes disease throughout life. Understanding the biology of infection, including virus binding to the cell surface, should help develop antiviral drugs or vaccines. The RSV F and G glycoproteins bind cell surface heparin sulfate proteoglycans (HSPGs) through heparin-binding domains. The G protein also has a CX3C chemokine motif which binds to the fractalkine receptor CX3CR1. G protein binding to CX3CR1 is not important for infection of immortalized cell lines, but reportedly is so for primary human airway epithelial cells (HAECs), the primary site for human infection. We studied the role of CX3CR1 in RSV infection with CX3CR1-transfected cell lines and HAECs with variable percentages of CX3CR1-expressing cells, and the effect of anti-CX3CR1 antibodies or a mutation in the RSV CX3C motif. Immortalized cells lacking HSPGs had low RSV binding and infection, which was increased markedly by CX3CR1 transfection. CX3CR1 was expressed primarily on ciliated cells, and ∼50 % of RSV-infected cells in HAECs were CX3CR1+. HAECs with more CX3CR1-expressing cells had a proportional increase in RSV infection. Blocking G binding to CX3CR1 with anti-CX3CR1 antibody or a mutation in the CX3C motif significantly decreased RSV infection in HAECs. The kinetics of cytokine production suggested that the RSV/CX3CR1 interaction induced RANTES (regulated on activation normal T-cell expressed and secreted protein), IL-8 and fractalkine production, whilst it downregulated IL-15, IL1-RA and monocyte chemotactic protein-1. Thus, the RSV G protein/CX3CR1 interaction is likely important in infection and infection-induced responses of the airway epithelium, the primary site of human infection. PMID:26297201

  11. Seasonal variations in Clostridium difficile infections are associated with influenza and respiratory syncytial virus activity independently of antibiotic prescriptions: a time series analysis in Quebec, Canada.

    PubMed

    Gilca, Rodica; Fortin, Elise; Frenette, Charles; Longtin, Yves; Gourdeau, Marie

    2012-02-01

    Seasonal variations in Clostridium difficile-associated diarrhea (CDAD), with a higher incidence occurring during winter months, have been reported. Although winter epidemics of respiratory viruses may be temporally associated with an increase in CDAD morbidity, we hypothesized that this association is mainly due to increased antibiotic use for respiratory infections. The objective of this study was to evaluate the effect of the two most frequent respiratory viruses (influenza virus and respiratory syncytial virus [RSV]) and antibiotics prescribed for respiratory infections (fluoroquinolones and macrolides) on the CDAD incidence in hospitals in the province of Québec, Canada. A multivariable Box-Jenkins transfer function model was built to relate monthly CDAD incidence to the monthly percentage of positive tests for influenza virus and RSV and monthly fluoroquinolone and macrolide prescriptions over a 4-year period (January 2005 to December 2008). Analysis showed that temporal variations in CDAD incidence followed temporal variations for influenza virus (P = 0.043), RSV (P = 0.004), and macrolide prescription (P = 0.05) time series with an average delay of 1 month and fluoroquinolone prescription time series with an average delay of 2 months (P = 0.01). We conclude that influenza virus and RSV circulation is independently associated with CDAD incidence after controlling for fluoroquinolone and macrolide use. This association was observed at an aggregated level and may be indicative of other phenomena occurring during wintertime.

  12. Simultaneous detection of influenza A, influenza B, and respiratory syncytial viruses and subtyping of influenza A H3N2 virus and H1N1 (2009) virus by multiplex real-time PCR.

    PubMed

    Chen, Yu; Cui, Dawei; Zheng, Shufa; Yang, Shigui; Tong, Jia; Yang, Dagan; Fan, Jian; Zhang, Jie; Lou, Bin; Li, Xuefen; Zhuge, Xiaoling; Ye, Bo; Chen, Baode; Mao, Weilin; Tan, Yajun; Xu, Genyun; Chen, Zhenjin; Chen, Nan; Li, Lanjuan

    2011-04-01

    A multiplex real-time PCR assay was developed to simultaneously detect and discriminate influenza A virus subtypes, including novel H1N1 (2009) and seasonal H3N2 virus, influenza B virus, and respiratory syncytial virus (RSV) in a single test tube, with detection sensitivity and specificity of 99% and 100%, respectively, for the four pathogens. PMID:21270233

  13. Preventing Cleavage of the Respiratory Syncytial Virus Attachment Protein in Vero Cells Rescues the Infectivity of Progeny Virus for Primary Human Airway Cultures

    PubMed Central

    Corry, Jacqueline; Johnson, Sara M.; Cornwell, Jessica

    2015-01-01

    ABSTRACT All live attenuated respiratory syncytial virus (RSV) vaccines that have advanced to clinical trials have been produced in Vero cells. The attachment (G) glycoprotein in virions produced in these cells is smaller than that produced in other immortalized cells due to cleavage. These virions are 5-fold less infectious for primary well-differentiated human airway epithelial (HAE) cell cultures. Because HAE cells are isolated directly from human airways, Vero cell-grown vaccine virus would very likely be similarly inefficient at initiating infection of the nasal epithelium following vaccination, and therefore, a larger inoculum would be required for effective vaccination. We hypothesized that Vero cell-derived virus containing an intact G protein would be more infectious for HAE cell cultures. Using protease inhibitors with increasing specificity, we identified cathepsin L to be the protease responsible for cleavage. Our evidence suggests that cleavage occurs in the late endosome or lysosome during endocytic recycling. Cathepsin L activity was 100-fold greater in Vero cells than in HeLa cells. In addition, cathepsin L was able to cleave the G protein in Vero cell-grown virions but not in HeLa cell-grown virions, suggesting a difference in G-protein posttranslational modification in the two cell lines. We identified by mutagenesis amino acids important for cleavage, and these amino acids included a likely cathepsin L cleavage site. Virus containing a modified, noncleavable G protein produced in Vero cells was 5-fold more infectious for HAE cells in culture, confirming our hypothesis and indicating the value of including such a mutation in future live attenuated RSV vaccines. IMPORTANCE Worldwide, RSV is the second leading infectious cause of infant death, but no vaccine is available. Experimental live attenuated RSV vaccines are grown in Vero cells, but during production the virion attachment (G) glycoprotein is cleaved. Virions containing a cleaved G protein

  14. Respiratory syncytial virus (RSV)

    MedlinePlus

    ... RSV often spreads quickly in crowded households and day care centers. The virus can live for a half ... The following increase the risk for RSV: Attending day care Being near tobacco smoke Having school-aged brothers ...

  15. Respiratory Syncytial Virus (RSV)

    MedlinePlus

    ... best Order bereavement materials News Moms Need Blog News & Media News Videos Mission stories Ambassadors Spotlights Tools & ... about vaccines that help keep baby healthy. GO News Moms Need Blog Read about what moms and ...

  16. Identification of cellular proteins that interact with Newcastle Disease Virus and human Respiratory Syncytial Virus by a two-dimensional virus overlay protein binding assay (VOPBA).

    PubMed

    Holguera, Javier; Villar, Enrique; Muñoz-Barroso, Isabel

    2014-10-13

    Although it is well documented that the initial attachment receptors for Newcastle Disease Virus (NDV) and Respiratory Syncytial Virus (RSV) are sialic acid-containing molecules and glycosaminoglycans respectively, the exact nature of the receptors for both viruses remains to be deciphered. Moreover, additional molecules at the host cell surface might be involved in the entry mechanism. With the aim of identifying the cellular proteins that interact with NDV and RSV at the cell surface, we performed a virus overlay protein binding assay (VOPBA). Cell membrane lysates were separated by two dimensional (2D) gel electrophoresis and electrotransferred to PVDF membranes, after which they were probed with high viral concentrations. NDV interacted with a Protein Disulfide Isomerase from chicken fibroblasts. In the case of RSV, we detected 15 reactive spots, which were identified as six different proteins, of which nucleolin was outstanding. We discuss the possible role of PDI and nucleolin in NDV and RSV entry, respectively.

  17. Optimization of Potent and Selective Quinazolinediones: Inhibitors of Respiratory Syncytial Virus That Block RNA-Dependent RNA-Polymerase Complex Activity

    PubMed Central

    2015-01-01

    A quinazolinedione-derived screening hit 2 was discovered with cellular antiviral activity against respiratory syncytial virus (CPE EC50 = 2.1 μM), moderate efficacy in reducing viral progeny (4.2 log at 10 μM), and marginal cytotoxic liability (selectivity index, SI ∼ 24). Scaffold optimization delivered analogs with improved potency and selectivity profiles. Most notable were compounds 15 and 19 (EC50 = 300–500 nM, CC50 > 50 μM, SI > 100), which significantly reduced viral titer (>400,000-fold), and several analogs were shown to block the activity of the RNA-dependent RNA-polymerase complex of RSV. PMID:25399509

  18. Comparison of a newly developed enzyme-linked immunosorbent assay with complement fixation and neutralisation tests for serology of bovine respiratory syncytial virus infections.

    PubMed

    Westenbrink, F; Brinkhof, J M; Straver, P J; Quak, J; De Leeuw, P W

    1985-05-01

    An indirect double antibody sandwich enzyme-linked immunosorbent assay (ELISA) was developed for the detection and titration of serum antibodies to bovine respiratory syncytial virus (BRSV). The ELISA was compared with a complement fixation (CF) test and a test for virus neutralising antibody in serum (virus neutralisation [VN] test). Testing sera collected in dairy herds revealed the closest correlation between the results of the ELISA and the CF test with respect to BRSV antibody titres. The VN test detected BRSV antibodies in a higher percentage of acute phase sera compared to the other two tests in field samples and in early bleedings of experimentally infected calves. However, the VN test was less effective in making a diagnosis of BRSV infections on the basis of a significant titre increase in paired sera. For this purpose the ELISA was found to be the most sensitive test.

  19. An investigation of the genetic basis of increased susceptibility to neutralization by anti-fusion glycoprotein antibody arising on passage of human respiratory syncytial virus in cell culture.

    PubMed

    Hiriote, W; Gias, E L Michael; Welsh, S H; Toms, G L

    2015-01-01

    Human respiratory syncytial virus isolates have previously been shown to exhibit resistance to neutralization by anti-fusion glycoprotein antibodies that is lost on passage in cell culture. Early passage resistant and late passage susceptible stocks of two virus isolates from different epidemics were cloned by plaque purification. Early passage stocks of both isolates yielded predominantly neutralization resistant clones while late passage stocks yielded predominantly susceptible clones. On further characterization of resistant and susceptible clones, resistant virus yields were lower and they were relatively resistant to both neutralization and fusion inhibition by anti-F murine monoclonal antibodies and were also resistant to neutralization by human sera and by Palivizumab. The full genome of resistant and susceptible clones from one of the isolates was sequenced. Four differences, confirmed by sequencing sister clones, were found between resistant and susceptible clones, one in each of the SH, G, F, and L genes.

  20. Venezuelan Equine Encephalitis Virus Replicon Particles Encoding Respiratory Syncytial Virus Surface Glycoproteins Induce Protective Mucosal Responses in Mice and Cotton Rats▿

    PubMed Central

    Mok, Hoyin; Lee, Sujin; Utley, Thomas J.; Shepherd, Bryan E.; Polosukhin, Vasiliy V.; Collier, Martha L.; Davis, Nancy L.; Johnston, Robert E.; Crowe, James E.

    2007-01-01

    Respiratory syncytial virus (RSV) is an important viral pathogen that causes severe lower respiratory tract infection in infants, the elderly, and immunocompromised individuals. There are no licensed RSV vaccines to date. To prevent RSV infection, immune responses in both the upper and lower respiratory tracts are required. Previously, immunization with Venezuelan equine encephalitis virus replicon particles (VRPs) demonstrated effectiveness in inducing mucosal protection against various pathogens. In this study, we developed VRPs encoding RSV fusion (F) or attachment (G) glycoproteins and evaluated the immunogenicity and efficacy of these vaccine candidates in mice and cotton rats. VRPs, when administered intranasally, induced surface glycoprotein-specific virus neutralizing antibodies in serum and immunoglobulin A (IgA) antibodies in secretions at the respiratory mucosa. In addition, fusion protein-encoding VRPs induced gamma interferon (IFN-γ)-secreting T cells in the lungs and spleen, as measured by reaction with an H-2Kd-restricted CD8+ T-cell epitope. In animals vaccinated with F protein VRPs, challenge virus replication was reduced below the level of detection in both the upper and lower respiratory tracts following intranasal RSV challenge, while in those vaccinated with G protein VRPs, challenge virus was detected in the upper but not the lower respiratory tract. Close examination of histopathology of the lungs of vaccinated animals following RSV challenge revealed no enhanced inflammation. Immunization with VRPs induced balanced Th1/Th2 immune responses, as measured by the cytokine profile in the lungs and antibody isotype of the humoral immune response. These results represent an important first step toward the use of VRPs encoding RSV proteins as a prophylactic vaccine for RSV. PMID:17928349

  1. Enhanced immunogenicity of a respiratory syncytial virus (RSV) F subunit vaccine formulated with the adjuvant GLA-SE in cynomolgus macaques.

    PubMed

    Patton, Kathryn; Aslam, Shahin; Shambaugh, Cindy; Lin, Rui; Heeke, Darren; Frantz, Chris; Zuo, Fengrong; Esser, Mark T; Paliard, Xavier; Lambert, Stacie L

    2015-08-26

    Respiratory syncytial virus (RSV) causes significant disease in elderly adults, but an effective vaccine is not yet available. We have previously reported that vaccines consisting of engineered respiratory syncytial virus soluble fusion protein (RSV sF) adjuvanted with glucopyranosyl lipid A (GLA) in an oil-in-water emulsion (stable emulsion [SE]) induce RSV F-specific T and B cell responses in mice and rats that protect from viral challenge. Here, we evaluated the immunogenicity of GLA-SE adjuvanted RSV sF vs unadjuvanted RSV sF vaccines in cynomolgus macaques (Macaca fascicularis). RSV F-specific IgG, RSV neutralizing antibodies, and RSV F-specific T cell IFNγ ELISPOT responses induced by GLA-SE adjuvanted RSV sF peaked at week 6 at significantly higher levels than achieved by unadjuvanted RSV sF and remained detectable through week 24, demonstrating response longevity. Two weeks after a week 24 booster immunization, humoral and cellular responses reached levels similar to those seen at the earlier peak response. Importantly, the GLA-SE adjuvanted RSV sF vaccine induced cross-neutralizing antibodies to other RSV A and B strains as well as F-specific IgA and IgG memory B cells. GLA-SE adjuvanted RSV sF was also demonstrated to drive a Th1-biased response characterized by more IFNγ than IL-4. This study indicates that a GLA-SE adjuvanted RSV sF vaccine induces robust humoral and Th1-biased cellular immunity in non-human primates and may benefit human populations at risk for RSV disease.

  2. Characterization of bovine respiratory syncytial virus proteins and mRNAs and generation of cDNA clones to the viral mRNAs.

    PubMed Central

    Lerch, R A; Stott, E J; Wertz, G W

    1989-01-01

    We have characterized the proteins and mRNAs of bovine respiratory syncytial (BRS) virus strain 391-2 and constructed cDNA clones corresponding to 9 of the 10 BRS virus mRNAs. The proteins of BRS virus-infected cells were compared with the proteins from human respiratory syncytial (HRS) virus-infected cells. Nine proteins specific to BRS virus-infected cells, corresponding to nine HRS virus proteins, were identified. Only a BRS virus polymerase protein remains to be identified. The BRS virus G glycoprotein showed major antigenic differences from the HRS virus G glycoprotein by immunoprecipitation and Western (immuno-) blot analysis, whereas the BRS virus F, N, M, and P proteins showed antigenic cross-reactivity with their HRS virus counterparts. Analysis of RNAs from BRS virus-infected cells showed virus-specific RNAs which had electrophoretic mobilities similar to those of mRNAs of HRS virus but which hybridized poorly or not at all with HRS virus-specific probes in Northern (RNA) blot analysis. To analyze the BRS virus RNAs further, cDNA clones to the BRS virus mRNAs were generated. Nine separate groups of clones were identified and shown to correspond to nine BRS virus mRNAs by Northern blot analysis. A 10th BRS virus large mRNA was identified by analogy with the HRS virus polymerase mRNA. These data show that like HRS virus, BRS virus has 10 genes coding for 10 mRNAs. Images PMID:2911122

  3. Performance of a Novel Point-of-Care Molecular Assay for Detection of Influenza A and B Viruses and Respiratory Syncytial Virus (Enigma MiniLab) in Children with Acute Respiratory Infection

    PubMed Central

    Douthwaite, Sam T.; Walker, Charlotte; Adams, Elisabeth J.; Mak, Catherine; Vecino Ortiz, Andres; Martinez-Alier, Nuria

    2015-01-01

    The performance of the Enigma MiniLab assay for influenza A and B viruses and respiratory syncytial virus (RSV) was compared to a centralized laboratory respiratory virus panel. The positive and negative percent agreement for influenza A virus, influenza B virus, and RSV were 79.2% (95% confidence interval [95% CI], 57.8 to 92.9%) and 99.4% (95% CI, 98.4 to 99.9), 100% (95% CI, 47.8 to 100%) and 100% (95% CI, 99.3 to 100%), 98.5% (95% CI, 94.6 to 99.8%) and 94.5% (95% CI, 91.9 to 96.4%), respectively. PMID:26560540

  4. Respiratory syncytial virus infection in infants with acute leukemia: a retrospective survey of the Japanese Pediatric Leukemia/Lymphoma Study Group.

    PubMed

    Hatanaka, Michiki; Miyamura, Takako; Koh, Katsuyoshi; Taga, Takashi; Tawa, Akio; Hasegawa, Daisuke; Kajihara, Ryosuke; Adachi, Souichi; Ishii, Eiichi; Tomizawa, Daisuke

    2015-12-01

    Respiratory syncytial virus (RSV) can cause life-threatening complications of lower respiratory tract infection (LRTI) in young children with malignancies, but reports remain limited. We performed a retrospective nationwide survey to clarify the current status of RSV disease among infants with hematological malignancies. Clinical course, treatment, and outcome of patients with hematological malignancies who suffered from RSV infections at the age of <24 months during anti-tumor therapy from April 2006 to March 2009 were investigated by sending a questionnaire to all member institutions of the Japanese Pediatric Leukemia/Lymphoma Study Group (JPLSG). Twelve patients with acute leukemia were identified as having experienced RSV disease. The primary diseases were acute myeloid leukemia (n = 8) and acute lymphoblastic leukemia (n = 4). RSV infection occurred pre- or during induction therapy (n = 8) and during consolidation therapy (n = 4). Eight patients developed LRTI, four of whom had severe pneumonia or acute respiratory distress syndrome; these four patients died despite receiving intensive care. In our survey, the prognosis of RSV disease in pediatric hematological malignancies was poor, and progression of LRTI in particular was associated with high mortality. In the absence of RSV-specific therapy, effective prevention and treatment strategies for severe RSV disease must be investigated.

  5. Pro-Inflammatory Cytokines in Nasopharyngeal Aspirate From Hospitalized Children With Respiratory Syncytial Virus Infection With or Without Rhinovirus Bronchiolitis, and Use of the Cytokines as Predictors of Illness Severity

    PubMed Central

    Díaz, Patricia V.; Valdivia, Gonzalo; Gaggero, Aldo A.; Bono, M.R.; Zepeda, Guillermo; Rivas, Mabel; Uasapud, Paola; Pinto, Ricardo A.; Boza, M. Lina; Guerrero, Julia

    2015-01-01

    Abstract Respiratory syncytial virus (RSV) and human rhinovirus (HRV) respiratory infection in children induce production of inflammatory interleukins (ILs) in the respiratory epithelium. As IL(s) determine the severity of illness, the purpose of this study was to identify the pro-inflammatory IL(s) that could be predictor(s) of clinical severity. One hundred and fifteen patients <2 years old with bronchiolitis due to RSV and /or HRV and 38 controls were selected from a hospital and an outpatient clinic. Clinical data of all patients were recorded. Severity was defined by the number of days with oxygen need. Nasopharyngeal aspirates (NPA) were collected to perform viral diagnosis by quantitative reverse transcription and polymerase chain reaction (qRT-PCR) and to quantify ILs: TNF-α, IL-10, IL-6, IL-1β, and IL-8, by flow cytometry. Simple and multiple regression and receiver operating characteristic (ROC) curves were used for statistical analysis. Of the patients selected 60 were single RSV, 28 RSV associated to HRV, and 27 single HRV. All patients (115) showed significantly higher IL levels when compared with controls. Levels of IL-6, IL-1β, and IL-8 detected in NPA from RSV single and associated to HRV were significantly higher than HRV infected and positively associated with days requiring O2. Levels of IL-6, IL-1β, and IL-8 detected in NPA from patients infected with RSV only or with both RSV and HRV are increased, and any of those 3 cytokines may have a predictive value for the number of days with need of supplemental oxygen. PMID:26426613

  6. Respiratory syncytial virus a and b display different temporal patterns in a 4-year prospective cross-sectional study among children with acute respiratory infection in a tropical city

    PubMed Central

    Bouzas, Maiara L.; Oliveira, Juliana R.; Fukutani, Kiyoshi F.; Borges, Igor C.; Barral, Aldina; Van der Gucht, Winke; Wollants, Elke; Van Ranst, Marc; de Oliveira, Camila I.; Van Weyenbergh, Johan; Nascimento-Carvalho, Cristiana M.

    2016-01-01

    Abstract Respiratory syncytial virus (RSV) is one of the most common etiological agents of childhood respiratory infections globally. Information on seasonality of different antigenic groups is scarce. We aimed to describe the frequency, seasonality, and age of children infected by RSV antigenic groups A (RSVA) and B (RSVB) among children with ARI in a 4-year period. Children (6–23 months old) with respiratory infection for ≤7 days were enrolled in a prospective cross-sectional study, from September, 2009 to October, 2013, in Salvador, in a tropical region of Brazil. Upon recruitment, demographic, clinical data, and nasopharyngeal aspirates (NPA) were collected. A multiplex quantitative real-time polymerase chain reaction (RT-PCR) with a group-specific primer and probeset for RSVA and RSVB was used. Seasonal distribution of infection by RSV different antigenic groups was evaluated by Prais-Wisten regression. Of 560 cases, the mean age was 11.4 ± 4.5 months and there were 287 (51.3%) girls. Overall, RSV was detected in 139 (24.8%; 95% CI: 21.4%–28.5%) cases, RSVA in 74 (13.2%; 95% CI: 10.6%–16.2%) cases, and RSVB in 67 (12.0%; 95% CI: 9.5%–14.9%) cases. Two (0.4%; 95% CI: 0.06%–1.2%) cases had coinfection. RSVA frequency was 9.6%, 18.4%, 21.6%, and 3.1% in 2010, 2011, 2012, and 2013, respectively. RSVB frequency was 19.2%, 0.7%, 1.4%, and 35.4% in the same years. RSVA was more frequently found from August to January than February to July (18.2% vs. 6.4%, P < 0.001). RSVB was more frequently found (P < 0.001) between March and June (36.0%) than July to October (1.0%) or November to February (1.6%). RSVB infection showed seasonal distribution and positive association with humidity (P = 0.02) whereas RSVA did not. RSVA was more common among children ≥1-year-old (17.8% vs. 1.8%; P = 0.02), as opposed to RSVB (11.5% vs. 12.2%; P = 0.8). One quarter of patients had RSV infection. RSVA compromised more frequently children aged ≥1 year. RSVA

  7. Formulation of the respiratory syncytial virus fusion protein with a polymer-based combination adjuvant promotes transient and local innate immune responses and leads to improved adaptive immunity.

    PubMed

    Sarkar, Indranil; Garg, Ravendra; van Drunen Littel-van den Hurk, Sylvia

    2016-09-30

    Respiratory syncytial virus (RSV) causes serious upper and lower respiratory tract infections in newborns and infants. Presently, there is no licensed vaccine against RSV. We previously reported the safety and efficacy of a novel vaccine candidate (ΔF/TriAdj) in rodent and lamb models following intranasal immunization. However, the effects of the vaccine on the innate immune system in the upper and lower respiratory tracts, when delivered intranasally, have not been characterized. In the present study, we found that ΔF/TriAdj triggered transient production of chemokines, cytokines and interferons in the nasal tissues and lungs of BALB/c mice. The types of chemokines produced were consistent with the populations of immune cells recruited, i.e. dendritic cells, macrophages and neutrophils, in the nose-associated lymphoid tissue (NALT), lung and their draining lymph nodes of the ΔF/TriAdj-immunized group. In addition, ΔF/TriAdj stimulated cellular activation with generation of mucosal and systemic antibody responses, and conferred complete protection from viral infection in the lungs upon RSV challenge. The effect of ΔF/TriAdj was short-lived in the nasal tissues and more prolonged in the lungs. In addition, both innate and adaptive immune responses were lower when mice were immunized with ΔF alone. These results suggest that ΔF/TriAdj modulates the innate mucosal environment in both upper and lower respiratory tracts, which contributes to robust adaptive immune responses and long-term protective efficacy of this novel vaccine formulation. PMID:27591951

  8. Design and validation of small interfering RNA on respiratory syncytial virus M2-2 gene: A potential approach in RNA interference on viral replication.

    PubMed

    Chin, V K; Atika Aziz, Nur A; Hudu, Shuaibu A; Harmal, Nabil S; Syahrilnizam, A; Jalilian, Farid A; Zamberi, S

    2016-10-01

    Human respiratory syncytial virus (RSV) is the leading cause of severe lower respiratory tract infection in infants and young children globally and is a significant pathogen of the elderly and immunocompromised. The M2-2 protein of respiratory syncytial virus (RSV) is particularly important in regulation of viral RNA transcription and replication that could be a potential anti-viral candidate against RSV infection. In this study, we designed and validated siRNAs that specifically target the RSV M2-2 gene. Four siRNAs targeting different regions of the M2-2 gene were designed using web tool. In-vitro evaluation of silencing effect was performed by using RSV infected Vero cell line. Viral M2-2 linked GFP recombinant plasmid was co-transfected with non-targeted siRNA, Pooled siRNA, siRNA 1, siRNA 2, siRNA 3 and siRNA 4 using synthetic cationic polymer. The silencing effect of M2-2 gene at the protein level was measured both qualitatively and quantitatively by using fluorescence microscopy and flow cytometry. Meanwhile, the silencing effect at the mRNA level was assessed by using RT-qPCR. This study showed that all four designed siRNAs can effectively and efficiently silence M2-2 gene. siRNA 2 showed the highest (98%) silencing effect on protein level and siRNA 4 with 83.1% at the mRNA level. The viral assay showed no significant cytopathic effects observed after 6days post-infection with siRNAs. In conclusion, this study showed the effectiveness of siRNA in silencing M2-2 gene both at the protein and mRNA level which could potentially be used as a novel therapeutic agent in the treatment of RSV infection. However, further study is warranted to investigate the silencing effect of M2-2 protein and inhibition of RSV infection. PMID:27432115

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

    PubMed

    Lerch, R A; Anderson, K; Amann, V L; Wertz, G W

    1991-03-01

    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

  10. Design and validation of small interfering RNA on respiratory syncytial virus M2-2 gene: A potential approach in RNA interference on viral replication.

    PubMed

    Chin, V K; Atika Aziz, Nur A; Hudu, Shuaibu A; Harmal, Nabil S; Syahrilnizam, A; Jalilian, Farid A; Zamberi, S

    2016-10-01

    Human respiratory syncytial virus (RSV) is the leading cause of severe lower respiratory tract infection in infants and young children globally and is a significant pathogen of the elderly and immunocompromised. The M2-2 protein of respiratory syncytial virus (RSV) is particularly important in regulation of viral RNA transcription and replication that could be a potential anti-viral candidate against RSV infection. In this study, we designed and validated siRNAs that specifically target the RSV M2-2 gene. Four siRNAs targeting different regions of the M2-2 gene were designed using web tool. In-vitro evaluation of silencing effect was performed by using RSV infected Vero cell line. Viral M2-2 linked GFP recombinant plasmid was co-transfected with non-targeted siRNA, Pooled siRNA, siRNA 1, siRNA 2, siRNA 3 and siRNA 4 using synthetic cationic polymer. The silencing effect of M2-2 gene at the protein level was measured both qualitatively and quantitatively by using fluorescence microscopy and flow cytometry. Meanwhile, the silencing effect at the mRNA level was assessed by using RT-qPCR. This study showed that all four designed siRNAs can effectively and efficiently silence M2-2 gene. siRNA 2 showed the highest (98%) silencing effect on protein level and siRNA 4 with 83.1% at the mRNA level. The viral assay showed no significant cytopathic effects observed after 6days post-infection with siRNAs. In conclusion, this study showed the effectiveness of siRNA in silencing M2-2 gene both at the protein and mRNA level which could potentially be used as a novel therapeutic agent in the treatment of RSV infection. However, further study is warranted to investigate the silencing effect of M2-2 protein and inhibition of RSV infection.

  11. Nonglycosylated G-Protein Vaccine Protects against Homologous and Heterologous Respiratory Syncytial Virus (RSV) Challenge, while Glycosylated G Enhances RSV Lung Pathology and Cytokine Levels

    PubMed Central

    Fuentes, Sandra; Coyle, Elizabeth M.; Golding, Hana

    2015-01-01

    ABSTRACT New efforts are under way to develop a vaccine against respiratory syncytial virus (RSV) that will provide protective immunity without the potential for vaccine-associated disease enhancement such as that observed in infants following vaccination with formalin-inactivated RSV vaccine. In addition to the F fusion protein, the G attachment surface protein is a target for neutralizing antibodies and thus represents an important vaccine candidate. However, glycosylated G protein expressed in mammalian cells has been shown to induce pulmonary eosinophilia upon RSV infection in a mouse model. In the current study, we evaluated in parallel the safety and protective efficacy of the RSV A2 recombinant unglycosylated G protein ectodomain (amino acids 67 to 298) expressed in Escherichia coli (REG) and those of glycosylated G produced in mammalian cells (RMG) in a mouse RSV challenge model. Vaccination with REG generated neutralizing antibodies against RSV A2 in 7/11 BALB/c mice, while RMG did not elicit neutralizing antibodies. Total serum binding antibodies against the recombinant proteins (both REG and RMG) were measured by surface plasmon resonance (SPR) and were found to be >10-fold higher for REG- than for RMG-vaccinated animals. Reduction of lung viral loads to undetectable levels after homologous (RSV-A2) and heterologous (RSV-B1) viral challenge was observed in 7/8 animals vaccinated with REG but not in RMG-vaccinated animals. Furthermore, enhanced lung pathology and elevated Th2 cytokines/chemokines were observed exclusively in animals vaccinated with RMG (but not in those vaccinated with REG or phosphate-buffered saline [PBS]) after homologous or heterologous RSV challenge. This study suggests that bacterially produced unglycosylated G protein could be developed alone or as a component of a protective vaccine against RSV disease. IMPORTANCE New efforts are under way to develop vaccines against RSV that will provide protective immunity without the potential

  12. DDA adjuvant induces a mixed Th1/Th2 immune response when associated with BBG2Na, a respiratory syncytial virus potential vaccine.

    PubMed

    Klinguer-Hamour, Christine; Libon, Christine; Plotnicky-Gilquin, Hélène; Bussat, Marie Claire; Revy, Lydie; Nguyen, Thien; Bonnefoy, Jean Yves; Corvaïa, Nathalie; Beck, Alain

    2002-06-21

    Human respiratory syncytial virus (hRSV) is one of the most common causes of respiratory infection in infants and the elderly. Previous attempts to vaccinate children against RSV failed and the induction of an aberrant Th2-type immune response was shown to induce severe to fatal pulmonary disease characterised in part by eosinophilia. BBG2Na is a promising human RSV subunit vaccine candidate which successfully passed phase II clinical trials in adults in association with Adju-Phos((R)). However, this formulation is not the most suitable for use in children since aluminium salts are known to induce a Th2-based immune response. In this study, we describe a potent and safe adjuvant formulation for BBG2Na in dimethyldioctadecylammonium bromide (DDA) that induces a mixed Th1/Th2 immune response in BALB/c mice. Furthermore, BBG2Na showed the same protective efficacy against RSV challenge when formulated either in DDA or in alum in mice and cotton rats. PMID:12034101

  13. A randomized controlled trial of motavizumab versus palivizumab for the prophylaxis of serious respiratory syncytial virus disease in children with hemodynamically significant congenital heart disease.

    PubMed

    Feltes, Timothy F; Sondheimer, Henry M; Tulloh, Robert M R; Harris, Brian S; Jensen, Kathryn M; Losonsky, Genevieve A; Griffin, M Pamela

    2011-08-01

    Children with hemodynamically significant congenital heart disease (CHD) are at risk for serious respiratory syncytial virus (RSV) disease. This study was designed to assess the safety and tolerability of motavizumab versus palivizumab in children with CHD and was not powered for efficacy. Patients (n = 1236) aged ≤24 mo were randomized to receive five monthly doses (15 mg/kg) of motavizumab or palivizumab during the RSV season. Adverse events (AEs) and serious AEs (SAEs) were recorded through 30 d after the last dose. RSV hospitalizations and RSV outpatient medically attended lower respiratory tract infections (MALRI; season 2) were summarized. Approximately 93 and 50% of patients reported an AE or SAE, respectively. Skin events occurred in 19.3% of motavizumab recipients and 16.2% of palivizumab recipients. Rates of hospitalizations and RSV MALRI were similar between treatment groups [relative risk (RR): 0.75; 95% CI, 0.34-1.59 and RR: 0.49; 95% CI, 0.10-1.99, respectively; both p > 0.05]. Motavizumab and palivizumab had similar safety profiles in children with hemodynamically significantly CHD; with the exception of skin events which were increased in motavizumab recipients. Safety and efficacy were consistent with another study comparing motavizumab with palivizumab in premature infants without CHD. PMID:21522037

  14. Respiratory Syncytial Virus whole-genome sequencing identifies convergent evolution of sequence duplication in the C-terminus of the G gene

    PubMed Central

    Schobel, Seth A.; Stucker, Karla M.; Moore, Martin L.; Anderson, Larry J.; Larkin, Emma K.; Shankar, Jyoti; Bera, Jayati; Puri, Vinita; Shilts, Meghan H.; Rosas-Salazar, Christian; Halpin, Rebecca A.; Fedorova, Nadia; Shrivastava, Susmita; Stockwell, Timothy B.; Peebles, R. Stokes; Hartert, Tina V.; Das, Suman R.

    2016-01-01

    Respiratory Syncytial Virus (RSV) is responsible for considerable morbidity and mortality worldwide and is the most important respiratory viral pathogen in infants. Extensive sequence variability within and between RSV group A and B viruses and the ability of multiple clades and sub-clades of RSV to co-circulate are likely mechanisms contributing to the evasion of herd immunity. Surveillance and large-scale whole-genome sequencing of RSV is currently limited but would help identify its evolutionary dynamics and sites of selective immune evasion. In this study, we performed complete-genome next-generation sequencing of 92 RSV isolates from infants in central Tennessee during the 2012–2014 RSV seasons. We identified multiple co-circulating clades of RSV from both the A and B groups. Each clade is defined by signature N- and O-linked glycosylation patterns. Analyses of specific RSV genes revealed high rates of positive selection in the attachment (G) gene. We identified RSV-A viruses in circulation with and without a recently reported 72-nucleotide G gene sequence duplication. Furthermore, we show evidence of convergent evolution of G gene sequence duplication and fixation over time, which suggests a potential fitness advantage of RSV with the G sequence duplication. PMID:27212633

  15. Influenza virus vaccine expressing fusion and attachment protein epitopes of respiratory syncytial virus induces protective antibodies in BALB/c mice.

    PubMed

    Bian, Chengrong; Liu, Shuzhen; Liu, Na; Zhang, Guangzhou; Xing, Li; Song, Yingwei; Duan, Yueqiang; Gu, Hongjing; Zhou, Ya; Zhang, Peirui; Li, Zhiwei; Zhang, Keming; Wang, Zhaohai; Zhang, Shaogeng; Wang, Xiliang; Yang, Penghui

    2014-04-01

    Respiratory syncytial virus (RSV) is an important viral pathogen that causes life-threatening respiratory infections in both infants and the elderly; no vaccines are at present available. In this report, we examined the use of influenza virus as a vehicle for production of an experimental RSV vaccine. We used reverse genetics to generate a recombinant influenza A virus with epitopes from the RSV fusion (F) and attachment (G) proteins (rFlu/RSV/F+G) in the influenza virus nonstructural (NS1) protein gene. Expression of RSV F+G epitope proteins was confirmed by Western blotting, and no changes in viral morphology were evident following examination by electron microscopy. BALB/c mice immunized intranasally with rFlu/RSV/F+G showed viral-specific antibody responses against both influenza and RSV. Total IgG, IgG1, IgG2a and IgA were measured in mice immunized with rFlu/RSV/F+G, revealing robust cellular and mucosal immune responses. Furthermore, we found that rFlu/RSV/F+G conferred protection against subsequent influenza and RSV challenges, showing significant decreases in viral replication and obvious attenuation of histopathological changes associated with viral infections. These findings suggest that rFlu/RSV/F+G is a promising vaccine candidate, which should be further assessed using cotton rat and primate models. PMID:24509239

  16. Natural history of human respiratory syncytial virus inferred from phylogenetic analysis of the attachment (G) glycoprotein with a 60-nucleotide duplication.

    PubMed

    Trento, Alfonsina; Viegas, Mariana; Galiano, Mónica; Videla, Cristina; Carballal, Guadalupe; Mistchenko, Alicia S; Melero, José A

    2006-01-01

    A total of 47 clinical samples were identified during an active surveillance program of respiratory infections in Buenos Aires (BA) (1999 to 2004) that contained sequences of human respiratory syncytial virus (HRSV) with a 60-nucleotide duplication in the attachment (G) protein gene. This duplication was analogous to that previously described for other three viruses also isolated in Buenos Aires in 1999 (A. Trento et al., J. Gen. Virol. 84:3115-3120, 2003). Phylogenetic analysis indicated that BA sequences with that duplication shared a common ancestor (dated about 1998) with other HRSV G sequences reported worldwide after 1999. The duplicated nucleotide sequence was an exact copy of the preceding 60 nucleotides in early viruses, but both copies of the duplicated segment accumulated nucleotide substitutions in more recent viruses at a rate apparently higher than in other regions of the G protein gene. The evolution of the viruses with the duplicated G segment apparently followed the overall evolutionary pattern previously described for HRSV, and this genotype has replaced other prevailing antigenic group B genotypes in Buenos Aires and other places. Thus, the duplicated segment represents a natural tag that can be used to track the dissemination and evolution of HRSV in an unprecedented setting. We have taken advantage of this situation to reexamine the molecular epidemiology of HRSV and to explore the natural history of this important human pathogen.

  17. Direct whole-genome deep-sequencing of human respiratory syncytial virus A and B from Vietnamese children identifies distinct patterns of inter- and intra-host evolution.

    PubMed

    Do, Lien Anh Ha; Wilm, Andreas; Van Doorn, H Rogier; Lam, Ha Minh; Sim, Shuzhen; Sukumaran, Rashmi; Tran, Anh Tuan; Nguyen, Bach Hue; Tran, Thi Thu Loan; Tran, Quynh Huong; Vo, Quoc Bao; Dac, Nguyen Anh Tran; Trinh, Hong Nhien; Nguyen, Thi Thanh Hai; Binh, Bao Tinh Le; Le, Khanh; Nguyen, Minh Tien; Thai, Quang Tung; Vo, Thanh Vu; Ngo, Ngoc Quang Minh; Dang, Thi Kim Huyen; Cao, Ngoc Huong; Tran, Thu Van; Ho, Lu Viet; Farrar, Jeremy; Jong, Menno de; Chen, Swaine; Nagarajan, Niranjan; Bryant, Juliet E; Hibberd, Martin L

    2015-12-01

    Human respiratory syncytial virus (RSV) is the major cause of lower respiratory tract infections in children ,2 years of age. Little is known about RSV intra-host genetic diversity over the course of infection or about the immune pressures that drive RSV molecular evolution. We performed whole-genome deep-sequencing on 53 RSV-positive samples (37 RSV subgroup A and 16 RSV subgroup B) collected from the upper airways of hospitalized children in southern Vietnam over two consecutive seasons. RSV A NA1 and RSV B BA9 were the predominant genotypes found in our samples, consistent with other reports on global RSV circulation during the same period. For both RSV A and B, the M gene was the most conserved, confirming its potential as a target for novel therapeutics. The G gene was the most variable and was the only gene under detectable positive selection. Further, positively selected sites inG were found in close proximity to and in some cases overlapped with predicted glycosylation motifs, suggesting that selection on amino acid glycosylation may drive viral genetic diversity. We further identified hotspots and coldspots of intra-host genetic diversity in the RSV genome, some of which may highlight previously unknown regions of functional importance.

  18. Assembly and Immunological Properties of Newcastle Disease Virus-Like Particles Containing the Respiratory Syncytial Virus F and G Proteins ▿

    PubMed Central

    McGinnes, Lori W.; Gravel, Kathryn A.; Finberg, Robert W.; Kurt-Jones, Evelyn A.; Massare, Michael J.; Smith, Gale; Schmidt, Madelyn R.; Morrison, Trudy G.

    2011-01-01

    Human respiratory syncytial virus (RSV) is a serious respiratory pathogen in infants and young children as well as elderly and immunocompromised populations. However, no RSV vaccines are available. We have explored the potential of virus-like particles (VLPs) as an RSV vaccine candidate. VLPs composed entirely of RSV proteins were produced at levels inadequate for their preparation as immunogens. However, VLPs composed of the Newcastle disease virus (NDV) nucleocapsid and membrane proteins and chimera proteins containing the ectodomains of RSV F and G proteins fused to the transmembrane and cytoplasmic domains of NDV F and HN proteins, respectively, were quantitatively prepared from avian cells. Immunization of mice with these VLPs, without adjuvant, stimulated robust, anti-RSV F and G protein antibody responses. IgG2a/IgG1 ratios were very high, suggesting predominantly TH1 responses. In contrast to infectious RSV immunization, neutralization antibody titers were robust and stable for 4 months. Immunization with a single dose of VLPs resulted in the complete protection of mice from RSV replication in lungs. Upon RSV intranasal challenge of VLP-immunized mice, no enhanced lung pathology was observed, in contrast to the pathology observed in mice immunized with formalin-inactivated RSV. These results suggest that these VLPs are effective RSV vaccines in mice, in contrast to other nonreplicating RSV vaccine candidates. PMID:20980510

  19. A new real-time RT-qPCR assay for the detection, subtyping and quantification of human respiratory syncytial viruses positive- and negative-sense RNAs.

    PubMed

    Essaidi-Laziosi, Manel; Lyon, Matthieu; Mamin, Aline; Fernandes Rocha, Mélanie; Kaiser, Laurent; Tapparel, Caroline

    2016-09-01

    Human respiratory syncytial virus (RSV) is a major health problem and the main cause of hospitalization due to bronchiolitis. RSV is divided into two antigenic subgroups, RSV-A and -B that co-circulate worldwide. Rapid and sensitive detection is desirable for proper patient handling while assessment of viral load may help to evaluate disease severity and progression. Finally RSV subtyping is needed to determine the prevalence and pathogenicity of each RSV subgroup, as well as their sensitivity to treatment. In this study, we took into account the most recent circulating RSV variants and designed two quantitative TaqMan one-step RT-PCR assays to detect and quantify both RSV subgroups separately. Standard dilutions of transcripts of positive and negative polarities were included in the assay validation to assess potential differences in sensitivity on negative-sense genomes and positive-sense RNAs. In addition, RSV detection in respiratory specimens of different types and sampled in different populations was compared to commercially available RSV diagnostic tools. Altogether, the RSV-A and -B assays revealed sensitive and quantitative over a wide range of viral loads, with a slight improved sensitivity of the RSV-B assay on positive sense transcripts, and allowed accurate RSV subtyping. We thus provide a useful tool for both RSV diagnostics and research. PMID:27180039

  20. Respiratory Syncytial Virus whole-genome sequencing identifies convergent evolution of sequence duplication in the C-terminus of the G gene.

    PubMed

    Schobel, Seth A; Stucker, Karla M; Moore, Martin L; Anderson, Larry J; Larkin, Emma K; Shankar, Jyoti; Bera, Jayati; Puri, Vinita; Shilts, Meghan H; Rosas-Salazar, Christian; Halpin, Rebecca A; Fedorova, Nadia; Shrivastava, Susmita; Stockwell, Timothy B; Peebles, R Stokes; Hartert, Tina V; Das, Suman R

    2016-01-01

    Respiratory Syncytial Virus (RSV) is responsible for considerable morbidity and mortality worldwide and is the most important respiratory viral pathogen in infants. Extensive sequence variability within and between RSV group A and B viruses and the ability of multiple clades and sub-clades of RSV to co-circulate are likely mechanisms contributing to the evasion of herd immunity. Surveillance and large-scale whole-genome sequencing of RSV is currently limited but would help identify its evolutionary dynamics and sites of selective immune evasion. In this study, we performed complete-genome next-generation sequencing of 92 RSV isolates from infants in central Tennessee during the 2012-2014 RSV seasons. We identified multiple co-circulating clades of RSV from both the A and B groups. Each clade is defined by signature N- and O-linked glycosylation patterns. Analyses of specific RSV genes revealed high rates of positive selection in the attachment (G) gene. We identified RSV-A viruses in circulation with and without a recently reported 72-nucleotide G gene sequence duplication. Furthermore, we show evidence of convergent evolution of G gene sequence duplication and fixation over time, which suggests a potential fitness advantage of RSV with the G sequence duplication. PMID:27212633

  1. Conformational studies of a short linear peptide corresponding to a major conserved neutralizing epitope of human respiratory syncytial virus fusion glycoprotein.

    PubMed

    Toiron, C; López, J A; Rivas, G; Andreu, D; Melero, J A; Bruix, M

    1996-10-01

    The conformational properties of a 21-residue peptide, corresponding to amino acids 255 to 275 (F255-275) of the human respiratory syncytial virus fusion (F) glycoprotein have been studied by CD and nmr spectroscopy. This peptide includes residues 262, 268, and 272 of the F polypeptide that are essential for integrity of most epitopes that mapped into a major antigenic site of the F molecule. CD data indicate that F255-275 adopts a random coil conformation in aqueous solution at low peptide concentrations. However, as the concentration of peptide is increased, a higher percentage of peptide molecules adopts an organized structure. This effect can be more easily observed when trifluoroethanol (30%) is added to peptide solutions, giving rise to CD spectra that resemble those of alpha-helix structures. These conformational changes were confirmed by nmr spectroscopy. The nuclear Overhauser effects observed in 30% trifluoroethanol/ water together with the conformational H alpha chemical shift data allowed us to propose a structural model of helix-loop-helix for the peptide in solution. In addition, these helical regions contain the amino acid residues essential for epitope integrity in the native F molecule. These results give new insights into the antigenic structure of the respiratory syncytical virus F glycoprotein. PMID:8837519

  2. Rapid and highly sensitive qualitative real-time assay for detection of respiratory syncytial virus A and B using NASBA and molecular beacon technology.

    PubMed

    Deiman, B; Schrover, C; Moore, C; Westmoreland, D; van de Wiel, P

    2007-12-01

    The performance of a sensitive and specific qualitative respiratory syncytial virus (RSV) assay based on NASBA technology and real-time molecular beacon detection is presented. Very low detection limits for both RSV A and RSV B were determined: 95% detection hit-rate of 95 and 47 copies/input in isolation for RSV A and RSV B, respectively. RSV was detected in a wide variety of clinical samples including respiratory swabs, nasopharyngeal aspirates (NPA), bronchoalveolar lavages (BAL), endotracheal secretions, and sputum samples. In total 779 clinical samples were tested and a valid result was obtained for 765 (RSV NASBA assay), 765 (cell culture), and 529 (rapid direct immunofluorescence testing (IF)) samples. Of these samples, 229 (RSV NASBA assay), 61 (cell culture), and 122 (IF) samples were positive for RSV. In addition, 106 samples were reported as RSV negative using the NOW RSV assay (Binax). Subsequent testing using the RSV NASBA assay demonstrated that 32 (30%) of these samples were RSV positive. The RSV NASBA assay includes a homologous internal control, which offers a high degree of standardization and quality control. When the RSV NASBA assay was performed on the NucliSens EasyQ platform (bioMérieux), test results of 48 sample extracts were obtained in less than 2h.

  3. Intranasal immunization with a replication-deficient adenoviral vector expressing the fusion glycoprotein of respiratory syncytial virus elicits protective immunity in BALB/c mice

    SciTech Connect

    Fu, Yuanhui; He, Jinsheng; Zheng, Xianxian; Wu, Qiang; Zhang, Mei; Wang, Xiaobo; Wang, Yan; Xie, Can; Tang, Qian; Wei, Wei; Wang, Min; Song, Jingdong; Qu, Jianguo; Zhang, Ying; Wang, Xin; Hong, Tao

    2009-04-17

    Human respiratory syncytial virus (RSV) is a serious pediatric pathogen of the lower respiratory tract worldwide. There is currently no clinically approved vaccine against RSV infection. Recently, it has been shown that a replication-deficient first generation adenoviral vector (FGAd), which encodes modified RSV attachment glycoprotein (G), elicits long-term protective immunity against RSV infection in mice. The major problem in developing such a vaccine is that G protein lacks MHC-I-restricted epitopes. However, RSV fusion glycoprotein (F) is a major cytotoxic T-lymphocyte epitope in humans and mice, therefore, an FGAd-encoding F (FGAd-F) was constructed and evaluated for its potential as an RSV vaccine in a murine model. Intranasal (i.n.) immunization with FGAd-F generated serum IgG, bronchoalveolar lavage secretory IgA, and RSV-specific CD8+ T-cell responses in BALB/c mice, with characteristic balanced or mixed Th1/Th2 CD4+ T-cell responses. Serum IgG was significantly elevated after boosting with i.n. FGAd-F. Upon challenge, i.n. immunization with FGAd-F displayed an effective protective role against RSV infection. These results demonstrate FGAd-F is able to induce effective protective immunity and is a promising vaccine regimen against RSV infection.

  4. Comparative evaluation of real-time PCR and conventional RT-PCR during a 2 year surveillance for influenza and respiratory syncytial virus among children with acute respiratory infections in Kolkata, India, reveals a distinct seasonality of infection.

    PubMed

    Agrawal, Anurodh S; Sarkar, Mehuli; Chakrabarti, Sekhar; Rajendran, K; Kaur, Harpreet; Mishra, Akhilesh C; Chatterjee, Mrinal K; Naik, Trailokya N; Chadha, Mandeep S; Chawla-Sarkar, Mamta

    2009-12-01

    Acute respiratory tract infections (ARTIs) are one of the most common causes of morbidity and mortality in young children worldwide. Influenza virus and respiratory syncytial virus (RSV) are the predominant aetiological agents during seasonal epidemics, and thus rapid and sensitive molecular tests for screening for such agents and timely identification of epidemics are required. This study compared real-time quantitative PCR (qPCR) with conventional RT-PCR for parallel identification of influenza A virus (IAV) or influenza B virus (IBV) and RSV. A total of 1091 respiratory samples was examined from children with suspected ARTIs between January 2007 and December 2008. Of these, 275 (25.21 %) were positive for either influenza or RSV by qPCR compared with 262 (24 .01%) positive by RT-PCR. Overall, IAV, IBV and RSV were detected in 121 (11.09 %), 59 (5.41 %) and 95 (8.71 %) samples, respectively. In spite of overlapping clinical symptoms, RSV and influenza virus showed distinct seasonal peaks. IAV correlated positively and RSV negatively with rainfall and temperature. No distinct seasonality was observed in IBV infections. This is, to the best of our knowledge, the first report of a systemic surveillance of respiratory viruses with seasonal correlation and prevalence rates from eastern India. This 2 year comparative analysis also confirmed the feasibility of using qPCR in developing countries, which will not only improve the scope for prevention of epidemics, but will also provide crucial epidemiological data from tropical regions.

  5. From the American Academy of Pediatrics: Policy statements--Modified recommendations for use of palivizumab for prevention of respiratory syncytial virus infections.

    PubMed

    2009-12-01

    Palivizumab was licensed in June 1998 by the US Food and Drug Administration for prevention of serious lower respiratory tract disease caused by respiratory syncytial virus (RSV) in pediatric patients who are at increased risk of severe disease. Safety and efficacy have been established for infants born at or before 35 weeks' gestation with or without chronic lung disease of prematurity and for infants and children with hemodynamically significant heart disease. The American Academy of Pediatrics (AAP) published a policy statement on the use of palivizumab in November 1998 (American Academy of Pediatrics, Committee on Infectious Diseases and Committee on Fetus and Newborn. Pediatrics. 1998;102[5]:1211-1216) and revised it in December 2003 (American Academy of Pediatrics, Committee on Infectious Diseases and Committee on Fetus and Newborn. Pediatrics. 2003;112[6 pt 1]:1442-1446), and an AAP technical report on palivizumab was published in 2003 (Meissner HC, Long SS; American Academy of Pediatrics, Committee on Infectious Diseases and Committee on Fetus and Newborn. Pediatrics. 2003;112[6 pt 1]:1447-1452). On the basis of the availability of additional data regarding seasonality of RSV disease as well as the limitations in available data on risk factors for identifying children who are at increased risk of serious RSV lower respiratory tract disease, AAP recommendations for immunoprophylaxis have been updated in an effort to ensure optimal balance of benefit and cost from this expensive intervention. This statement updates and replaces the 2003 AAP statement and the 2006 Red Book and is consistent with the 2009 Red Book recommendations.

  6. Insurance Status and the Risk of Severe Respiratory Syncytial Virus Disease in United States Preterm Infants Born at 32–35 Weeks Gestational Age

    PubMed Central

    Franklin, Jeremy A.; Anderson, Evan J.; Wu, Xionghua; Ambrose, Christopher S.; Simões, Eric A. F.

    2016-01-01

    Background. Database studies have identified that public health insurance status is associated with an increased risk of severe respiratory syncytial virus (RSV) disease in US infants. However, these studies did not adjust for the presence of other risk factors and did not evaluate the risk in preterm infants. Methods. In this study, we evaluate the independent association between public insurance and severe RSV disease outcomes adjusting for other risk factors. The prospective, observational RSV Respiratory Events among Preterm Infants Outcomes and Risk Tracking (REPORT) study was conducted over 2 consecutive RSV seasons at 188 US clinical sites that enrolled preterm infants born at 32–35 wGA who had not received RSV immunoprophylaxis with palivizumab. Adjusted incidence rates per 100 infant-seasons of the RSV-associated endpoints of outpatient lower respiratory tract infection (LRI), emergency department (ED) visits, RSV hospitalizations (RSVHs), and intensive care unit admissions during peak RSV season (November–March) were compared for infants with private and public insurance. Results. Of 1642 evaluable infants enrolled in the REPORT study, 50.1% had private insurance and 49.9% had public health insurance. Adjusted rates of RSV outpatient LRIs were similar; however, rates of ED visits (hazard ratio [HR], 2.04; 95% confidence interval [CI], 1.20–3.45) were higher for subjects with public insurance, with a similar but nonsignificant trend observed for hospitalization (HR, 1.61; 95% CI, .93–2.78). Conclusions. Socioeconomic status, as evaluated by public versus private healthcare insurance, is a significant independent risk factor for ED use in US preterm infants and may contribute to increased RSVHs in this population. PMID:27704018

  7. From the American Academy of Pediatrics: Policy statements--Modified recommendations for use of palivizumab for prevention of respiratory syncytial virus infections.

    PubMed

    2009-12-01

    Palivizumab was licensed in June 1998 by the US Food and Drug Administration for prevention of serious lower respiratory tract disease caused by respiratory syncytial virus (RSV) in pediatric patients who are at increased risk of severe disease. Safety and efficacy have been established for infants born at or before 35 weeks' gestation with or without chronic lung disease of prematurity and for infants and children with hemodynamically significant heart disease. The American Academy of Pediatrics (AAP) published a policy statement on the use of palivizumab in November 1998 (American Academy of Pediatrics, Committee on Infectious Diseases and Committee on Fetus and Newborn. Pediatrics. 1998;102[5]:1211-1216) and revised it in December 2003 (American Academy of Pediatrics, Committee on Infectious Diseases and Committee on Fetus and Newborn. Pediatrics. 2003;112[6 pt 1]:1442-1446), and an AAP technical report on palivizumab was published in 2003 (Meissner HC, Long SS; American Academy of Pediatrics, Committee on Infectious Diseases and Committee on Fetus and Newborn. Pediatrics. 2003;112[6 pt 1]:1447-1452). On the basis of the availability of additional data regarding seasonality of RSV disease as well as the limitations in available data on risk factors for identifying children who are at increased risk of serious RSV lower respiratory tract disease, AAP recommendations for immunoprophylaxis have been updated in an effort to ensure optimal balance of benefit and cost from this expensive intervention. This statement updates and replaces the 2003 AAP statement and the 2006 Red Book and is consistent with the 2009 Red Book recommendations. PMID:19736258

  8. Induction of mucosal and systemic immunity against respiratory syncytial virus by inactivated virus supplemented with TLR9 and NOD2 ligands.

    PubMed

    Shafique, Muhammad; Wilschut, Jan; de Haan, Aalzen

    2012-01-11

    Respiratory syncytial virus (RSV) infection is the most important viral cause of severe respiratory disease in infants and children worldwide and also forms a serious threat in the elderly. The development of RSV vaccine, however, has been hampered by the disastrous outcome of an earlier trial using an inactivated and parenterally administered RSV vaccine which did not confer protection but rather primed for enhanced disease upon natural infection. Mucosal administration does not seem to prime for enhanced disease, but non-replicating RSV antigen does not induce a strong mucosal immune response. We therefore investigated if mucosal immunization with inactivated RSV supplemented with innate receptor ligands, TLR9 (CpG ODN) and NOD2 (L18-MDP) through the upper or total respiratory tract is an effective and safe approach to induce RSV-specific immunity. Our data show that beta-propiolactone (BPL) inactivated RSV (BPL-RSV) supplemented with CpG ODN and L18-MDP potentiates activation of antigen-presenting cells (APC) in vitro, as demonstrated by NF-κB induction in a model APC cell line. In vivo, BPL-RSV supplemented with CpG ODN/L18-MDP ligands induces local IgA responses and augments Th1-signature IgG2a subtype responses after total respiratory tract (TRT), but less efficient after upper respiratory tract (intranasal, IN) immunization. Addition of TLR9/NOD2 ligands to the inactivated RSV also promoted affinity maturation of RSV-specific IgG antibodies and shifted T cell responses from mainly IL-5-secreting cells to predominantly IFN-γ-producing cells, indicating a Th1-skewed response. This effect was seen for both IN and TRT immunization. Finally, BPL-RSV supplemented with TLR9/NOD2 ligands significantly improved the protection efficacy against a challenge with infectious virus, without stimulating enhanced disease as evidenced by lack of eotaxin mRNA expression and eosinophil infiltration in the lung. We conclude that mucosal immunization with inactivated RSV

  9. Epidemiology of respiratory syncytial virus in children in Cyprus during three consecutive winter seasons (2010-2013): age distribution, seasonality and association between prevalent genotypes and disease severity.

    PubMed

    Panayiotou, C; Richter, J; Koliou, M; Kalogirou, N; Georgiou, E; Christodoulou, C

    2014-11-01

    This study reports the epidemiology of respiratory syncytial virus (RSV) in hospitalized children in Cyprus over three successive seasons (2010-2013) and the association between prevalent genotypes and disease severity. RSV infections had a circulation pattern from December to March. Most RSV-positive children (83%) were aged <2 years. Genotyping of RSV isolates showed that during the first winter season of the study (2010-2011), the only RSV genotype circulating was GA2 (RSV-A), followed by genotype BA (RSV-B) in the next winter season with only few sporadic cases of GA2. During the last winter season of the study (2012-2013) the newly emerged RSV genotype ON1 (RSV-A) was virtually the only circulating genotype. Children infected with genotype ON1 suffered a significantly milder illness compared to infections with genotypes GA2 and BA with a higher percentage of BA-infected children requiring oxygen. Our findings are in contrast to the majority of published reports that suggest RSV-A causes more severe illness than RSV-B. Therefore, further investigation of the association between RSV genotypes and disease severity is required, as it might affect treatment strategies in the future.

  10. Protein kinase R is a novel mediator of CD40 signaling and plays a critical role in modulating immunoglobulin expression during respiratory syncytial virus infection.

    PubMed

    Thakur, Sheetal A; Zalinger, Zachary B; Johnson, Teresa R; Imani, Farhad

    2011-12-01

    Effective immunoglobulin responses play a vital role in protection against most pathogens. However, the molecular mediators and mechanisms responsible for signaling and selective expression of immunoglobulin types remain to be elucidated. Previous studies in our laboratory have demonstrated that protein kinase R (PKR) plays a crucial role in IgE responses to double-stranded RNA (dsRNA) in vitro. In this study, we show that PKR plays a critical role in IgG expression both in vivo and in vitro. PKR(-/-) mice show significantly altered serum IgG levels during respiratory syncytial virus (RSV) infection. IgG2a expression is particularly sensitive to a lack of PKR and is below the detection level in mock- or RSV-infected PKR(-/-) mice. Interestingly, we show that upon activation by anti-CD40 and gamma interferon (IFN-γ), B cells from PKR(-/-) mice show diminished major histocompatibility complex class II (MHC II), CD80, and CD86 levels on the cell surface compared to wild-type (WT) mice. Our data also show that PKR is necessary for optimal expression of adhesion molecules, such as CD11a and ICAM-1, that are necessary for homotypic aggregation of B cells. Furthermore, in this report we demonstrate for the first time that upon CD40 ligation, PKR is rapidly phosphorylated and activated, indicating that PKR is an early and novel downstream mediator of CD40 signaling pathways.

  11. Determination of the disulfide bond arrangement of human respiratory syncytial virus attachment (G) protein by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    PubMed Central

    Gorman, J. J.; Ferguson, B. L.; Speelman, D.; Mills, J.

    1997-01-01

    The attachment protein or G protein of the A2 strain of human respiratory syncytial virus (RSV) was digested with trypsin and the resultant peptides separated by reverse-phase high-performance liquid chromatography (HPLC). One tryptic peptide produced a mass by matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS) corresponding to residues 152-187 with the four Cys residues of the ectodomain (residues 173, 176, 182, and 186) in disulfide linkage and absence of glycosylation. Sub-digestion of this tryptic peptide with pepsin and thermolysin produced peptides consistent with disulfide bonds between Cys173 and Cys186 and between Cys176 and Cys182. Analysis of ions produced by post-source decay of a peptic peptide during MALDI-TOF-MS revealed fragmentation of peptide bonds with minimal fission of an inter-chain disulfide bond. Ions produced by this unprecedented MALDI-induced post-source fragmentation corroborated the existence of the disulfide arrangement deduced from mass analysis of proteolysis products. These findings indicate that the ectodomain of the G protein has a non-glycosylated subdomain containing a "cystine noose." PMID:9194191

  12. Comparison of affinity chromatography and adsorption to vaccinia virus recombinant infected cells for depletion of antibodies directed against respiratory syncytial virus glycoproteins present in a human immunoglobulin preparation.

    PubMed

    Sastre, Patricia; Melero, José A; García-Barreno, Blanca; Palomo, Concepción

    2005-06-01

    Antibodies directed against human respiratory syncytial virus (HRSV) glycoproteins were depleted from a commercial immunoglobulin preparation (RespiGam) by two different methods. The first method consisted of repeated adsorption of RespiGam to Sepharose beads with covalently bound soluble forms of the two major viral glycoproteins (F or G). The second method consisted of adsorption of immunoglobulins to live cells expressing F or G glycoproteins on their surfaces after infection with vaccinia virus recombinants. While the first method removed efficiently antibodies that reacted with F and/or G glycoproteins by ELISA, it was inefficient in the elimination of anti-HRSV neutralizing antibodies. In contrast, the second method removed efficiently anti-HRSV antibodies that both reacted by ELISA and neutralized virus infectivity. These results confirm that human neutralizing antibodies are directed exclusively against HRSV F and G glycoproteins, and, they raise the possibility that F and G glycoproteins inserted into cell membranes differ antigenically from their soluble forms linked covalently to Sepharose beads.

  13. Epidemiology of respiratory syncytial virus in children in Cyprus during three consecutive winter seasons (2010-2013): age distribution, seasonality and association between prevalent genotypes and disease severity.

    PubMed

    Panayiotou, C; Richter, J; Koliou, M; Kalogirou, N; Georgiou, E; Christodoulou, C

    2014-11-01

    This study reports the epidemiology of respiratory syncytial virus (RSV) in hospitalized children in Cyprus over three successive seasons (2010-2013) and the association between prevalent genotypes and disease severity. RSV infections had a circulation pattern from December to March. Most RSV-positive children (83%) were aged <2 years. Genotyping of RSV isolates showed that during the first winter season of the study (2010-2011), the only RSV genotype circulating was GA2 (RSV-A), followed by genotype BA (RSV-B) in the next winter season with only few sporadic cases of GA2. During the last winter season of the study (2012-2013) the newly emerged RSV genotype ON1 (RSV-A) was virtually the only circulating genotype. Children infected with genotype ON1 suffered a significantly milder illness compared to infections with genotypes GA2 and BA with a higher percentage of BA-infected children requiring oxygen. Our findings are in contrast to the majority of published reports that suggest RSV-A causes more severe illness than RSV-B. Therefore, further investigation of the association between RSV genotypes and disease severity is required, as it might affect treatment strategies in the future. PMID:24476750

  14. Cell fusion assay by expression of respiratory syncytial virus (RSV) fusion protein to analyze the mutation of palivizumab-resistant strains.

    PubMed

    Yasui, Yosuke; Yamaji, Yoshiaki; Sawada, Akihito; Ito, Takashi; Nakayama, Tetsuo

    2016-05-01

    Respiratory syncytial virus (RSV) consists of fusion (F), glyco (G), and small hydrophobic (SH) proteins as envelope proteins, and infects through cell fusion. F protein is expressed on the surface of infected cells, and induces cell fusion. In the present report, expression plasmids of the F, G and SH proteins were constructed and cell fusion activity was investigated under T7 RNA polymerase. F protein alone induced cell fusion at a lower concentration than previously reported, and co-expression of F and SH proteins induced cell fusion more efficiently than F protein alone. Palivizumab is the only prophylactic agent against RSV infection. Palivizumab-resistant strains having mutations of the F protein of K272E and S275F were reported. These mutations were introduced into an F-expression plasmid, and exhibited no inhibition of cell fusion with palivizumab. Among the RSV F protein mutants, N276S has been reported to have partial resistance against palivizumab, but the F expression plasmid with the N276S mutation showed a reduction in cell fusion in the presence of palivizumab, showing no resistance to palivizumab. The present expression system was useful for investigating the mechanisms of RSV cell fusion. PMID:26794681

  15. Ginseng diminishes lung disease in mice immunized with formalin-inactivated respiratory syncytial virus after challenge by modulating host immune responses.

    PubMed

    Lee, Jong Seok; Cho, Min Kyoung; Hwang, Hye Suk; Ko, Eun-Ju; Lee, Yu-Na; Kwon, Young-Man; Kim, Min-Chul; Kim, Ki-Hye; Lee, Young-Tae; Jung, Yu-Jin; Kang, Sang-Moo

    2014-11-01

    Formalin-inactivated respiratory syncytial virus (FI-RSV) immunization is known to cause severe pulmonary inflammatory disease after subsequent RSV infection. Ginseng has been used in humans for thousands of years due to its potential health benefits. We investigated whether ginseng would have immune modulating effects on RSV infection in mice previously immunized with FI-RSV. Oral administration of mice with ginseng increased IgG2a isotype antibody responses to FI-RSV immunization, indicating T-helper type 1 (Th1) immune responses. Ginseng-treated mice that were nonimmunized or previously immunized with FI-RSV showed improved protection against RSV challenge compared with control mice without ginseng treatment. Ginseng-mediated improved clinical outcomes after live RSV infection were evidenced by diminished weight losses, decreased interleukin-4 cytokine production but increased interferon-γ production, modulation of CD3 T-cell populations toward a Th1 response, and reduced inflammatory response. Ginseng-mediated protective host immune modulation against RSV pulmonary inflammation was observed in different strains of wild-type and mutant mice. These results indicate that ginseng can modulate host immune responses to FI-RSV immunization and RSV infection, resulting in protective effects against pulmonary inflammatory disease.

  16. Molecular evolution of the hypervariable region of the attachment glycoprotein gene in human respiratory syncytial virus subgroup B genotypes BA9 and BA10.

    PubMed

    Nagasawa, Koo; Hirano, Eiko; Kobayashi, Miho; Ryo, Akihide; Oishi, Kazunori; Obuchi, Masatsugu; Ishiwada, Naruhiko; Noda, Masahiro; Kuroda, Makoto; Shimojo, Naoki; Kimura, Hirokazu

    2015-12-01

    We studied the molecular evolution of the C-terminal 3rd hypervariable region in the attachment glycoprotein gene of human respiratory syncytial virus subgroup B (HRSV-B) genotypes BA9 and BA10. We performed time-scaled phylogenetic analyses using Bayesian Markov chain Monte Carlo methods. We also performed a genetic distance analysis (p-distance analysis), positive and negative selection analyses, and a Bayesian skyline plot (BSP) analysis. We found that genotype BA9 diverged from the common ancestor of genotypes BA7, BA8, and BA10, while genotype BA10 diverged from the ancestor of genotypes BA7 and BA8. Strains of both genotypes were distributed worldwide. BA9 and BA10 diverged between 1999 and 2001. Both BA9 and BA10 evolved rapidly (about 4.8×10(-3)substitutions/site/year) and formed three distinct lineages in a 10-year period. BA10 strains belonging to lineage 3 had large genetic distances (p-distance>0.07). Thus, it may be possible to classify these strains as a new genotype, BA11. No positive selection site was detected in either genotype. Phylodynamic analyses showed that the effective population size of BA10 decreased gradually since 2010 and BA9 slightly decreased since 2009. The results suggested that the recently prevalent HRSV-B genotypes BA9 and BA10 evolved uniquely, leading to epidemics of HRSV-B worldwide over a 15-year period. PMID:26408340

  17. Evaluation of Measles Vaccine Virus as a Vector to Deliver Respiratory Syncytial Virus Fusion Protein or Epstein-Barr Virus Glycoprotein gp350

    PubMed Central

    Mok, Hoyin; Cheng, Xing; Xu, Qi; Zengel, James R; Parhy, Bandita; Zhao, Jackie; Wang, C. Kathy; Jin, Hong

    2012-01-01

    Live attenuated recombinant measles vaccine virus (MV) Edmonston-Zagreb (EZ) strain was evaluated as a viral vector to express the ectodomains of fusion protein of respiratory syncytial virus (RSV F) or glycoprotein 350 of Epstein-Barr virus (EBV gp350) as candidate vaccines for prophylaxis of RSV and EBV. The glycoprotein gene was inserted at the 1st or the 3rd position of the measles virus genome and the recombinant viruses were generated. Insertion of the foreign gene at the 3rd position had a minimal impact on viral replication in vitro. RSV F or EBV gp350 protein was secreted from infected cells. In cotton rats, EZ-RSV F and EZ-EBV gp350 induced MV- and insert-specific antibody responses. In addition, both vaccines also induced insert specific interferon gamma (IFN-γ) secreting T cell response. EZ-RSV F protected cotton rats from pulmonary replication of RSV A2 challenge infection. In rhesus macaques, although both EZ-RSV F and EZ-EBV gp350 induced MV specific neutralizing antibody responses, only RSV F specific antibody response was detected. Thus, the immunogenicity of the foreign antigens delivered by measles vaccine virus is dependent on the nature of the insert and the animal models used for vaccine evaluation. PMID:22383906

  18. Inability to evoke a long-lasting protective immune response to respiratory syncytial virus infection in mice correlates with ineffective nasal antibody responses.

    PubMed

    Singleton, Richard; Etchart, Nathalie; Hou, Sam; Hyland, Lisa

    2003-11-01

    Long-lasting protective antibody is not normally generated in children following primary respiratory syncytial virus (RSV) infection, frequently leading to reinfection. We used the BALB/c mouse model to examine the role of the nasal-associated lymphoid tissue and the bone marrow in the generation of RSV-specific long-lasting plasma cells, with a view to further understanding the mechanisms responsible for the poorly sustained RSV antibody levels following primary infection. We show here that substantial numbers of RSV-specific plasma cells were generated in the bone marrow following challenge, which were maintained thereafter. In contrast, in the nasal-associated lymphoid tissue, RSV-specific plasma cell numbers waned quickly both after primary infection and after challenge and were not maintained at a higher level after boosting. These data indicate that the inability to generate a robust local mucosal response in the nasal tissues may contribute substantially to the likelihood of subsequent reinfection and that the presence of serum anti-RSV antibody without local protection is not enough to protect against reinfection. PMID:14557616

  19. Evaluation of Nanogen MGB Alert Detection Reagents in a multiplex real-time PCR for influenza virus types A and B and respiratory syncytial virus.

    PubMed

    Hymas, Weston C; Hillyard, David R

    2009-03-01

    A multiplex real-time RT-PCR assay that detects influenza A, influenza B and respiratory syncytial virus (RSV) using the MGB Alert Influenza A&B/RSV Detection Reagent RUO (Nanogen, San Diego, CA) was developed. The Nanogen detection reagents consist of PCR primers and minor groove binder-conjugated hybridization probes for each virus and an internal control. Virus typing was determined by post-PCR melt curve analysis. A non-competitive armored RNA internal control was co-extracted with each sample to monitor nucleic acid extraction and RT-PCR. The assay was evaluated using a collection of culture, DFA and RT-PCR (Hexaplex, Prodesse, Waukesha, WI) positive and negative samples. The real-time multiplex assay detected 236 of 237 positive specimens for a 99% correlation. Of 30 Hexaplex negative samples tested, the multiplex real-time assay detected an additional 7 positives confirmed using additional PCR assays. Melt curve analysis for each virus produced average melting peaks of 60.4 degrees C, 66.7 degrees C and 69.4 degrees C for influenza A, influenza B and RSV respectively. Sequence analysis of 7 influenza A samples producing aberrant melt curves, confirmed the presence of a single nucleotide polymorphism beneath the influenza A probe. The limit of detection of each virus in 4 different sample types was measured to be between 7 and 806 copies. Overall, the multiplexed real-time RT-PCR assay was sensitive, robust and easy to use.

  20. Ten years of global evolution of the human respiratory syncytial virus BA genotype with a 60-nucleotide duplication in the G protein gene.

    PubMed

    Trento, Alfonsina; Casas, Inmaculada; Calderón, Ana; Garcia-Garcia, Maria L; Calvo, Cristina; Perez-Breña, Pilar; Melero, José A

    2010-08-01

    The emergence of natural isolates of human respiratory syncytial virus group B (HRSV-B) with a 60-nucleotide (nt) duplication in the G protein gene in Buenos Aires, Argentina, in 1999 (A. Trento et al., J. Gen. Virol. 84:3115-3120, 2003) and their dissemination worldwide allowed us to use the duplicated segment as a natural tag to examine in detail the evolution of HRSV during propagation in its natural host. Viruses with the duplicated segment were all clustered in a new genotype, named BA (A. Trento et al., J. Virol. 80:975-984, 2006). To obtain information about the prevalence of these viruses in Spain, we tested for the presence of the duplicated segment in positive HRSV-B clinical samples collected at the Severo Ochoa Hospital (Madrid) during 12 consecutive epidemics (1996-1997 to 2007-2008). Viruses with the 60-nt duplication were found in 61 samples, with a high prevalence relative to the rest of B genotypes in the most recent seasons. Global phylogenetic and demographic analysis of all G sequences containing the duplication, collected across five continents up until April 2009, revealed that the prevalence of the BA genotype increased gradually until 2004-2005, despite its rapid dissemination worldwide. After that date and coinciding with a bottleneck effect on the population size, a relatively new BA lineage (BA-IV) replaced all other group B viruses, suggesting further adaptation of the BA genotype to its natural host.

  1. Characterization of the CD8{sup +} T cell responses directed against respiratory syncytial virus during primary and secondary infection in C57BL/6 mice

    SciTech Connect

    Lukens, Michael V.; Claassen, Erwin A.W.; Graaff, Patricia M.A. de; Dijk, Mariska E.A. van; Hoogerhout, Peter; Toebes, Mireille; Schumacher, Ton N.; Most, Robbert G. van der; Kimpen, Jan L.L.; Bleek, Grada M. van . E-mail: g.vanbleek@umcutrecht.nl

    2006-08-15

    The BALB/c mouse model for human respiratory syncytial virus infection has contributed significantly to our understanding of the relative role for CD4{sup +} and CD8{sup +} T cells to immune protection and pathogenic immune responses. To enable comparison of RSV-specific T cell responses in different mouse strains and allow dissection of immune mechanisms by using transgenic and knockout mice that are mostly available on a C57BL/6 background, we characterized the specificity, level and functional capabilities of CD8{sup +} T cells during primary and secondary responses in lung parenchyma, airways and spleens of C57BL/6 mice. During the primary response, epitopes were recognized originating from the matrix, fusion, nucleo- and attachment proteins, whereas the secondary response focused predominantly on the matrix epitope. C57BL/6 mice are less permissive for hRSV infection than BALB/c mice, yet we found CD8{sup +} T cell responses in the lungs and bronchoalveolar lavage, comparable to the responses described for BALB/c mice.

  2. Combined virus-like particle and fusion protein-encoding DNA vaccination of cotton rats induces protection against respiratory syncytial virus without causing vaccine-enhanced disease.

    PubMed

    Hwang, Hye Suk; Lee, Young-Tae; Kim, Ki-Hye; Park, Soojin; Kwon, Young-Man; Lee, Youri; Ko, Eun-Ju; Jung, Yu-Jin; Lee, Jong Seok; Kim, Yu-Jin; Lee, Yu-Na; Kim, Min-Chul; Cho, Minkyoung; Kang, Sang-Moo

    2016-07-01

    A safe and effective vaccine against respiratory syncytial virus (RSV) should confer protection without causing vaccine-enhanced disease. Here, using a cotton rat model, we investigated the protective efficacy and safety of an RSV combination vaccine composed of F-encoding plasmid DNA and virus-like particles containing RSV fusion (F) and attachment (G) glycoproteins (FFG-VLP). Cotton rats with FFG-VLP vaccination controlled lung viral replication below the detection limit, and effectively induced neutralizing activity and antibody-secreting cell responses. In comparison with formalin inactivated RSV (FI-RSV) causing severe RSV disease after challenge, FFG-VLP vaccination did not cause weight loss, airway hyper-responsiveness, IL-4 cytokines, histopathology, and infiltrates of proinflammatory cells such as eosinophils. FFG-VLP was even more effective in preventing RSV-induced pulmonary inflammation than live RSV infections. This study provides evidence that FFG-VLP can be developed into a safe and effective RSV vaccine candidate. PMID:27123586

  3. Replacement of the respiratory syncytial virus nonstructural proteins NS1 and NS2 by the V protein of parainfluenza virus 5

    SciTech Connect

    Tran, Kim C.; He, Biao; Teng, Michael N.

    2007-11-10

    Paramyxoviruses have been shown to produce proteins that inhibit interferon production and signaling. For human respiratory syncytial virus (RSV), the nonstructural NS1 and NS2 proteins have been shown to have interferon antagonist activity through an unknown mechanism. To understand further the functions of NS1 and NS2, we generated recombinant RSV in which both NS1 and NS2 were replaced by the PIV5 V protein, which has well-characterized IFN antagonist activities ({delta}NS1/2-V). Expression of V was able to partially inhibit IFN responses in {delta}NS1/2-V-infected cells. In addition, the replication kinetics of {delta}NS1/2-V were intermediate between {delta}NS1/2 and wild-type (rA2) in A549 cells. However, expression of V did not affect the ability of {delta}NS1/2-V to activate IRF3 nuclear translocation and IFN{beta} transcription. These data indicate that V was able to replace some of the IFN inhibitory functions of the RSV NS1 and NS2 proteins, but also that NS1 and NS2 have functions in viral replication beyond IFN antagonism.

  4. Antigen-dependent proliferation and cytokine induction in respiratory syncytial virus-infected cotton rats reflect the presence of effector-memory T cells

    SciTech Connect

    Richter, Bettina W.M.; Onuska, Jaya M.; Niewiesk, Stefan; Prince, Gregory A.; Eichelberger, Maryna C. . E-mail: MarynaE@virionsystems.com

    2005-06-20

    Respiratory syncytial virus (RSV) is a major cause of lower airway disease in infants and children. Immunity to RSV is not long lasting, resulting in re-occurring infections throughout life. Effective long-lived immunity results when central-memory T cells that proliferate vigorously and secrete IL-2 are present. In contrast, effector-memory T cells that mainly produce IFN-{gamma}, facilitate virus clearance but are not long lived. To identify the type of memory response induced after RSV-A (Long) infection, we characterized the kinetics of the antigen-specific immune response and identified the types of cytokines induced. RSV-specific lymphocytic proliferation following primary and secondary infection was similar, and in both cases responses waned within a short period of time. In addition, mRNA for IFN-{gamma} but not IL-2 was induced in RSV-specific CD4{sup +} T cells. This supports the idea that the presence of effector-memory rather than central-memory T cells contributes to the ineffectiveness of the immune response to RSV.

  5. Correlation of cytotoxic activity in lungs to recovery of normal and gamma-irradiated cotton rats from respiratory syncytial virus infection

    SciTech Connect

    Sun, C.S.; Wyde, P.R.; Knight, V.

    1983-10-01

    Cotton rats (Sigmodon hispidus) that were exposed to 300, 600, or 900 rads of gamma irradiation and inoculated intranasally 2 days later with respiratory syncytial virus (RSV) exhibited prolonged virus shedding and delayed humoral and cytotoxic immune responses compared with comparably inoculated nonirradiated control rats. In nonirradiated animals and in animals exposed to 300 and 600 rads, levels of virus declined and then disappeared from the lungs during the period in which cytotoxic activity was maximal in the lungs of these animals. In contrast, in the group of cotton rats exposed to 900 rads of irradiation, local cytotoxic activity remained low throughout the 11-day observation period, and virus was not eliminated from the lungs. Although virus-neutralizing antibodies in serum and lavage fluids from these animals may have been involved, correlation of antibody concentrations with virus clearance from lungs was not as evident. These data suggest that cytotoxic effector cells have a positive role in eliminating RSV from the lungs of unprimed cotton rats.

  6. Potent inhibition of respiratory syncytial virus replication using a 2-5A-antisense chimera targeted to signals within the virus genomic RNA

    PubMed Central

    Player, Mark R.; Barnard, Dale L.; Torrence, Paul F.

    1998-01-01

    The 2-5A system is a recognized mechanistic component of the antiviral action of interferon. Interferon-induced 2-5A synthetase generates 2-5A, which, in turn, activates the latent constitutive RNase L that degrades viral RNA. Chemical conjugation of 2-5A to an antisense oligonucleotide can target the 2-5A-dependent RNase L to the antisense-specified RNA and effect its selective destruction. Such a 2-5A-antisense chimera (NIH351) has been developed that targets a consensus sequence within the respiratory syncytial virus (RSV) genomic RNA. NIH351 was 50- to 90-fold more potent against RSV strain A2 than was ribavirin, the presently approved drug for clinical management of RSV infection. It was similarly active against a variety of RSV strains of both A and B subgroups and possessed a cell culture selectivity index comparable to ribavirin. In addition, the anti-RSV activity of NIH351 was shown to be virus-specific and a result of a true antisense effect, because a scrambled nucleotide sequence in the antisense domain of NIH351 caused a significant decrease in antiviral activity. The 2-5A system’s RNase L was implicated in the mechanism of action of NIH351 because a congener with a disabled 2-5A moiety was of greatly reduced anti-RSV effectiveness. These findings represent an innovative approach to the control of RSV replication. PMID:9671772

  7. Protection and mechanism of action of a novel human respiratory syncytial virus vaccine candidate based on the extracellular domain of small hydrophobic protein

    PubMed Central

    Schepens, Bert; Sedeyn, Koen; Vande Ginste, Liesbeth; De Baets, Sarah; Schotsaert, Michael; Roose, Kenny; Houspie, Lieselot; Van Ranst, Marc; Gilbert, Brian; van Rooijen, Nico; Fiers, Walter; Piedra, Pedro; Saelens, Xavier

    2014-01-01

    Infections with human respiratory syncytial virus (HRSV) occur globally in all age groups and can have devastating consequences in young infants. We demonstrate that a vaccine based on the extracellular domain (SHe) of the small hydrophobic (SH) protein of HRSV, reduced viral replication in challenged laboratory mice and in cotton rats. We show that this suppression of viral replication can be transferred by serum and depends on a functional IgG receptor compartment with a major contribution of FcγRI and FcγRIII. Using a conditional cell depletion method, we provide evidence that alveolar macrophages are involved in the protection by SHe-specific antibodies. HRSV-infected cells abundantly express SH on the cell surface and are likely the prime target of the humoral immune response elicited by SHe-based vaccination. Finally, natural infection of humans and experimental infection of mice or cotton rats does not induce a strong immune response against HRSV SHe. Using SHe as a vaccine antigen induces immune protection against HRSV by a mechanism that differs from the natural immune response and from other HRSV vaccination strategies explored to date. Hence, HRSV vaccine candidates that aim at inducing protective neutralizing antibodies or T-cell responses could be complemented with a SHe-based antigen to further improve immune protection. PMID:25298406

  8. Effect of Brazilian Propolis on Exacerbation of Respiratory Syncytial Virus Infection in Mice Exposed to Tetrabromobisphenol A, a Brominated Flame Retardant

    PubMed Central

    Takeshita, Tomomi; Toyama, Satomi; Hayashi, Yuya; Honda, Shiori; Sakamoto, Shuichi; Matsuoka, Sayuri; Hidaka, Muneaki; Tsutsumi, Shigetoshi; Yasukawa, Ken; Park, Yong Kun

    2013-01-01

    Tetrabromobisphenol A (TBBPA), a brominated flame retardant, has been found to exacerbate pneumonia in respiratory syncytial virus- (RSV-) infected mice. We examined the effect of Brazilian propolis (AF-08) on the exacerbation of RSV infection by TBBPA exposure in mice. Mice were fed a powdered diet mixed with 1% TBBPA alone, 0.02% AF-08 alone, or 1% TBBPA and 0.02% AF-08 for four weeks and then intranasally infected with RSV. TBBPA exposure increased the pulmonary virus titer and level of IFN-γ, a representative marker of pneumonia due to RSV infection, in the lungs of infected mice without toxicity. AF-08 was significantly effective in reducing the virus titers and IFN-γ level increased by TBBPA exposure. Also, AF-08 significantly reduced proinflammatory cytokine (TNF-α and IL-6) levels in the lungs of RSV-infected mice with TBBPA exposure, but Th2 cytokine (IL-4 and IL-10) levels were not evidently increased. Neither TBBPA exposure nor AF-08 treatment affected the anti-RSV antibody production in RSV-infected mice. In flow cytometry analysis, AF-08 seemed to be effective in reducing the ratio of pulmonary CD8a+ cells in RSV-infected mice with TBBPA exposure. TBBPA and AF-08 did not exhibit anti-RSV activity in vitro. Thus, AF-08 probably ameliorated pneumonia exacerbated by TBBPA exposure in RSV-infected mice by limiting excess cellular immune responses. PMID:24250719

  9. Influence of antigen conformation and mode of presentation on the antibody and protective responses against human respiratory syncytial virus: relevance for vaccine development.

    PubMed

    Melero, José A

    2016-10-01

    Human respiratory syncytial virus (hRSV) remains one of the most prevalent human pathogens for which a vaccine is still missing. After several decades of hesitant efforts, particularly after the harmful effects of a formalin-inactivated hRSV vaccine trial in the 1960s, hRSV vaccine development has received new impetus from structure-based studies of its main protective antigen: the fusion (F) glycoprotein. This article reviews studies done with hRSV F, either in pieces (e.g. epitopes) or as soluble or membrane-anchored molecules folded in different conformations or presented under different forms. Knowledge gained from these studies has provided the basis for novel vaccines that are now in different phases of development and has generated tools and reagents for developing other control measures such as prophylactic or therapeutic antibodies against this virus, which remains the most important cause of hospitalization in infants and one of the leading global causes of infant mortality.

  10. WHO consultation on Respiratory Syncytial Virus Vaccine Development Report from a World Health Organization Meeting held on 23-24 March 2015.

    PubMed

    Modjarrad, Kayvon; Giersing, Birgitte; Kaslow, David C; Smith, Peter G; Moorthy, Vasee S

    2016-01-01

    Respiratory syncytial virus (RSV) is a globally prevalent cause of lower respiratory infection in neonates and infants. Despite its disease burden, a safe and effective RSV vaccine has remained elusive. In recent years, improved understanding of RSV biology and innovations in immunogen design has resulted in the advancement of multiple vaccine candidates into the clinical development pipeline. Given the growing number of vaccines in clinical trials, the rapid pace at which they are being tested, and the likelihood that an RSV vaccine will reach the commercial market in the next 5-10 years, consensus and guidance on clinical development pathways and licensure routes are needed now, before large-scale efficacy trials commence. In pursuit of this aim, the World Health Organization convened the first RSV vaccine consultation in 15 years on the 23rd and 24th of March, 2015 in Geneva, Switzerland. The meeting's primary objective was to provide guidance on clinical endpoints and development pathways for vaccine trials with a focus on considerations of low- and middle-income countries. Meeting participants reached consensus on candidate case definitions for RSV disease, considerations for clinical efficacy endpoints, and the clinical development pathway for active and passive immunization trials in maternal and pediatric populations. The strategic focus of this meeting was on the development of high quality, safe and efficacious RSV preventive interventions for global use and included: (1) maternal/passive immunization to prevent RSV disease in infants less than 6 months; (2) pediatric immunization to prevent RSV disease in infants and young children once protection afforded by maternal immunization wanes. PMID:26100926

  11. TLR2/MyD88/NF-κB Pathway, Reactive Oxygen Species, Potassium Efflux Activates NLRP3/ASC Inflammasome during Respiratory Syncytial Virus Infection

    PubMed Central

    Mgbemena, Victoria; Tsai, Su-Yu; Chang, Te-Hung; Berton, Michael T.; Morris, Ian R.; Allen, Irving C.; Ting, Jenny P.-Y.; Bose, Santanu

    2012-01-01

    Human respiratory syncytial virus (RSV) constitute highly pathogenic virus that cause severe respiratory diseases in newborn, children, elderly and immuno-compromised individuals. Airway inflammation is a critical regulator of disease outcome in RSV infected hosts. Although “controlled” inflammation is required for virus clearance, aberrant and exaggerated inflammation during RSV infection results in development of inflammatory diseases like pneumonia and bronchiolitis. Interleukin-1β (IL-1β) plays an important role in inflammation by orchestrating the pro-inflammatory response. IL-1β is synthesized as an immature pro-IL-1β form. It is cleaved by activated caspase-1 to yield mature IL-1β that is secreted extracellularly. Activation of caspase-1 is mediated by a multi-protein complex known as the inflammasome. Although RSV infection results in IL-1β release, the mechanism is unknown. Here in, we have characterized the mechanism of IL-1β secretion following RSV infection. Our study revealed that NLRP3/ASC inflammasome activation is crucial for IL-1β production during RSV infection. Further studies illustrated that prior to inflammasome formation; the “first signal” constitutes activation of toll-like receptor-2 (TLR2)/MyD88/NF-κB pathway. TLR2/MyD88/NF-κB signaling is required for pro-IL-1β and NLRP3 gene expression during RSV infection. Following expression of these genes, two “second signals” are essential for triggering inflammasome activation. Intracellular reactive oxygen species (ROS) and potassium (K+) efflux due to stimulation of ATP-sensitive ion channel promote inflammasome activation following RSV infection. Thus, our studies have underscored the requirement of TLR2/MyD88/NF-κB pathway (first signal) and ROS/potassium efflux (second signal) for NLRP3/ASC inflammasome formation, leading to caspase-1 activation and subsequent IL-1β release during RSV infection. PMID:22295065

  12. Respiratory Syncytial Virus and Other Viral Infections among Children under Two Years Old in Southern Vietnam 2009-2010: Clinical Characteristics and Disease Severity

    PubMed Central

    Bryant, Juliet E.; Tran, Anh Tuan; Nguyen, Bach Hue; Tran, Thi Thu Loan; Tran, Quynh Huong; Vo, Quoc Bao; Tran Dac, Nguyen Anh; Trinh, Hong Nhien; Nguyen, Thi Thanh Hai; Le Binh, Bao Tinh; Le, Khanh; Nguyen, Minh Tien; Thai, Quang Tung; Vo, Thanh Vu; Ngo, Ngoc Quang Minh; Dang, Thi Kim Huyen; Cao, Ngoc Huong; Tran, Thu Van; Ho, Lu Viet; Farrar, Jeremy; de Jong, Menno; van Doorn, H. Rogier

    2016-01-01

    Background Despite a high burden of respiratory syncytial virus (RSV) infections among children, data on demographic and clinical characteristics of RSV are scarce in low and middle income countries. This study aims to describe the viral etiologies, the demographic, epidemiological, and clinical characteristics of children under two years of age who were hospitalized with a lower respiratory tract infections (LRTI), focusing on RSV (prevalence, seasonality, subgroups, viral load) and its association with disease severity. Methods A prospective study among children under two years of age, hospitalized with LRTI was conducted in two referral pediatric hospitals in Ho Chi Minh City, Vietnam, from May 2009 to December 2010. Socio-demographic, clinical data and nasopharyngeal swabs were collected on enrolment and discharge. Multiplex real-time RT-PCR (13 viruses) and quantitative RSV RT-PCR were used to identify viral pathogens, RSV load and subgroups. Results Among 632 cases, 48% were RSV positive. RSV infections occurred at younger age than three other leading viral infections i.e rhinovirus (RV), metapneumovirus (MPV), parainfluenza virus (PIV-3) and were significantly more frequent in the first 6 months of life. Clinical severity score of RSV infection was significantly higher than PIV-3 but not for RV or MPV. In multivariate analysis, RV infection was significantly associated with severity while RSV infection was not. Among RSV infections, neither viral load nor viral co-infections were significantly associated with severity. Young age and having fever at admission were significantly associated with both RSV and LRTI severity. A shift in RSV subgroup predominance was observed during two consecutive rainy seasons but was not associated with severity. Conclusion We report etiologies, the epidemiological and clinical characteristics of LRTI among hospitalized children under two years of age and risk factors of RSV and LRTI severity. PMID:27500954

  13. Replication-Competent Influenza Virus and Respiratory Syncytial Virus Luciferase Reporter Strains Engineered for Co-Infections Identify Antiviral Compounds in Combination Screens

    PubMed Central

    Yan, Dan; Weisshaar, Marco; Lamb, Kristen; Chung, Hokyung K; Lin, Michael Z; Plemper, Richard K

    2016-01-01

    Myxoviruses such as influenza A virus (IAV) and respiratory syncytial virus (RSV) are major human pathogens, mandating the development of novel therapeutics. To establish a high-throughput screening protocol for the simultaneous identification of pathogen- and host-targeted hit candidates against either or both pathogens, we have attempted coinfection of cells with IAV and RSV. However, viral replication kinetics were incompatible, RSV signal window was low, and an IAV-driven minireplicon reporter assay used in initial screens narrowed the host cell range and restricted to single-cycle infections. To overcome these limitations, we developed an RSV strain carrying firefly luciferase fused to an innovative universal small-molecule assisted shut-off domain, which boosted assay signal window, and a hyperactive fusion protein that synchronized IAV and RSV reporter expression kinetics and suppresses the identification of RSV entry inhibitors sensitive to a recently reported RSV pan-resistance mechanism. Combined with a replication-competent recombinant IAV strain harboring nano-luciferase, the assay performed well on a human respiratory cell line and supports multi-cycle infections. Miniaturized to 384-well format, the protocol was validated through screening of a set of the NIH Clinical Collection (NCC) in quadruplicate. These test screens demonstrated favorable assay parameters and reproducibility. Application to a LOPAC library of bioactive compounds in a proof-of-concept campaign detected licensed anti-myxovirus therapeutics, ribavirin and the neuraminidase inhibitor zanamivir, and identified two unexpected RSV-specific hit candidates, Fenretinide and the opioid receptor antagonist BNTX-7. Hits were evaluated in direct and orthogonal dose-response counterscreens using a standard recRSV reporter strain expressing renilla luciferase. PMID:26307636

  14. Recombinant Human Respiratory Syncytial Virus (RSV) Monoclonal Antibody Fab is Effective Therapeutically when Introduced Directly into the Lungs of RSV-Infected Mice

    NASA Astrophysics Data System (ADS)

    Crowe, James E., Jr.; Murphy, Brian R.; Chanock, Robert M.; Williamson, R. Anthony; Barbas, Carlos F., III; Burton, Dennis R.

    1994-02-01

    Previously, recombinant human respiratory syncytial virus (RSV) monoclonal antibody Fabs were generated by antigen selection from random combinatorial libraries displayed at the tip of filamentous phage. Two such Fabs, which exhibited high binding affinity for RSV F glycoprotein (a major protective antigen), were evaluated for therapeutic efficacy in infected mice just before or at the time of peak virus replication in the lungs. Fab 19, which neutralized RSV infectivity with high efficiency in tissue culture, was effective therapeutically when delivered directly into the lungs by intranasal instillation under anesthesia. In contrast, RSV Fab 126, which failed to neutralize virus in cell culture, did not exhibit a therapeutic effect under these conditions. The amount of Fab 19 required to effect a 5000- to 12,000-fold reduction in titer of RSV in the lungs within 24 hr was rather small. In four separate experiments, a single instillation of 12.9-50 μg of RSV Fab 19 was sufficient to achieve such a reduction in pulmonary virus in a 25g mouse. The use of Fabs instead of the whole immunoglobulin molecules from which they are derived reduced the protein content of a therapeutic dose. This is important because the protein load that can be delivered effectively into the lungs is limited. The therapeutic effect of a single treatment with Fab 19 was not sustained, so that a rebound in pulmonary virus titer occurred on the 2nd day after treatment. This rebound in pulmonary RSV titer could be prevented by treating infected mice with a single dose of Fab 19 daily for 3 days. These observations suggest that human monoclonal Fabs grown in Escherichia coli may prove useful in the treatment of serious RSV disease as well as diseases caused by other viruses where replication in vivo is limited primarily to the lumenal lining of the respiratory tract.

  15. Examining strain diversity and phylogeography in relation to an unusual epidemic pattern of respiratory syncytial virus (RSV) in a long-term refugee camp in Kenya

    PubMed Central

    2014-01-01

    Background A recent longitudinal study in the Dadaab refugee camp near the Kenya-Somalia border identified unusual biannual respiratory syncytial virus (RSV) epidemics. We characterized the genetic variability of the associated RSV strains to determine if viral diversity contributed to this unusual epidemic pattern. Methods For 336 RSV positive specimens identified from 2007 through 2011 through facility-based surveillance of respiratory illnesses in the camp, 324 (96.4%) were sub-typed by PCR methods, into 201 (62.0%) group A, 118 (36.4%) group B and 5 (1.5%) group A-B co-infections. Partial sequencing of the G gene (coding for the attachment protein) was completed for 290 (89.5%) specimens. These specimens were phylogenetically analyzed together with 1154 contemporaneous strains from 22 countries. Results Of the 6 epidemic peaks recorded in the camp over the period, the first and last were predominantly made up of group B strains, while the 4 in between were largely composed of group A strains in a consecutive series of minor followed by major epidemics. The Dadaab group A strains belonged to either genotype GA2 (180, 98.9%) or GA5 (2, < 1%) while all group B strains (108, 100%) belonged to BA genotype. In sequential epidemics, strains within these genotypes appeared to be of two types: those continuing from the preceding epidemics and those newly introduced. Genotype diversity was similar in minor and major epidemics. Conclusion RSV strain diversity in Dadaab was similar to contemporaneous diversity worldwide, suggested both between-epidemic persistence and new introductions, and was unrelated to the unusual epidemic pattern. PMID:24690157

  16. One-step multiplex real time RT-PCR for the detection of bovine respiratory syncytial virus, bovine herpesvirus 1 and bovine parainfluenza virus 3

    PubMed Central

    2012-01-01

    Background Detection of respiratory viruses in veterinary species has traditionally relied on virus detection by isolation or immunofluorescence and/or detection of circulating antibody using ELISA or serum neutralising antibody tests. Multiplex real time PCR is increasingly used to diagnose respiratory viruses in humans and has proved to be superior to traditional methods. Bovine respiratory disease (BRD) is one of the most common causes of morbidity and mortality in housed cattle and virus infections can play a major role. We describe here a one step multiplex reverse transcriptase quantitative polymerase chain reaction (mRT-qPCR) to detect the viruses commonly implicated in BRD. Results A mRT-qPCR assay was developed and optimised for the simultaneous detection of bovine respiratory syncytial virus (BRSV), bovine herpes virus type 1 (BoHV-1) and bovine parainfluenza virus type 3 (BPI3 i & ii) nucleic acids in clinical samples from cattle. The assay targets the highly conserved glycoprotein B gene of BoHV-1, nucleocapsid gene of BRSV and nucleoprotein gene of BPI3. This mRT-qPCR assay was assessed for sensitivity, specificity and repeatability using in vitro transcribed RNA and recent field isolates. For clinical validation, 541 samples from clinically affected animals were tested and mRT-qPCR result compared to those obtained by conventional testing using virus isolation (VI) and/or indirect fluorescent antibody test (IFAT). Conclusions The mRT-qPCR assay was rapid, highly repeatable, specific and had a sensitivity of 97% in detecting 102 copies of BRSV, BoHV-1 and BPI3 i & ii. This is the first mRT-qPCR developed to detect the three primary viral agents of BRD and the first multiplex designed using locked nucleic acid (LNA), minor groove binding (MGB) and TaqMan probes in one reaction mix. This test was more sensitive than both VI and IFAT and can replace the aforesaid methods for virus detection during outbreaks of BRD. PMID:22455597

  17. CD4+ T-Cell-Mediated Antiviral Protection of the Upper Respiratory Tract in BALB/c Mice following Parenteral Immunization with a Recombinant Respiratory Syncytial Virus G Protein Fragment

    PubMed Central

    Plotnicky-Gilquin, Hélène; Robert, Alain; Chevalet, Laurent; Haeuw, Jean-Francois; Beck, Alain; Bonnefoy, Jean-Yves; Brandt, Christian; Siegrist, Claire-Anne; Nguyen, Thien Ngoc; Power, Ultan F.

    2000-01-01

    We analyzed the protective mechanisms induced against respiratory syncytial virus subgroup A (RSV-A) infection in the lower and upper respiratory tracts (LRT and URT) of BALB/c mice after intraperitoneal immunization with a recombinant fusion protein incorporating residues 130 to 230 of RSV-A G protein (BBG2Na). Mother-to-offspring antibody (Ab) transfer and adoptive transfer of BBG2Na-primed B cells into SCID mice demonstrated that Abs are important for LRT protection but have no effect on URT infection. In contrast, RSV-A clearance in the URT was achieved in a dose-dependent fashion after adoptive transfer of BBG2Na-primed T cells, while it was abolished in BBG2Na-immunized mice upon in vivo depletion of CD4+, but not CD8+, T cells. Furthermore, the conserved RSV-A G protein cysteines and residues 193 and 194, overlapping the recently identified T helper cell epitope on the G protein (P. W. Tebbey et al., J. Exp. Med. 188:1967–1972, 1998), were found to be essential for URT but not LRT protection. Taken together, these results demonstrate for the first time that CD4+ T cells induced upon parenteral immunization with an RSV G protein fragment play a critical role in URT protection of normal mice against RSV infection. PMID:10729118

  18. Characteristics and Their Clinical Relevance of Respiratory Syncytial Virus Types and Genotypes Circulating in Northern Italy in Five Consecutive Winter Seasons.

    PubMed

    Esposito, Susanna; Piralla, Antonio; Zampiero, Alberto; Bianchini, Sonia; Di Pietro, Giada; Scala, Alessia; Pinzani, Raffaella; Fossali, Emilio; Baldanti, Fausto; Principi, Nicola

    2015-01-01

    In order to investigate the genetic diversity and patterns of the co-circulating genotypes of respiratory syncytial virus (RSV) and their possible relationships with the severity of RSV infection, we studied all of the RSV-positive nasopharyngeal samples collected from children during five consecutive winters (2009-2010, 2010-2011, 2011-2012, 2012-2013 and 2013-2014). The RSVs were detected using the respiratory virus panel fast assay and single-tube RT-PCR, their nucleotides were sequenced, and they were tested for positive selection. Of the 165 positive samples, 131 (79.4%) carried RSV-A and 34 (20.6%) RSV-B; both groups co-circulated in all of the study periods, with RSV-A predominating in all the seasons except for winter 2010-2011, which had a predominance of RSV-B. Phylogenetic analysis of the RSV-A sequences identified genotypes NA1 and ON1, the second replacing the first during the last two years of the study period. The RSV-B belonged to genotypes BA9 and BA10. BA9 was detected in all the years of the study whereas BA only desultorily. Comparison of the subjects infected by RSV-A and RSV-B types did not reveal any significant differences, but the children infected by genotype A/NA1 more frequently had lower respiratory tract infections (p<0.0001) and required hospitalisation (p = 0.007) more often than those infected by genotype A/ON1. These findings show that RSV has complex patterns of circulation characterised by the periodical replacement of the predominant genotypes, and indicate that the circulation and pathogenic role of the different RSV strains should be investigated as each may have a different impact on the host. A knowledge of the correlations between types, genotypes and disease severity may also be important in order to be able to include the more virulent strains in future vaccines.

  19. Characteristics and Their Clinical Relevance of Respiratory Syncytial Virus Types and Genotypes Circulating in Northern Italy in Five Consecutive Winter Seasons

    PubMed Central

    Esposito, Susanna; Piralla, Antonio; Zampiero, Alberto; Bianchini, Sonia; Di Pietro, Giada; Scala, Alessia; Pinzani, Raffaella; Fossali, Emilio; Baldanti, Fausto; Principi, Nicola

    2015-01-01

    In order to investigate the genetic diversity and patterns of the co-circulating genotypes of respiratory syncytial virus (RSV) and their possible relationships with the severity of RSV infection, we studied all of the RSV-positive nasopharyngeal samples collected from children during five consecutive winters (2009–2010, 2010–2011, 2011–2012, 2012–2013 and 2013–2014). The RSVs were detected using the respiratory virus panel fast assay and single-tube RT-PCR, their nucleotides were sequenced, and they were tested for positive selection. Of the 165 positive samples, 131 (79.4%) carried RSV-A and 34 (20.6%) RSV-B; both groups co-circulated in all of the study periods, with RSV-A predominating in all the seasons except for winter 2010–2011, which had a predominance of RSV-B. Phylogenetic analysis of the RSV-A sequences identified genotypes NA1 and ON1, the second replacing the first during the last two years of the study period. The RSV-B belonged to genotypes BA9 and BA10. BA9 was detected in all the years of the study whereas BA only desultorily. Comparison of the subjects infected by RSV-A and RSV-B types did not reveal any significant differences, but the children infected by genotype A/NA1 more frequently had lower respiratory tract infections (p<0.0001) and required hospitalisation (p = 0.007) more often than those infected by genotype A/ON1. These findings show that RSV has complex patterns of circulation characterised by the periodical replacement of the predominant genotypes, and indicate that the circulation and pathogenic role of the different RSV strains should be investigated as each may have a different impact on the host. A knowledge of the correlations between types, genotypes and disease severity may also be important in order to be able to include the more virulent strains in future vaccines. PMID:26047100

  20. Baculovirus vectors expressing F proteins in combination with virus-induced signaling adaptor (VISA) molecules confer protection against respiratory syncytial virus infection.

    PubMed

    Zhang, Yuan; Qiao, Lei; Hu, Xiao; Zhao, Kang; Zhang, Yanwen; Chai, Feng; Pan, Zishu

    2016-01-01

    Baculovirus has been exploited for use as a novel vaccine vector. To investigate the feasibility and efficacy of recombinant baculoviruses (rBVs) expressing respiratory syncytial virus (RSV) fusion (F) proteins, four constructs (Bac-tF/64, Bac-CF, Bac-CF/tF64 and Bac-CF/tF64-VISA) were generated. Bac-tF64 displays the F ectodomain (tF) on the envelope of rBVs, whereas Bac-CF expresses full-length F protein in transduced mammalian cells. Bac-CF/tF64 not only displays tF on the envelope but also expresses F in cells. Bac-CF/tF64-VISA comprises Bac-CF/tF64 harboring the virus-induced signaling adaptor (VISA) gene. After administration to BALB/c mice, all four vectors elicited RSV neutralizing antibody (Ab), systemic Ab (IgG, IgG1, and IgG2a), and cytokine responses. Compared with Bac-tF64, mice inoculated with Bac-CF and Bac-CF/tF64 exhibited an increased mixed Th1/Th2 cytokine response, increased ratios of IgG2a/IgG1 antibody responses, and reduced immunopathology upon RSV challenge. Intriguingly, co-expression of VISA reduced Th2 cytokine (IL-4, IL-5, and IL-10) production induced by Bac-CF/tF64, thus relieving lung pathology upon a subsequent RSV challenge. Our results indicated that the Bac-CF/tF64 vector incorporated with the VISA molecule may provide an effective vaccine strategy for protection against RSV.

  1. Small interfering RNAs targeted to interleukin-4 and respiratory syncytial virus reduce airway inflammation in a mouse model of virus-induced asthma exacerbation.

    PubMed

    Khaitov, Musa R; Shilovskiy, Igor P; Nikonova, Aleksandra A; Shershakova, Nadezda N; Kamyshnikov, Oleg Y; Babakhin, Alexander A; Zverev, Vitaly V; Johnston, Sebastian L; Khaitov, Rakhim M

    2014-07-01

    Asthma exacerbations are caused primarily by viral infections. Antisense and small interfering RNA (siRNA) technologies have gained attention as potential antiasthma and antiviral approaches. In this study we analyzed whether gene silencing of interleukin (IL)-4 expression and respiratory syncytial virus (RSV) replication by RNA interference is able to suppress allergen- and virus-induced responses in a mouse model of virus-induced asthma exacerbation. Knockdown efficacy of IL-4 siRNA molecules was analyzed in the human HEK293T cell line by cotransfection of six different siRNAs with a plasmid carrying mouse IL-4. The most potent siRNA was then used in a mouse model of RSV-induced asthma exacerbation. BALB/c mice were sensitized intraperitoneally with o