Human rhinovirus in experimental infection after peroral Lactobacillus rhamnosus GG consumption, a pilot study.

BACKGROUND: Data has emerged on possible beneficial effects of probiotics in respiratory tract viral infections, but it is unclear if the promising positive effects evidenced are due to a reduced viral load during infections. The aims of this work were to investigate the effect of peroral probiotic Lactobacillus rhamnosus GG (American Type Culture Collection [ATCC], Accession No. 53103) consumption on human rhinovirus (HRV) load in nasopharyngeal lavage samples in experimental HRV infection, and to correlate viral load to clinical symptoms. METHODS: Intranasal HRV A39 inoculation was performed on 59 adults, who had consumed juice enriched with live or heat-inactivated L. rhamnosus GG or control juice for 3 weeks prior to inoculation in a randomized, controlled, pilot trial setting. Nasopharyngeal lavage samples and symptom data were analyzed on day 0 before inoculation, and on days 2 and 5. Samples were subjected to quantitative HRV detection by polymerase chain reaction (PCR). RESULTS: Before inoculation 9 of 59 (15%) samples presented with another HRV strain than the studied A39. There was a tendency toward the lowest HRV loads in the L. rhamnosus GG groups and the highest in placebo group (log10 copies/mL, 95% confidence interval [CI], 6.20 [5.18 to 7.40] in live, 6.30 [4.91 to 7.08] in inactivated L. rhamnosus GG, and 7.25 [5.81 to 7.52] in placebo group, p = 0.57 in day 2) in the wild-type excluded population. The HRV load positively correlated with the symptom scores on days 2 and 5 (correlation coefficient 0.61 [p < 0.001] and 0.28 [p = 0.034], respectively). CONCLUSION: Results did not show statistical differences in viral loads in subjects using L. rhamnosus GG when compared to placebo. HRV load positively correlated with the total symptom scores.

Characterization of the nasopharyngeal microbiota in health and during rhinovirus challenge.

BACKGROUND: The bacterial communities of the nasopharynx play an important role in upper respiratory tract infections (URTIs). Our study represents the first survey of the nasopharynx during a known, controlled viral challenge. We aimed to gain a better understanding of the composition and dynamics of the nasopharyngeal microbiome during viral infection. METHODS: Rhinovirus illnesses were induced by self-inoculation using the finger to nose or eye natural transmission route in ten otherwise healthy young adults. Nasal lavage fluid samples (NLF) samples were collected at specific time points before, during, and following experimental rhinovirus inoculation. Bacterial DNA from each sample (N = 97 from 10 subjects) was subjected to 16S rRNA sequencing by amplifying the V1-V2 hypervariable region followed by sequencing using the 454-FLX platform. RESULTS: This survey of the nasopharyngeal microbiota revealed a highly complex microbial ecosystem. Taxonomic composition varied widely between subjects and between time points of the same subject. We also observed significantly higher diversity in not infected individuals compared to infected individuals. Two genera - Neisseria and Propionibacterium - differed significantly between infected and not infected individuals. Certain phyla, including Firmicutes, Actinobacteria, and Proteobacteria, were detected in all samples. CONCLUSIONS: Our results reveal the complex and diverse nature of the nasopharyngeal microbiota in both healthy and viral-challenged adults. Although some phyla were common to all samples, differences in levels of diversity and selected phyla were detected between infected and uninfected participants. Deeper, species-level metagenomic sequencing in a larger sample is warranted.

Bacteria in the nose of young adults during wellness and rhinovirus colds: detection by culture and microarray methods in 100 nasal lavage specimens.

BACKGROUND: Patients  with  viral respiratory infections/viral rhinitis/common colds are often treated with antibiotic; however, there is little information on whether or how bacterial microbiota in the nose and nasopharynx might influence the course of viral illnesses. METHODS: To initiate investigation of possible interaction between viral respiratory illness and microbiota of the nose/nasopharynx, we used microarray technology to examine 100 nasal lavage fluid (NLF) samples for bacterial species and recorded the bacterial titer of culturable bacteria. Rhinovirus illnesses were induced by self-inoculation using the "finger to nose or eye natural transmission route" in 10 otherwise healthy young adults. NLF samples were collected during wellness and at specific time points following experimental rhinovirus inoculation. RESULTS: The rhinovirus infection rate was 70%. There were no consistent changes in the prevalence of different bacterial species determined by microarray and bacterial titer by culture methods during rhinovirus infection. The bacterial profile in NLF samples showed high variability between volunteers but low variability in multiple NLFs obtained before and following infection from the same volunteer. Streptococcus epidermidis/coagulase-negative staphylococcus (CNS) were identified in all 10 subjects. One or more bacterial sinus/otitis pathogens were identified by microarray in 6 of the 10 volunteers. The microarray identified a few bacteria not included in traditional bacterial cultures. CONCLUSION: Our pilot study showed that each of the 10 volunteers had a unique bacterial profile in the nose by microarray analysis and that bacterial load did not change during experimental rhinovirus colds. Larger scale studies are warranted.

16S rRNA survey revealed complex bacterial communities and evidence of bacterial interference on human adenoids.

Adenoid microbiota plays an important role in the development of various infectious and non-infectious diseases of the upper airways, such as otitis media, adenotonsillitis, rhinosinusitis and adenoid hypertrophy. Studies have suggested that adenoids could act as a potential reservoir of opportunistic pathogens. However, previous bacterial surveys of adenoids were mainly culture based and therefore might only provide an incomplete and potentially biased assessment of the microbial diversity. To develop an in-depth and comprehensive understanding of the adenoid microbial communities and test the 'pathogen reservoir hypothesis', we carried out a 16S rRNA based, culture-independent survey of bacterial communities on 67 human adenoids removed by surgery. Our survey revealed highly diverse adenoid bacterial communities distinct from those of other body habitats. Despite large interpersonal variations, adenoid microbiota shared a core set of taxa and can be classified into at least five major types based on its bacterial species composition. Our results support the 'pathogen reservoir hypothesis' as we found common pathogens of otitis media to be both prevalent and abundant. Co-occurrence analyses revealed evidence consistent with the bacterial interference theory in that multiple common pathogens showed 'non-coexistence' relationships with non-pathogenic members of the commensal microflora.

Occurrence of acute otitis media during colds in children younger than four years.

To determine how frequently acute otitis media (AOM) occurs, we enrolled children between 6 months and 3 years of age who returned several weeks before and 6 to 10 times during a cold for tympanometry and photography of the tympanic membrane. American Academy of Pediatrics (AAP) criteria were used to diagnose AOM. Children visited their physicians at their discretion. AOM occurred in 17 (55%) of 31 colds; in 12 (100%) colds with pre-existing middle ear effusion (MEE); and in 5 (26%) of 19 colds with no pre-existing MEE (P < 0.0001). Four patients received antibiotics from their physicians. Of 17 children with AOM, 12 did not seek care. AOM is common during colds, particularly with pre-existing MEE.

Rhinovirus infections in the upper airway.

The majority of cold and flulike illnesses are caused by human rhinoviruses (HRVs). Improved detection of HRV has shown that HRVs are also associated with more serious illness, such as exacerbation of asthma, wheezing illnesses in children, chronic obstructive pulmonary disease, cardiopulmonary disease, and fatal pneumonia in immune-compromised patients. HRV is a major cause of acute viral respiratory tract infections in hospitalized children and is among the leading causes of childhood mortality worldwide. Detection of the HRV genome by reverse transcriptase-polymerase chain reaction and genomic sequencing has brought to light a new clade, HRV-C, to the already recognized HRV-A and HRV-B clades. The clinical complications related to all rhinovirus infections include acute otitis media, acute sinusitis, and acute bronchitis. The enormous public health implications from those diseases far overshadow those of the common cold. This article provides an overview of the pathogenesis of rhinovirus infection in the upper airways. Most research has been done in young healthy adults with self-limiting experimental and natural rhinovirus infections, and this may set the stage for understanding rhinovirus infections in the ear, sinus, and lower airways.

Bacterial pathogens of acute sinusitis in the osteomeatal complex during common colds and wellness.

BACKGROUND: Pathogenic bacteria have been cultured from the osteomeatal complex (OMC) in one-third of adults with apparent acute bacterial sinusitis; however, it is not known whether bacteria are present in the OMC during uncomplicated viral colds in adults. METHODS: Adult volunteers were recruited for a study during wellness and at the time of acute common cold. Swab cultures were obtained from the OMC and from the nasopharynx by 2 routes (through the nose and through the mouth). Swab eluates were inoculated on selective agars to detect S. pneumoniae, H. influenzae, and M. catarrhalis. RESULTS: Bacterial pathogens were detected in the OMC more frequently during common colds than during wellness (31% vs 8%, p < 0.008). Pathogens detected in the OMC were always present in the nasopharynx of the subject. CONCLUSION: Bacterial pathogens are present in the OMC in a subgroup of adult patients with uncomplicated upper respiratory illness/common cold. The nasopharynx appears to be the reservoir for bacterial pathogens in the OMC.

Rhinovirus genome evolution during experimental human infection.

Human rhinoviruses (HRVs) evolve rapidly due in part to their error-prone RNA polymerase. Knowledge of the diversity of HRV populations emerging during the course of a natural infection is essential and represents a basis for the design of future potential vaccines and antiviral drugs. To evaluate HRV evolution in humans, nasal wash samples were collected daily for five days from 15 immunocompetent volunteers experimentally infected with a reference stock of HRV-39. In parallel, HeLa-OH cells were inoculated to compare HRV evolution in vitro. Nasal wash in vivo assessed by real-time PCR showed a viral load that peaked at 48-72 h. Ultra-deep sequencing was used to compare the low-frequency mutation populations present in the HRV-39 inoculum in two human subjects and one HeLa-OH supernatant collected 5 days post-infection. The analysis revealed hypervariable mutation locations in VP2, VP3, VP1, 2C and 3C genes and conserved regions in VP4, 2A, 2B, 3A, 3B and 3D genes. These results were confirmed by classical sequencing of additional samples, both from inoculated volunteers and independent cell infections, and suggest that HRV inter-host transmission is not associated with a strong bottleneck effect. A specific analysis of the VP1 capsid gene of 15 human cases confirmed the high mutation incidence in this capsid region, but not in the antiviral drug-binding pocket. We could also estimate a mutation frequency in vivo of 3.4x10(-4) mutations/nucleotides and 3.1x10(-4) over the entire ORF and VP1 gene, respectively. In vivo, HRV generate new variants rapidly during the course of an acute infection due to mutations that accumulate in hot spot regions located at the capsid level, as well as in 2C and 3C genes.

Impact of oseltamivir treatment on the incidence and course of acute otitis media in children with influenza.

OBJECTIVE: Acute otitis media (AOM) is the most common complication of pediatric influenza, and imposes a substantial health care burden. We examined the influence of oseltamivir treatment on the incidence and course of AOM in children with influenza. METHODS: In the original study, 695 children 1-12 years who presented within 48h of the onset of influenza-like symptoms were randomized to oseltamivir (2mg/kg) or placebo given twice daily for 5 days. AOM was assessed at enrollment and days 3, 6 (+/-1), 10 (+/-2) and 28 (+/-7). AOM was clinically diagnosed by the participating primary care provider, supported by tympanometry when possible. We performed a retrospective analysis of those participants with laboratory-confirmed influenza (LCI). Assessments included the incidence and clinical course of new AOM cases. RESULTS: In all, 452 children had LCI; 217 received oseltamivir and 235 placebo. AOM was diagnosed on or after study day 3 at a significantly lower frequency in the oseltamivir versus placebo group (12.4% versus 21.7%; relative risk [RR]: 0.57 [95% CI: 0.37, 0.88], respectively). Treatment effects were greatest for children 1-2 years (RR=0.42 [95% CI: 0.20, 0.89]) and 3-5 years (RR=0.45 [95% CI: 0.19, 1.04]), in whom the incidence of AOM was highest. CONCLUSIONS: Oseltamivir treatment significantly reduces the emergence of new AOM infections in children with LCI; effects are most pronounced in those <5 years. CLINICAL TRIAL NUMBER: WV15758.

Nasal secretion concentrations of IL-5, IL-6, and IL-10 in children with and without upper respiratory tract viruses.

OBJECTIVE: To determine if levels of interleukin (IL) 5, IL-6, and IL-10 or their ratios in nasal secretion are diagnostic of viral upper respiratory tract infections (vURTIs) and coldlike illnesses (CLIs) in children. DESIGN: Longitudinal study of children for vURTIs, CLIs, and concentrations and ratios of nasal cytokines. SETTING: Outpatient assessments of children. PARTICIPANTS: A total of 224 children, aged 1 to 9 years. MAIN OUTCOME MEASURES: Concentrations of IL-5, IL-6, and IL-10 in nasal secretions, vURTIs diagnosed by polymerase chain reaction (PCR) detection of upper respiratory tract viruses, and concurrent CLIs diagnosed by parents. RESULTS: Of 1269 secretion samples, 552 (43.5%) were collected during a vURTI (PCR findings positive for an assayed virus [PCR(+)]). A concurrent CLI was diagnosed for 34% of the PCR(+) samples and for 18% of the samples found to be negative by PCR analysis (PCR(-)). Cytokine concentrations and ratios were highly variable and skewed to the lower values. The significance of the cytokine concentrations and ratios as discriminators of groups defined by the presence or absence of virus and of subgroups defined by the presence or absence of a CLI was evaluated using receiver operating characteristic curves. All measures were significant discriminators of the PCR(+) vs PCR(-) groups, and most were significant discriminators of the paired CLI subgroups. The concentration of IL-6 and the IL-5/IL-6 ratio were the best discriminators across all groups and subgroups. However, the sensitivities and specificities of those discriminators at the best cutoff values were on the order of 0.7 for the most extreme pairwise comparison (PCR(+)CLI(+) vs PCR(-)CLI(-)) and lower for the other comparison groups. CONCLUSION: The low sensitivities and specificities for cytokine-based assignment of specimens to the paired groups and subgroups limit their usefulness for diagnosis of infection or illness.

Decreased rhinovirus shedding after intranasal oxymetazoline application in adults with induced colds compared with intranasal saline.

BACKGROUND: Intranasal oxymetazoline (OMZ) is used as a decongestant during common colds. Recently, intracellular adhesion molecule (ICAM) 1 receptor expression in vitro has been shown to be diminished by OMZ. ICAM-1 is the major receptor used by rhinovirus to gain entry to human cells. The objective of this study was to assess the effect of OMZ on geometric mean titer of rhinovirus in nasal lavage fluid after rhinovirus inoculation. METHODS: Volunteers with antibody titers of ≤1:4 to rhinovirus type 39 were enrolled in a randomized, reference-controlled, double-blind study. Beginning 3 hours after intranasal challenge with 100-300 tissue culture infectious dose (TCID)50 of virus, subjects received active 0.05% OMZ (45 µL containing 22.5 µg of OMZ hydrochloride in citrate buffer) or reference control (physiological saline solution [PSS]) three times daily for 5 days. Rhinovirus was detected in fibroblast cultures. RESULTS: Geometric mean viral titer (log10) in 34 rhinovirus-infected subjects receiving OMZ was 1.49 on day 2 compared with 2.24 in the 38 infected subjects receiving PSS (p = 0.04). On day 3, the mean titers were 1.45 and 2.08, respectively. Median length of viral shedding was 3.3 days (OMZ) and 3.4 (PSS). Duration of clinical illness was 6.1 days in both groups. CONCLUSION: Topical OMZ decreased viral titer on day 2 during experimental rhinovirus infection in normal volunteers.

Acute cough: a diagnostic and therapeutic challenge.

BACKGROUND: Acute cough is one of the most common complaints prompting patient visits to healthcare professionals. Despite the broad repercussions of acute cough on patient quality of life, school and work productivity, and public health resources, research on this condition is minimal, as are the available treatment options. Many patients use over-the-counter medicines, which are often ineffective for symptom relief. Some therapies may achieve antitussive activity, but at the expense of unpleasant or intolerable side effects. UNMET NEEDS: When considering the treatments currently available for the management of acute cough, the multiple limitations of such treatments are quite apparent. Most of these treatments lack clinically proven efficacy and reliability to support their use. This reinforces the need for the generation of quality scientific data from well-performed clinical trials. Hopefully, the result will be the development of safer, more effective and more reliable therapeutic options in the management of acute cough. COUGH ASSESSMENT AND MANAGEMENT: Acute cough can be due to a variety of causes, and it is worthwhile to consider these pathogenic factors in some detail. It is also important to be familiar with the effects that acute cough has on patients' quality of life, work productivity, and the healthcare system; proper awareness of these effects may contribute to better understanding of the social impact of cough. In reference to the available treatments for the management of acute cough, adequate knowledge of the type of over-the-counter and prescription products in the market, as well as their mode of action and advantages/disadvantages, may provide expanded pharmacotherapeutic opportunities and facilitate better clinical decisions. However, due to the drawbacks of current treatment options, ideas for future cough management and newer products need to be considered and tested. CONCLUSION: In view of the socio-economic impact of acute cough and the limitations of available treatments, a renewed interest in the management of acute cough needs to be encouraged. The current strategies for acute cough management need to be reassessed, with a focus on developing new, reliable products and formulations with proven efficacy and safety.

Location of bacterial biofilm in the mucus overlying the adenoid by light microscopy.

OBJECTIVE: To determine the location of bacteria and biofilm in adenoid tissue and in mucus overlying the adenoid. DESIGN: Adenoids removed in 1 piece were oriented to the cephalic and caudal ends. Mucus was fixed by the gradual addition of Carnoy fluid. Consecutive histologic sections were stained with periodic acid-Schiff for visualization of the exopolysaccharide matrix, Giemsa for visualization of bacteria and cells, and fluorescent in situ hybridization with a universal probe for visualization of bacteria. SETTING: Department of Otolaryngology-Head and Neck Surgery, University of Virginia. PARTICIPANTS: We obtained adenoids from children 10 years or younger who had chronic adenotonsillitis or obstructive sleep apnea. Twenty-seven adenoids were used to develop the fixation method. We examined histologic sections from 9 of 10 adenoids fixed using the final fixation protocol. One adenoid that was missing the surface epithelium was excluded from further evaluation. MAIN OUTCOME MEASURE: Identification of bacteria by light microscopy. RESULTS: Bacteria in large numbers were present in the mucus overlying the surface of all 9 adenoids; bacteria were not found in the parenchyma of the adenoids below the epithelial surface. Bacterial biofilms were present on 8 of the 9 adenoids. Sessile (attached) biofilm was present on the caudal end of only 1 adenoid. Multiple planktonic (unattached) biofilms were present on 7 adenoids, always in areas not subject to mucus flow. Biofilms were most common on the caudal portions of adenoids. CONCLUSIONS: Bacteria of the adenoid reside in secretions on the surface and in crypts. Biofilms, predominantly planktonic, were present on 8 of 9 adenoids excised because of hypertrophy. Whether biofilms have a role in the causation of adenoid hypertrophy is not known.

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 TNFalpha (-308), IL-10(-1082, -819, -592), IL-6 (-174) and IFN-gamma (+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 TNFalpha 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 TNFalpha 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.

Daily tympanometry as a functional measure of middle ear status and Eustachian tube function.

OBJECTIVE: Tympanometry is a relatively simple method to assess middle ear (ME) status and pressure. Daily, serial tympanometric measurements may contain information on the constitutional efficiency of Eustachian tube function (ETF). A study was conducted to determine family compliance with an effort-intensive protocol that requires daily tympanometry done on each child, daily symptom recording by a parent and weekly visits with study personnel and to abstract measures that summarize the data as they relate to ETF. METHODS: Longitudinal 6-month, daily follow-up on 249 children from 123 families by parent-recorded bilateral tympanometry and cold-like illnesses. RESULTS: Study attrition was minimal with 90% of the enrolled families completing 90% of the study period. Parental compliance with daily tympanometry was less. Overall, 67% of the families were 80% compliant and 51% were 90% compliant. Two measures (aMEP, average middle ear pressure; vMEP, standard deviation of MEP) were evaluated for summaries of the longitudinal tympanometric results. These measures were not bilaterally independent or mutually independent and were affected by cold-burden and age. The best summary measure of constitutional ET efficiency may be a dimensional map of vMEP on aMEP with subsequent subdivision of the dimensional space into zones with functional interpretations. CONCLUSION: Long-term studies that require daily parental recording of tympanometry and the presence/absence of cold-like illness are feasible. It is expected that compliance with the required procedures can be increased substantially by pre-selection of candidate families. Preliminary data analyses suggest that the longitudinal tympanometric data contains important information on constitutional ETF.

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.

Upper respiratory virus detection without parent-reported illness in children is virus-specific.

BACKGROUND: Viral upper respiratory tract infection (vURI) may or may not present with a cold/flu-like illness (CFLI). OBJECTIVES: For common upper respiratory viruses that cause vURIs, to determine the relative frequencies of virus detection by PCR in subjects with and without CFLIs. STUDY DESIGN: Prospective follow-up of 170 children aged 1-8.6 years through the CFLI season by daily parental diary for CFLI episodes and nasal secretion sampling using PCR assays for adenovirus, coronavirus (types 229E and OC43), influenza virus (types A and B), parainfluenza (types 1-3) virus, rhinovirus, and respiratory syncytial virus (RSV). RESULTS: Virus was detected in 415 of 956 independent assays: 425 CFLI episodes and 531 non-CFLI periods were sampled; samples from 270 (64%) CFLI episodes and 145 (27%) non-CFLI periods contained virus detected by PCR. Rhinovirus was most frequently detected at 64%, followed by mixed viruses at 12%, RSV at 7%, and the other viruses at 3-5% of all detections. About 85% of RSV, influenza A and adenovirus detections were associated with a CFLI, whereas less than 62% of other virus detections were associated with CFLI. CONCLUSIONS: The frequency of PCR virus detection without CFLI was different among viruses. This introduces virus-specific biases to estimating the frequencies of specific complications attributable to a vURI when ascertained by CFLI identification.

Daily tympanometry for high-resolution measurement of the time between onset of cold-like illness and middle ear effusion.

BACKGROUND: Tympanometry is a simple method to assess middle ear pressure (MEP) and the presence of middle ear effusion (MEE), a marker of otitis media (OM). OBJECTIVES: To determine whether daily parental tympanometry and illness sign recording in their children can be used to define the time between onsets of cold-like illness (CLI) and MEE at high resolution. STUDY DESIGN: Prospective, longitudinal, 7 month, daily follow-up on 169 children aged 1 through 8.6 years. METHODS: Tympanograms and illness were recorded daily by a parent. Tympanograms were examined, rejected if artifactual, and MEP data were entered into a database, with "flat tympanograms" coded as -400 mm H2O = MEE. The incidence and burden of CLIs (>2 days) were calculated, and for each CLI, the presence/absence of concurrent MEE (>2 days) was determined. For each child, the average MEP for CLI and nonCLI days was calculated. Paired CLI and tympanometric results were aligned and the days between event onsets determined. Stepwise regression was used to assign risk predictors for the measured outcomes. RESULTS: A total of 566 CLIs were recorded, and the average CLI burden/child was 16%. Age was a significant predictor of CLI incidence/child, and age, history of colds, and daily environment were predictors of CLI-burden/child. Of the 433 evaluable CLI episodes, MEE was a complication in 37%, and MEE with a CLI was predicted by age, OM history, and environment. MEP was significantly more negative during CLI episodes, and the magnitude was predicted by age, race, and OM history. The average difference in MEE-CLI onsets was 1.2 +/- 4.0 days; approximately 32% of MEEs occurred prior to CLI onset and 17% on the same day as CLI onset. CONCLUSIONS: CLIs adversely affect the middle ear-ambient pressure balance and are frequently associated with MEE episodes. The distribution in onsets between those events suggests that chemoprophylaxis of a child with a newly identified CLI to prevent MEE would have a low expected efficiency.

The incidence, prevalence and burden of OM in unselected children aged 1-8 years followed by weekly otoscopy through the "common cold" season.

BACKGROUND: There is a continuing interest in defining the incidence, prevalence and burden of otitis media (OM) in the individual and population for purposes of assigning "risk factors". Often overlooked in past studies are the contributions of cold-like illnesses (CLIs) and sampling interval to those estimates. OBJECTIVE: Describe the incidence of symptomatic (AOM) and asymptomatic (OME) OM, the prevalence of OM, the contribution of CLI incidence, burden and other OM "risk factors" to the incidence and burden of OM, and the effect of sampling interval on those measures in children. METHODS: 148 children (74 male; 131 white, aged 1.0-8.6 years) were followed from November 1 to April 30 by weekly pneumatic otoscopy to diagnose OM presence/absence and by daily parental diary to assign CLI episodes. Data for previously identified OM "risk factors" were collected on 127. Results were summarized using standard measures of incidence, prevalence and burden, and multiple regression techniques were used to identify OM "risk factors". RESULTS: The basal OM prevalence was 20% with peaks in December and March and the temporal pattern was correlated with CLI prevalence. The incidence of OME (per 27,232 child-days) was 317, AOM was 74 and CLI was 456. The seasonal pattern of AOM and OME incidences tracked and was correlated with that for CLIs. New OM episodes were usually of short duration (