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Find video protocols related to scientific articles indexed in Pubmed.
A multiplexed droplet digital PCR assay performs better than qPCR on inhibition prone samples.
Diagn. Microbiol. Infect. Dis.
PUBLISHED: 07-04-2014
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We demonstrate the development of a multiplex droplet digital PCR assay for human cytomegalovirus (CMV), human adenovirus species F, and an internal plasmid control that may be useful for PCR inhibition-prone clinical samples. This assay performs better on inhibition-prone stool samples than a quantitative PCR assay for CMV and is the first published clinical virology droplet digital PCR assay to incorporate an internal control.
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Parainfluenza virus lower respiratory tract disease after hematopoietic cell transplant: viral detection in the lung predicts outcome.
Clin. Infect. Dis.
PUBLISHED: 03-05-2014
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Parainfluenza virus (PIV) commonly infects patients following hematopoietic cell transplantation (HCT), frequently causing lower respiratory tract disease (LRTD). The definition of LRTD significantly differs among studies evaluating the impact of PIV after HCT.
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Nosocomial transmission of respiratory syncytial virus in an outpatient cancer center.
Biol. Blood Marrow Transplant.
PUBLISHED: 01-15-2014
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Respiratory syncytial virus (RSV) outbreaks in inpatient settings are associated with poor outcomes in cancer patients. The use of molecular epidemiology to document RSV transmission in the outpatient setting has not been well described. We performed a retrospective cohort study of 2 nosocomial outbreaks of RSV at the Seattle Cancer Care Alliance. Subjects included patients seen at the Seattle Cancer Care Alliance with RSV detected in 2 outbreaks in 2007-2008 and 2012 and all employees with respiratory viruses detected in the 2007-2008 outbreak. A subset of samples was sequenced using semi-nested PCR targeting the RSV attachment glycoprotein coding region. Fifty-one cases of RSV were identified in 2007-2008. Clustering of identical viral strains was detected in 10 of 15 patients (67%) with RSV sequenced from 2007 to 2008. As part of a multimodal infection control strategy implemented as a response to the outbreak, symptomatic employees had nasal washes collected. Of 254 employee samples, 91 (34%) tested positive for a respiratory virus, including 14 with RSV. In another RSV outbreak in 2012, 24 cases of RSV were identified; 9 of 10 patients (90%) had the same viral strain, and 1 (10%) had another viral strain. We document spread of clonal strains within an outpatient cancer care setting. Infection control interventions should be implemented in outpatient, as well as inpatient, settings to reduce person-to-person transmission and limit progression of RSV outbreaks.
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Respiratory Syncytial Virus in Hematopoietic Cell Transplant Recipients: Factors Determining Progression to Lower Respiratory Tract Disease.
J. Infect. Dis.
PUBLISHED: 12-23-2013
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Background.?Respiratory syncytial virus (RSV) lower respiratory tract disease (LRD) is a life-threatening complication in hematopoietic cell transplant (HCT) recipients. Lymphopenia has been associated with an increased risk of progression from upper respiratory tract infection (URI) to LRD.Methods.?This study retrospectively analyzed the significance of lymphocyte engraftment dynamics, lung function, smoking history, corticosteroids, antiviral treatment, viral subtypes and RSV-specific neutralizing antibodies for the progression to LRD in 181 HCT recipients with RSV URI.Results.?In multivariable models, smoking history, conditioning with high dose total body irradiation, and an absolute lymphocyte count (ALC) ?100/mm(3) at the time of URI onset were significantly associated with disease progression. No progression occurred in patients with ALCs of >1,000/mm(3) at URI onset. Lymphocyte engraftment dynamics were similar in progressors and non-progressors. Pre- and posttransplant donor and posttransplant recipient RSV subtype-specific neutralizing antibody levels, RSV viral subtypes, and corticosteroids also were not significantly associated with LRD progression.Conclusions.?Host and transplant related factors appear to determine the risk of progression to LRD more than viral factors. Dysfunctional cell-mediated immunity appears to be important in the pathogenesis of progressive RSV disease after HCT. A characterization of RSV-specific T cell immunity is warranted.
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Respiratory syncytial virus lower respiratory disease in hematopoietic cell transplant recipients: viral RNA detection in blood, antiviral treatment, and clinical outcomes.
Clin. Infect. Dis.
PUBLISHED: 09-24-2013
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Background.?Respiratory syncytial virus (RSV) pneumonia after hematopoietic cell transplant (HCT) is associated with severe morbidity. Although RSV RNA has been detected in serum from patients with RSV lower respiratory disease (LRD) after HCT, the association with clinical outcomes has not been well established in multivariable models. Additionally, the role of antiviral treatment in HCT recipients has not been previously analyzed in multivariable models. Methods.?We retrospectively identified HCT recipients with virologically confirmed RSV LRD and tested stored plasma/serum samples by quantitative reverse transcription polymerase chain reaction for RSV RNA. Risk factors for RSV RNA detection and the impact of RSV RNA in serum and antiviral therapy on outcomes were analyzed using multivariable Cox models. Results.?RSV RNA was detected in plasma or serum from 28 of 92 (30%) patients at a median of 24.5 days following HCT and 2 days following LRD. In multivariable models, neutropenia, monocytopenia, thrombocytopenia, and mechanical ventilation increased the risk of plasma/serum RSV RNA detection; lymphopenia and steroid use did not. RSV RNA detection increased the risk of overall mortality in multivariable models (adjusted hazard ratio [aHR], 2.09 [P = .02]), whereas treatment with aerosolized ribavirin decreased the risk of overall mortality and pulmonary death (aHR, 0.33 [P = .001] and aHR 0.31 [P = .003], respectively). Conclusions.?RSV RNA detection in plasma or serum may be a marker for lung injury and poor outcomes in HCT recipients with RSV LRD. Treatment with aerosolized ribavirin appeared to be protective against overall and pulmonary mortality.
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Comparison of the Simplexa FluA/B & RSV direct assay and laboratory-developed real-time PCR assays for detection of respiratory virus.
J. Clin. Microbiol.
PUBLISHED: 09-18-2013
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The results of the Focus Simplexa FluA/B & RSV Direct assay were compared to those of laboratory-developed reverse transcription PCR tests for 498 nasopharyngeal swabs. Concordance rates were 96.6% (476/493; ? = 0.91), 97.6% (481/493; ? = 0.47), and 99.2% (488/492; ? = 0.94) for influenza A, influenza B, and respiratory syncytial virus, respectively.
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Home Self-Collection of Nasal Swabs for Diagnosis of Acute Respiratory Virus Infections in Children With Cystic Fibrosis.
J Pediatric Infect Dis Soc
PUBLISHED: 05-21-2013
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Understanding the importance of respiratory viruses in children with cystic fibrosis (CF) has been limited because of challenges using clinic- or hospital-based diagnostic testing. We conducted a pilot study to assess feasibility of home self- (or parent-) collection of nasal swabs (NS).
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Mortality rates of human metapneumovirus and respiratory syncytial virus lower respiratory tract infections in hematopoietic cell transplantation recipients.
Biol. Blood Marrow Transplant.
PUBLISHED: 03-18-2013
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Human metapneumovirus (HMPV), a common respiratory virus, can cause severe disease in pre- and post-hematopoietic cell transplantation (HCT) recipients. We conducted a retrospective cohort analysis in HCT patients with HMPV (n = 23) or respiratory syncytial virus (n = 23) detected in bronchoalveolar lavage samples by reverse transcription PCR between 2006 and 2011 to determine disease characteristics and factors associated with outcome. Mortality rates at 100 days were 43% for both HMPV and respiratory syncytial virus lower respiratory tract disease. Steroid therapy, oxygen requirement >2 L or mechanical ventilation, and bone marrow as cell source were significant risk factors for overall and virus-related mortality in multivariable models, whereas the virus type was not. The presence of centrilobular/nodular radiographic infiltrates was a possible protective factor for mechanical ventilation. Thus, HMPV lower respiratory tract disease is associated with high mortality in HCT recipients. Earlier detection in combination with new antiviral therapy is needed to reduce mortality among HCT recipients.
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Comparison of a multiplex real-time PCR assay with a multiplex Luminex assay for influenza virus detection.
J. Clin. Microbiol.
PUBLISHED: 01-23-2013
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We describe the development of a multiplex reverse transcription-PCR (RT-PCR) with Luminex microarray hybridization for detection of influenza virus subtypes (FLULUM). Performance of FLULUM was evaluated by comparing it to our real-time RT-PCR influenza virus assay on samples collected during two influenza seasons. Both assays targeted the matrix genes of influenza virus A (FluA M) and influenza virus B (FluB M) and the hemagglutinin genes of seasonal H3N2 (H3) and H1N1 (H1) and 2009 pandemic H1N1 (2009 H1). We evaluated FLULUM on both the Luminex LX200 and the Luminex MagPix instruments. Compared to real-time PCR, FLULUM tested on 259 specimens submitted in the 2010-2011 season showed sensitivities of 97.3% for FluA M, 90.5% for 2009 H1, 96.9% for H3, and 88.9% for FluB M. No specimens were positive for seasonal H1. FLULUM tested on 806 specimens submitted in the 2011-2012 season showed a sensitivity of 100% for FluA M, 89.9% for 2009 H1, 96.4% for H3, and 95.6% for FluB M. No cross-reactivity was observed for other respiratory viruses. Analytical sensitivity was assessed by testing dilutions of specimens with high viral loads. The limits of detection of FLULUM were comparable to those of the real-time PCR assay for FluA M, FluB M, and H3. The limits of detection for seasonal H1 and 2009 H1 were 10-fold higher for the FLULUM assay compared to real-time PCR. The FLULUM is an economic assay with high clinical sensitivity and specificity. It is particularly suited to high-volume detection of influenza viruses.
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Molecular epidemiology of respiratory syncytial virus transmission in childcare.
J. Clin. Virol.
PUBLISHED: 01-10-2013
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Respiratory syncytial virus (RSV) is the most important cause of serious respiratory infections in young children. No prior studies using molecular techniques to examine RSV transmission in the community childcare setting have been performed.
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Epidemiology of multiple respiratory viruses in childcare attendees.
J. Infect. Dis.
PUBLISHED: 01-03-2013
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The identification of multiple viruses during respiratory illness is increasing with advances in rapid molecular testing; however, the epidemiology of respiratory viral coinfections is not well known.
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Detection of adeno-associated virus viremia in hematopoietic cell transplant recipients.
J. Infect. Dis.
PUBLISHED: 10-17-2011
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Adeno-associated virus (AAV) is widely considered to be nonpathogenic, but the clinical epidemiology of this virus is limited. By use of polymerase chain reaction assays, we investigated the incidence and clinical significance of AAV viremia in a population of consecutive recipients of a hematopoietic cell transplant (HCT). Four (2.8%) of 145 patients developed AAV viremia after HCT. Viremia was low level and transient in all patients. Two patients were admitted to the hospital and died in proximity to AAV viremia (<7 weeks between diagnosis and death); however, AAV was not detected in tissue specimens obtained at autopsy. Thus, AAV does not appear to play a pathogenic role in organ-specific illness, even in a highly immunocompromised population.
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Respiratory viruses in children with cystic fibrosis: viral detection and clinical findings.
Influenza Other Respir Viruses
PUBLISHED: 09-29-2011
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Viral detection from different respiratory sample types in children with cystic fibrosis (CF) is facilitated by available molecular methods, but optimum sampling strategies have not been identified. In addition, associations between viral detection and respiratory symptoms are not well described.
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Multiple versus single virus respiratory infections: viral load and clinical disease severity in hospitalized children.
Influenza Other Respir Viruses
PUBLISHED: 05-31-2011
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Molecular testing for viral pathogens has resulted in increasing detection of multiple viruses in respiratory secretions of ill children. The clinical impact of multiple virus infections on clinical presentation and outcome is unclear.
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Emergence of oseltamivir-resistant pandemic H1N1 in an immunocompetent child with severe status asthmaticus.
J Asthma
PUBLISHED: 05-23-2011
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Pandemic influenza A (H1N1) may cause severe illness in pediatric patient with chronic lung disease.
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H275Y mutant pandemic (H1N1) 2009 virus in immunocompromised patients.
Emerging Infect. Dis.
PUBLISHED: 04-08-2011
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Most oseltamivir-resistant pandemic (H1N1) 2009 viruses have been isolated from immunocompromised patients. To describe the clinical features, treatment, outcomes, and virologic data associated with infection from pandemic (H1N1) 2009 virus with H275Y mutation in immunocompromised patients, we retrospectively identified 49 hematology-oncology patients infected with pandemic (H1N1) 2009 virus. Samples from 33 of those patients were tested for H275Y genotype by allele-specific real-time PCR. Of the 8 patients in whom H275Y mutations was identified, 1 had severe pneumonia; 3 had mild pneumonia with prolonged virus shedding; and 4 had upper respiratory tract infection, of whom 3 had prolonged virus shedding. All patients had received oseltamivir before the H275Y mutation was detected; 1 had received antiviral prophylaxis. Three patients excreted resistant virus for >60 days. Emergence of oseltamivir resistance is frequent in immunocompromised patients infected with pandemic (H1N1) 2009 virus and can be associated with a wide range of clinical disease and viral kinetics.
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Shedding of pandemic (H1N1) 2009 virus among health care personnel, Seattle, Washington, USA.
Emerging Infect. Dis.
PUBLISHED: 04-08-2011
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The Centers for Disease Control and Prevention (CDC) recommends that health care personnel (HCP) infected with pandemic influenza (H1N1) 2009 virus not work until 24 hours after fever subsides without the use of antipyretics. During an influenza outbreak, we examined the association between viral shedding and fever among infected HCP. Participants recorded temperatures daily and provided nasal wash specimens for 2 weeks after symptom onset. Specimens were tested by using PCR and culture. When they met CDC criteria for returning to work, 12 of 16 HCP (75%) (95% confidence interval 48%-93%) had virus detected by PCR, and 9 (56%) (95% confidence interval 30%-80%) had virus detected by culture. Fever was not associated with shedding duration (p = 0.65). HCP might shed virus even when meeting CDC exclusion guidelines. Further research is needed to clarify the association between viral shedding, symptoms, and infectiousness.
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Emerging oseltamivir resistance in seasonal and pandemic influenza A/H1N1.
J. Clin. Virol.
PUBLISHED: 03-22-2011
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The emergence of oseltamivir resistance in seasonal and pandemic influenza A/H1N1 has created challenges for diagnosis and clinical management. This review discusses how clinical virology laboratories have handled diagnosis of oseltamivir-resistant H1N1 and what we have learned from clinical studies and case series. Immunocompetent patients infected with oseltamivir-resistant H1N1 have similar outcomes as patients infected with oseltamivir-susceptible H1N1. However, immunocompromised patients infected with oseltamivir-resistant H1N1 experience potentially more risks of complication and transmissibility with few therapeutic options.
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A prospective study of parainfluenza virus type 4 infections in children attending daycare.
Pediatr. Infect. Dis. J.
PUBLISHED: 02-15-2011
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Studies of parainfluenza virus type 4 (PIV-4) have been limited by difficulty in culturing. We prospectively studied a cohort of 225 young children attending daycare followed for 165 child-years, using polymerase chain reaction to detect 12 viruses, including PIV-4. PIV-4 was second only to PIV-3, occurring in 9 of 87 (10%) PIV+ illnesses. PIV-4 illnesses were not more severe and not associated with a specific clinical syndrome.
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Introduction of a novel parechovirus RT-PCR clinical test in a regional medical center.
J. Clin. Virol.
PUBLISHED: 02-07-2011
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Although data documenting the severity and frequency of human parechovirus (HPeV) infections have been published, detection of HPeV is not routinely performed in most clinical virology laboratories.
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Outpatient upper respiratory tract viral infections in children with malaria symptoms in Western Kenya.
Am. J. Trop. Med. Hyg.
PUBLISHED: 11-02-2010
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A cross-sectional study was performed in children 5 through 10 years of age presenting to outpatient clinics in Nyanza Province, Kenya, in which nasal swab and blood specimens were collected during the high malaria transmission season. Patients presenting with malaria-like symptoms within 4 days of fever onset were enrolled in the study. Plasmodium parasitemia was determined by blood smear microscopy. Nasal swabs were screened for a panel of respiratory viruses by polymerase chain reaction. Influenza A, rhinoviruses, and other respiratory viruses were detected in 18%, 26%, and 12% of 197 specimens, respectively. Four of 36 patients with influenza A had a positive malaria blood slide, compared with 20 of 52 patients with rhinovirus. A significant burden of disease caused by influenza A in febrile children during the study period was observed, highlighting the need for further research into the burden of influenza disease in regions where malaria is holoendemic.
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Molecular detection of rhinoviruses.
Expert Rev. Mol. Diagn.
PUBLISHED: 05-15-2010
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Rhinoviruses are a frequent cause of the common cold and more serious illnesses, such as otitis media, sinusitis and asthma exacerbations and, more infrequently, have been implicated in lower airway infections, including pneumonia and exacerbations of chronic obstructive pulmonary disease and cystic fibrosis. Diagnosis of rhinovirus infections is best achieved using sensitive and specific reverse-transcription (RT)-PCR assays. The article by Do et al. describes the development and validation of a new one-step, TaqMan RT-PCR assay targeting the rhinovirus noncoding region for the detection of rhinovirus RNA in respiratory specimens from children with upper respiratory symptoms. The assay was rapid and specific and sensitive compared with cell culture. The recent use of RT-PCR assays for the detection of rhinoviruses in clinical specimens has provided more accurate information about the disease burden and epidemiology of these ubiquitous viruses.
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Frequent and prolonged shedding of bocavirus in young children attending daycare.
J. Infect. Dis.
PUBLISHED: 04-27-2010
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Little is known about human bocavirus (HBoV) persistence and shedding and the association between HBoV detection and the onset and resolution of respiratory symptoms.
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Respiratory virus pneumonia after hematopoietic cell transplantation (HCT): associations between viral load in bronchoalveolar lavage samples, viral RNA detection in serum samples, and clinical outcomes of HCT.
J. Infect. Dis.
PUBLISHED: 03-31-2010
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Few data exist on respiratory virus quantitation in lower respiratory samples and detection in serum from hematopoietic cell transplant (HCT) recipients with respiratory virus-associated pneumonia.
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Epidemiology of viral respiratory tract infections in a prospective cohort of infants and toddlers attending daycare.
J. Clin. Virol.
PUBLISHED: 01-13-2010
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The epidemiology of respiratory tract infections (RTIs) in a daycare cohort has not been explored using molecular techniques.
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Human rhinovirus and coronavirus detection among allogeneic hematopoietic stem cell transplantation recipients.
Blood
PUBLISHED: 12-30-2009
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Little is known about clinical and virologic manifestations of rhinovirus (HRV) and coronavirus (HCoV) infections after hematopoietic cell transplantation (HCT). We performed surveillance for 1 year and describe the natural history of these infections during the first 100 days after allogeneic HCT, when symptom surveys and upper respiratory samples were collected weekly. Samples were tested using RT-PCR for HRVs and HCoVs (OC43, 229E, HKU1, and NL63). Among 215 patients, 64 (30%) patients had 67 infections. Day 100 cumulative incidence estimate was 22.3% for HRV and 11.1% for HCoV. Median duration of viral shedding was 3 weeks; prolonged shedding of at least 3 months occurred in 6 of 45 patients with HRV and 3 of 22 with HCoV. Six patients with HRV and 9 with HCoV were asymptomatic. HRV infection was associated with rhinorrhea, congestion, postnasal drip, sputum, and cough; HCoV infection was not associated with respiratory symptoms or hepatic dysfunction. Lower respiratory infection developed in 2 patients with HRV before day 100, and 1 each with HRV and HCoV after day 100. HRV and HCoV infections are common in the first 100 days after HCT, viral shedding lasts more than 3 weeks in half, and lower respiratory infection is rare.
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Detection of bocavirus in saliva of children with and without respiratory illness.
J. Clin. Microbiol.
PUBLISHED: 09-30-2009
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We evaluated saliva samples from 149 children 2 to 11 years old for human bocavirus (hBoV) DNA. hBoV was detected in saliva samples at asymptomatic enrollment in 3% (5/149) and during respiratory illness in 2% (2/106) of the cases. hBoV was detected in only 1/149 asymptomatic and 0/106 illness nasal samples.
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Multiple viral respiratory pathogens in children with bronchiolitis.
Acta Paediatr.
PUBLISHED: 04-03-2009
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The aim of the study was to describe the frequency of viral pathogens and relative frequency of co-infections in nasal specimens obtained from young children with bronchiolitis receiving care at a childrens hospital.
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Self-collection of foam nasal swabs for respiratory virus detection by PCR among immunocompetent subjects and hematopoietic cell transplant recipients.
J. Clin. Microbiol.
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Self-collected foam nasal swabs (NS) obtained after instillation of saline spray were compared with nasal washes from immunocompetent subjects during 146 upper respiratory infections (URIs); the sensitivities for reverse transcription (RT)-PCR respiratory virus detection were 95% and 88%, respectively (P = 0.06). The sensitivities from NS collected with and without saline spray during 142 URIs from immunocompetent subjects were 96% and 86% (P = 0.004), respectively, and those from 140 URI samples from hematopoietic cell transplantation recipients were 88% and 85% (P = 0.56), respectively.
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Influenza viral RNA detection in blood as a marker to predict disease severity in hematopoietic cell transplant recipients.
J. Infect. Dis.
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Influenza RNA in blood (viremia) was detected in 9 of 79 (11.4%) hematopoietic cell transplant recipients with influenza, and was less frequently observed in patients with upper respiratory tract disease only and more frequently in patients infected with 2009 pandemic influenza A/H1N1 strain (versus seasonal strains). Viremia increased the risk of progression to lower respiratory tract disease (LRD), hypoxemia, respiratory failure, and overall and influenza-related death. Among patients with LRD, viremia was associated with increased hazards of overall and influenza-associated death (hazard ratio 3.5, 1.1-12). Thus, influenza viremia may serve as marker for overall poor outcome.
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WU and KI polyomaviruses in respiratory samples from allogeneic hematopoietic cell transplant recipients.
Emerging Infect. Dis.
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Data are limited regarding 2 new human polyomaviruses, KI polyomavirus (KIPyV) and WU polyomavirus (WUPyV), in immunocompromised patients. We used real-time PCR to test for these and 12 respiratory viruses in 2,732 nasal wash samples collected during the first year after allogeneic hematopoietic cell transplantation from 222 patients. Specimens were collected weekly until day 100; then at least every 3 months. One year after hematopoietic cell transplantation, the cumulative incidence estimate was 26% for KIPyV and 8% for WUPyV. Age <20 years predicted detection of KIPyV (hazard ratio [HR] 4.6) and WUPyV (HR 4.4), and detection of a respiratory virus in the previous 2 weeks predicted KIPyV detection (HR 3.4). Sputum production and wheezing were associated with detection of KIPyV in the past week and WUPyV in the past month. There were no associations with polyomavirus detection and acute graft versus host disease, cytomegalovirus reactivation, neutropenia, lymphopenia, hospitalization, or death.
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Comparison of FilmArray Respiratory Panel and laboratory-developed real-time reverse transcription-polymerase chain reaction assays for respiratory virus detection.
Diagn. Microbiol. Infect. Dis.
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The FilmArray Respiratory Panel (Idaho Technology) is a highly multiplexed respiratory virus real-time polymerase chain reaction (PCR) assay. Eighty-four respiratory viruses identified by laboratory-developed real-time reverse transcription-PCR assays (LDA) or by viral cultures were mixed and tested by FilmArray to assess its performance. FilmArray identified 72 (90%) of 80 viruses also detected by LDA. Six of the 8 viruses not detected by FilmArray had PCR cycle threshold values >35. Compared to LDA, FilmArray showed comparable sensitivity when used to test serial dilutions of virus mixtures and good agreement with negative samples. With the use of 1 FilmArray instrument, 7 clinical samples could be analyzed and reported in an 8-h shift compared to 20 using LDA and 1 real-time detection instrument. While the FilmArray was rapid and easy to use, its low throughput and qualitative results may be a disadvantage in some clinical settings.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.