Birgit Winther

Gene Expression Profiles during In Vivo Human Rhinovirus Infection Insights into the Host Response

RATIONALE Human rhinovirus infections cause colds and trigger exacerbations of lower airway diseases. OBJECTIVES To define changes in gene expression profiles during in vivo rhinovirus infections. METHODS Nasal epithelial scrapings were obtained before and during experimental rhinovirus infection, and gene expression was evaluated by microarray. Naturally acquired rhinovirus infections, cultured human epithelial cells, and short interfering RNA knockdown were used to further evaluate the role of viperin in rhinovirus infections. MEASUREMENTS AND MAIN RESULTS Symptom scores and viral titers were measured in subjects inoculated with rhinovirus or sham control, and changes in gene expression were assessed 8 and 48 hours after inoculation. Real-time reverse transcription-polymerase chain reaction for viperin and rhinoviruses was used in naturally acquired infections, and viperin mRNA levels and viral titers were measured in cultured cells. Rhinovirus-induced changes in gene expression were not observed 8 hours after viral infection, but 11,887 gene transcripts were significantly altered in scrapings obtained 2 days postinoculation. Major groups of up-regulated genes included chemokines, signaling molecules, interferon-responsive genes, and antivirals. Viperin expression was further examined and also was increased in naturally acquired rhinovirus infections, as well as in cultured human epithelial cells infected with intact, but not replication-deficient, rhinovirus. Knockdown of viperin with short interfering RNA increased rhinovirus replication in infected epithelial cells. CONCLUSIONS Rhinovirus infection significantly alters the expression of many genes associated with the immune response, including chemokines and antivirals. The data obtained provide insights into the host response to rhinovirus infection and identify potential novel targets for further evaluation.

Histopathologic Examination and Enumeration of Polymorphonuclear Leukocytes in the Nasal Mucosa during Experimental Rhinovirus Colds

The histology of the nasal mucosa was examined by serial scrape and punch biopsies in 20 rhinovirus infected volunteers and 10 sham inoculated controls. No morphologic changes could be detected in the epithelial or subepithelial portions in the mucosa of specimens from infected volunteers. There was a significant increase in the number of polymorphonuclear leukocytes (PMNs) in the nasal epithelium of the infected subjects early in the course of the cold compared to their pre-infection baseline. However, trauma to the nasal mucosa from repeated sampling led to an outpouring of PMNs into nasal mucus, making evaluation of the results difficult. The number of mast cells seen in the mucosal specimens of the infected subjects did not differ from that seen in controls.

Rhinosinusitis: Developing guidance for clinical trials

The Rhinosinusitis Initiative was developed by 5 national societies. The current guidance document is an expansion of the 2004 publication, “Rhinosinusitis: Establishing definitions for clinical research and patient care” and provides templates for clinical trials in antimicrobial, anti-inflammatory, and symptom-relieving therapies for the following: (1) acute presumed bacterial rhinosinusitis, (2) chronic rhinosinusitis (CRS) without nasal polyps, (3) CRS with nasal polyps, and (4) classic allergic fungal rhinosinusitis. In addition to the templates for clinical trials and proposed study designs, the Rhinosinusitis Initiative has developed 6 appendices, which address (1) health outcomes, (2) nasal endoscopy and staging of CRS, (3) radiologic imaging, (4) microbiology, (5) laboratory measures, and (6) biostatistical methods.

Temporal Relationships Between Colds, Upper Respiratory Viruses Detected by Polymerase Chain Reaction, and Otitis Media in Young Children Followed Through a Typical Cold Season

INTRODUCTION. Otitis media is a frequent complication of a viral upper respiratory tract infection, and the reported co-incidence of those diseases increases with assay sensitivity and sampling density. We determined the incidence of otitis-media complications in young children when referenced to cold-like illnesses and to concurrent virus recovery from the nasopharynx. METHODS. A total of 60 children from 24 families were followed from October 2003 through April 30, 2004, by daily parental recording of illness signs, weekly pneumatic otoscopic examinations, and periodic polymerase chain reaction assay of collected nasal fluids for common viruses. RESULTS. One hundred ninety-nine cold-like illnesses were observed, but a sample for virus assay was not collected concurrent with 71 episodes. Of the remainder, 73% of cold-like illnesses were temporally related to recovery of 1 or a combination of the assayed viruses, with rhinovirus predominating. For non–cold-like illness periods, 54 (18%) of 297 assays were positive for virus, and the virus frequency distribution was similar to that for cold-like illnesses. There were 93 diagnosed otitis-media episodes; 65 (70%) of these occurred during a cold-like illness. For the 79 otitis-media episodes with available nasal samples, 61 (77%) were associated with a positive virus result. In this population, the otitis-media complication rate for a cold-like illness was 33%. CONCLUSIONS. A cold-like illness was not a prerequisite for polymerase chain reaction detection of viruses in the nose and nasopharynx of young children. Viral detection by polymerase chain reaction in the absence of a cold-like illness is associated with complications in some subjects. Otitis media is a complication of viral infection both with and without concurrent cold-like illnesses, thus downwardly biasing coincidence estimates that use cold-based illnesses as the denominator.

Symptom Profile of Common Colds in School-Aged Children

Background: Signs and symptoms of a common cold reported in young children are those perceived by caretakers. Objective signs include cough, fever, and sneezing. Subjective symptoms include nasal congestion, feverishness, headache, and sore throat. School-aged children may provide a more accurate picture of the symptom profile during colds because they can self-report. Methods: Using preprinted diary sheets listing common signs and symptoms, diaries were kept for school-aged children for 10 days after onset of a cold. Nasopharyngeal aspirates were analyzed for respiratory viruses and potential bacterial pathogens. Results: Out of 81 colds studied, the most common signs were cough and sneezing, although the most common symptoms were nasal congestion and runny nose. Other symptoms, including feverishness and headache, were each reported in 15% of children at onset. The majority of children (73%) continued to be symptomatic 10 days after onset. Rhinovirus was detected in 46% and 1 or more potential bacterial pathogens (Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis) in 29% of episodes. Symptom profiles for rhinovirus illnesses and those in which potential pathogenic bacteria were detected were not different from the rest. Conclusion: The common cold in school-aged children is characterized by nasal congestion, cough, and runny nose. Signs and symptoms usually continue for at least 10 days.

Viral-induced rhinitis.

Upper respiratory viruses cause self-limited illness characterized by acute rhinitis. In rhinovirus colds the symptoms are thought to be caused by the host response rather than viral damage of the nasal epithelium. Rhinovirus triggers an inflammatory cascade, evidenced by the presence of inflammatory mediators (e.g., IL-8) and proinflammatory cytokines (e.g., kinins) in nasal secretions, which results in symptomatic illness. In contrast to rhinovirus and coronavirus, which do not cause discernible epithelial damage, influenza virus and adenovirus do damage the nasal epithelium. Appropriate antiviral therapy will depend on the causative virus. Treatment of rhinovirus colds may require an antiviral agent (e.g., interferon a) in combination with antiinflammatory medication.

Combined Antiviral-Antimediator Treatment for the Common Cold

Abstract A randomized, controlled, double-masked clinical trial was conducted with a combination antiviral-antimediator treatment for experimental rhinovirus colds. In all, 150 healthy men and women (aged 18–51 years) were randomly assigned to 1 of 3 groups: intranasal interferon (IFN)–α2b (6×106 U every 12 h × 3) plus oral chlorpheniramine (12 mg extended release) and ibuprofen (400 mg) every 12 h for 4.5 days (n=59 subjects); intranasal placebo plus oral chlorpheniramine and ibuprofen (n=61 subjects); or intranasal and oral placebos (n=30 subjects). Treatment was started 24 h after intranasal viral challenge. During the 4.5 days of treatment with IFN-α2b, chlorpheniramine, and ibuprofen, the daily mean total symptom score was reduced by 33%–73%, compared with placebo. Treatment reduced the severity of rhinorrhea, sneezing, nasal obstruction, sore throat, cough, and headache and reduced nasal mucus production, nasal tissue use, and virus concentrations in nasal secretions. IFN-α2b added to the effectiveness of chlorpheniramine and ibuprofen and was well tolerated

Weekly point prevalence of Streptococcus pneumoniae, Hemophilus influenzae and Moraxella catarrhalis in the upper airways of normal young children: effect of respiratory illness and season

The aim was to determine the effect of respiratory illness and season on carriage rates in the upper airways of Streptococcus pneumoniae, Hemophilus influenzae and Moraxella catarrhalis in normal children. Sixteen healthy children, 1–10 years old, amenable to weekly sampling were followed longitudinally for at least three seasons of the year. Respiratory symptoms were recorded daily; weekly nasal aspirate/wash samples were cultured on selective agars. Urea concentration in samples was used to define dilution of secretion. 68% of 950 samples were culture positive; 44% of positives had two or all three species. Each species was detected in about one third of samples. Bacteria were detected in 76% of samples during illness vs. 65% during wellness (p=0.004). Seasonal carriage rates varied from 56% in summer and fall to 85% in winter. There was a strong inverse correlation between dilution of secretion and bacterial detection rate in illness and wellness aspirate samples during the four seasons (r=−0.82, p=0.01). Detection of bacteria varied with the amount of secretion in the sample. This variation accounts for the apparent differences in bacterial carriage during illness vs. wellness and during different seasons.

Rate of concurrent otitis media in upper respiratory tract infections with specific viruses.

OBJECTIVE To estimate the coincidence of new otitis media (OM) for first nasopharyngeal detections of the more common viruses by polymerase chain reaction (PCR). New OM episodes are usually coincident with a viral upper respiratory tract infection (vURTI), but there are conflicting data regarding the association between specific viruses and OM. DESIGN Longitudinal (October-March), prospective follow-up of children for coldlike illness (CLI) by diary, middle ear status by pneumatic otoscopy, and vURTI by PCR. SETTING Academic medical centers. PARTICIPANTS A total of 102 families with at least 2 children aged between 1 and 5 years (213 children; mean [SD] age, 3.7 [1.5] years; 110 male; and 176 white) were recruited from the local communities at 2 study sites by advertisement. MAIN OUTCOME MEASURES New OM and CLI episodes and nasopharyngeal virus detections. RESULTS A total of 176 children (81%) had isolated PCR detection of at least 1 virus. The OM coincidence rates were 62 of 144 (44%) for rhinovirus, 15 of 27 (56%) for respiratory syncytial virus, 8 of 11 (73%) and 1 of 5 (20%) for influenza A and B, respectively, 6 of 12 (50%) for adenovirus, 7 of 18 (39%) for coronavirus, and 4 of 11 (36%) for parainfluenza virus detections (P = .37). For rhinovirus, new OM occurred in 50% of children with and 32% without a concurrent CLI (P = .15), and OM risk was predicted by OM and breastfeeding histories and by daily environment outside the home. CONCLUSIONS New OM was associated with nasopharyngeal detection of all assayed viruses irrespective of the presence or absence of a concurrent CLI. Differences among viruses were noted, but statistical significance was not achieved, possibly because of the low power associated with the small number of nonrhinovirus detections.

Cytokine polymorphisms predict the frequency of otitis media as a complication of rhinovirus and RSV infections in children.

Previous studies suggested that the otitis media (OM) complication rate of viral upper respiratory infection (vURI) is conditioned by genes affecting cytokine production. Two hundred and thirty children (114 male; 187 White, 25 Black; aged 1–9.3 years, average = 3.6 ± 1.6 years) were prospectively followed over the typical cold season for cold-like illness and OM. Nasopharyngeal secretion samples collected during cold-like illness and OM were assayed for upper respiratory viruses and buccal samples were assayed for TNFα (−308), IL-10(−1082, −819, −592), IL-6 (−174) and IFN-γ (+874) polymorphisms. Logistic regression was used to identify genotypes that predict OM coincident with RSV and rhinovirus (RV) infection. Of the 157 children with RV detection (79 male; 132 White, 13 Black, 12 Other; aged 3.6 ± 1.5 years), simple logistic regression identified age (B = −0.34, Z = −2.8, P < 0.01, OR = 0.71), IL-6 (B = −0.76, Z = −3.3, P < 0.01, OR = 0.47) and IL-10 (B = 0.49, Z = 2.0, P = 0.05, OR = 1.6) as significant predictors of OM coincidence. A more complex logistic regression model for RV detection that included selected OM risk factors identified these factors as well as the TNFα genotype, OM history, breastfeeding history and daily environment as significant predictors of OM coincidence. Of the 43 children with RSV detection (21 male; 35 White, 5 Black, 3 Other, aged 3.9 ± 1.7 years), logistic regression identified IL-10 (B = 1.05, Z = 2.0, P = 0.05, OR = 2.9) as a significant predictor of OM coincidence. New OM episodes coincident with evidence of RSV and RV infection were significantly more frequent in children with high production IL-10 phenotypes. The low production IL-6 and high production TNFα phenotypes also contributed to OM risk during RV detection. Cytokine polymorphisms may be one of an expectedly large number of genetic factors contributing to the known heritability of OM.