Tytti Vuorinen

Human bocavirus and acute wheezing in children.

Abstract Background. Human bocavirus is a newly discovered parvovirus. It has been detected primarily in children with acute lower respiratory tract infection, but its occurrence, clinical profile, and role as a causative agent of respiratory tract disease are not clear. Methods. We investigated the presence of human bocavirus by quantitative polymerase chain reaction of nasopharyngeal aspirate specimens and selected serum samples obtained from 259 children (median age, 1.6 years) who had been hospitalized for acute expiratory wheezing. The samples were analyzed for 16 respiratory viruses by polymerase chain reaction, virus culture, antigen detection, and serological assays. Results. At least 1 potential etiologic agent was detected in 95% of children, and >1 agent was detected in 34% of children. Human bocavirus was detected in 49 children (19%). A large proportion of the cases were mixed infections with other viruses, but human bocavirus was the only virus detected in 12 children (5%). High viral loads of human bocavirus were noted mainly in the absence of other viral agents, suggesting a causative role for acute wheezing. In addition, infections that had uncertain clinical relevance and low viral loads were prevalent. Human bocavirus DNA was frequently detected in serum specimens obtained from patients with acute wheezing, suggesting systemic infection. Conclusions. Human bocavirus is prevalent among children with acute wheezing and can cause systemic infection. Results suggest a model for bocavirus infection in which high viral loads are potentially associated with respiratory symptoms and low viral loads indicate asymptomatic shedding. Therefore, quantitative polymerase chain reaction analysis may be important for additional studies of human bocavirus.

Respiratory Picornaviruses and Respiratory Syncytial Virus as Causative Agents of Acute Expiratory Wheezing in Children

We studied the viral etiology of acute expiratory wheezing (bronchiolitis, acute asthma) in 293 hospitalized children in a 2-year prospective study in Finland. A potential causative viral agent was detected in 88% of the cases. Eleven different viruses were represented. Respiratory syncytial virus (RSV) (27%), enteroviruses (25%), rhinovirus (24%), and nontypable rhino/enterovirus (16%) were found most frequently. In infants, RSV was found in 54% and respiratory picornaviruses (rhinovirus and enteroviruses) in 42% of the cases. In older children, respiratory picornaviruses dominated (65% of children ages 1-2 years and 82% of children ages >3 years). Human metapneumovirus was detected in 4% of all children and in 11% of infants. To prevent and treat acute expiratory wheezing illnesses in children, efforts should be focused on RSV, enterovirus, and rhinovirus infections.

Burden of Influenza in Children in the Community

BACKGROUND Influenza vaccination of healthy children is encouraged because children are frequently hospitalized for influenza-attributable illnesses. However, most children with influenza are treated as outpatients, and scarce data are available on the burden of influenza in these children. METHODS We performed a prospective study of respiratory infections in preenrolled cohorts of children < or = 13 years old during 2 consecutive respiratory seasons (2231 child-seasons of follow-up). At any sign of respiratory infection, we examined the children and obtained a nasal swab for the detection of influenza. The parents filled out daily symptom diaries. Of all the enrollees, 94% remained active participants in the study. RESULTS The average annual rate of influenza was highest (179 cases/1000 children) among children < 3 years old. Acute otitis media developed as a complication of influenza in 39.7% of children < 3 years old. For every 100 influenza-infected children < 3 years old, there were 195 days of parental work loss (mean duration, 3.2 days). CONCLUSIONS Influenza causes a substantial burden of illness on outpatient children and their families. Vaccination of children < 3 years old might be beneficial for reducing the direct and indirect costs of influenza in children.

Coxsackievirus A6 and Hand, Foot, and Mouth Disease, Finland

During fall 2008, an outbreak of hand, foot, and mouth disease (HFMD) with onychomadesis (nail shedding) as a common feature occurred in Finland. We identified an unusual enterovirus type, coxsackievirus A6 (CVA6), as the causative agent. CVA6 infections may be emerging as a new and major cause of epidemic HFMD.

Co-circulation of coxsackieviruses A6 and A10 in hand, foot and mouth disease outbreak in Finland

BACKGROUND A nationwide outbreak of hand, foot and mouth disease (HFMD) occurred in Finland in autumn 2008. The outbreak was untypical since a considerable number of clinically diagnosed patients were adults. Furthermore, many of the patients suffered from onychomadesis several weeks after the acute phase of HFMD. OBJECTIVES Detection, identification and phylogenetic analysis of human enteroviruses (HEV) that caused the outbreak. STUDY DESIGN A total of 420 clinical specimens were obtained from 317 HFMD cases all over the country. The presence of HEV in the specimens was analysed by virus isolation and/or direct real-time RT-PCR; selected HEV strains were further typed by molecular methods. The genetic similarities of HEV strains were assessed by phylogenetic analyses on partial VP1 sequences. RESULTS HEV were detected in 212 HFMD cases, including both children and adults, throughout Finland. Two HEV types, coxsackieviruses A6 (CV-A6) and A10 (CV-A10), were identified as the causative agents of the outbreak. One genetic variant of CV-A6 predominated, but, additionally, three other genetically distinct CV-A6 strains were found. All CV-A10 strains segregated into one genetic cluster distinct from previously reported CV-A10 sequences. CONCLUSIONS The Finnish 2008 HFMD outbreak was caused by two infrequently detected, co-circulating, coxsackie A viruses. Our data suggest endemic circulation of both CV-A types in Northern Europe and that the outbreak was due to the emergence of new genetic variants of these viruses.

Bronchiolitis: age and previous wheezing episodes are linked to viral etiology and atopic characteristics.

Background: Diagnostic criteria for bronchiolitis are variable. Objective: To study how the risk factors for recurrent wheezing and asthma vary by different definitions of bronchiolitis. Methods: Viral etiology and atopic characteristics were studied in 259 hospitalized wheezing children (median age, 14 months; range, 0–36 months). The data were analyzed according to age (<6, <12, <24 and <36 months) and whether they had a history or no history of a previous wheezing episode. Sixteen viruses were detected by conventional and molecular methods. Atopic characteristics included the presence of eczema, specific and total IgE responses, blood eosinophil count, and modified asthma predictive index. Results: Evidence of respiratory virus infection was found in 93% of the cases and allergic sensitization in 26% of the cases. Rhinovirus infections and atopic characteristics (sensitization, blood eosinophil count, and modified asthma predictive index) increased by age and were significantly more common in children with recurrent wheezing episodes than in first-time wheezers in age categories of <24 and <36 months (P < 0.05 for all). Conclusions: In children with bronchiolitis, 2 clinical factors, age and number of previous wheezing episodes, are linked to inflammatory (atopy-related factors) and virologic risk factors of asthma (rhinovirus-associated disease). According to current US and UK guidelines, bronchiolitis includes wheezing children <24 months of age. Our observations suggest that the clinical definition should include only children with their first episode of wheezing.

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

Background: The role of systemic corticosteroids in the treatment of early childhood wheezing in children is not clear. Objective: We sought to determine whether prednisolone is effective in rhinovirus-induced early wheezing. Methods: We conducted a controlled trial comparing oral prednisolone (2 mg/kg per day in three divided doses for 3 days) with placebo in 78 hospitalized children (mean age, 1.1 year; standard deviation, 0.7) experiencing their first or second episode of wheezing induced by rhinovirus or respiratory syncytial virus. Mixed viral infections were excluded. Our primary end point was the time until the patient was ready for discharge; secondary end points included oxygen saturation during hospitalization, duration of symptoms, occurrence of relapses during the next 2 months and blood eosinophil counts at discharge and 2 weeks later. Results: In multivariate regression analysis, prednisolone did not influence the time until ready for discharge, but it decreased relapses during the subsequent 2-month period in rhinovirus-affected children (prednisolone versus placebo, 22% versus 56%; odds ratio, 19.06; 95% confidence interval, 2.52–144.03; P = 0.004) and in children with blood eosinophils ≥0.2 × 109/L (respectively, 24% versus 71%; odds ratio, 10.57; 95% confidence interval, 1.99–56.22; P = 0.006). Rhinovirus-affected children had more blood eosinophils on admission (mean, 0.44 versus 0.086 × 109/L), had a higher prevalence of atopy (44% versus 8%) and were older (mean, 1.4 versus 0.9 years, P < 0.001 for all) than respiratory syncytial virus-infected children. Conclusion: Prednisolone reduced relapses during a 2-month period after first episodes of wheezing associated with rhinovirus infection or blood eosinophils ≥0.2 × 109/L.

Diagnosis of Enteroviral Meningitis by Use of Polymerase Chain Reaction of Cerebrospinal Fluid, Stool, and Serum Specimens

BACKGROUND Because enteroviruses can be detected in various clinical samples during enteroviral meningitis, we analyzed the combined diagnostic utility of polymerase chain reaction (PCR) of cerebrospinal fluid (CSF), feces, and serum for detection of enterovirus in specimens obtained from adults with aseptic meningitis or encephalitis. METHODS PCR results were analyzed for 34 adults for whom enteroviral meningitis was diagnosed on the basis of virus isolation and antibody detection in our hospital during 1999-2003. PCR results were also analyzed for 77 adults with meningitis or encephalitis of another defined cause for whom this assay was used for diagnostic evaluation during that period. RESULTS Twenty-six (76%) of 34 CSF samples and 24 (96%) of 25 fecal samples collected from patients with enteroviral meningitis had positive PCR results. The diagnostic yield of the test was lower for CSF specimens obtained >2 days after clinical onset, compared with CSF collected < or =2 days after onset. Instead, PCR of feces was highly useful also later, because 12 of the 13 fecal specimens obtained 5-16 days after clinical onset had positive test results. None of 75 CSF samples and 2 of 48 fecal samples obtained from patients with nonenteroviral infection had positive PCR results. All serum samples were PCR negative. CONCLUSIONS PCR of fecal specimens obtained throughout the course of enteroviral meningitis had the highest clinical sensitivity for detecting enterovirus. It is recommended that, in addition to performance of CSF PCR, fecal samples collected from patients with suspected enteroviral meningitis should be tested by PCR, especially when the duration of symptoms is >2 days.

Cardiomyocyte apoptosis in experimental coxsackievirus B3 myocarditis

INTRODUCTION Viruses are known to induce apoptosis in their host cells. We studied whether cardiomyocyte apoptosis occurs upon coxsackievirus B3 (CVB3) infection and whether virus-associated apoptosis plays a role in the pathogenesis of experimental myocarditis. METHODS BALB/c mice were infected with two variants of CVB3 causing either mild or severe myocarditis. Myocardial and serum samples were collected from Day 1 to Day 14 after virus inoculation. Apoptosis was detected in myocardial tissue sections using the terminal transferase-mediated DNA nick end labelling (TUNEL) assay and staining of active caspase 3, and compared with the presence of infectious CVB3 and viral proteins in cardiomyocytes. RESULTS Compared with the noninfected control mice, infection with either CVB3 variant resulted in significantly increased cardiomyocyte apoptosis, which peaked on Day 5 after infection. At this time, the average percentages of apoptotic cardiomyocytes were 0.17% (SD 0.04; P=.03) and 0.77% (SD 0.11; P<.01) in mild and severe disease forms, respectively. The amount of apoptosis correlated with titers of infectious CVB3 in the heart muscle. Viral proteins were detected in the TUNEL-positive cells by immunohistochemistry. In the late stages of disease, apoptosis, together with inflammatory infiltrates persisted only in the severe disease form. CONCLUSIONS CVB3-associated myocardial damage involves cardiomyocyte apoptosis. In the early stages of the disease, it appears to be triggered by viral replication in the cardiomyocytes.

Cytomegalovirus Infection of the Heart Is Common in Patients with Fatal Myocarditis

BACKGROUND Although enteroviruses and adenoviruses are considered to be the leading causes of the usually mild clinical myocarditis, little is known about the etiology of severe or fatal myocarditis. METHODS We collected all available clinical records and myocardial autopsy samples for patients who had myocarditis recorded as the underlying cause of death in Finland during the period of 1970-1998. Findings for all available patients (20 men and 20 women; median age, 49 years) with myocarditis that fulfilled the Dallas criteria and who had sufficient data were included in the study. Twelve subjects who had died accidentally served as control subjects. Polymerase chain reaction (PCR) and in situ hybridization assays were used for detection of viral genomes (adenovirus, cytomegalovirus, enterovirus, human herpesvirus 6, influenza A and B viruses, parvovirus B19, and rhinovirus) in heart samples. RESULTS Viral nucleic acids were found in the hearts of 17 patients (43%), including cytomegalovirus (15 patients), parvovirus B19 (4 patients), enterovirus (1 patient), and human herpesvirus 6 (1 patient). In 4 patients, cytomegalovirus DNA was found in addition to parvovirus B19 or enterovirus genomes. No adenoviruses, rhinoviruses, or influenza viruses were detected in this study of fatal myocarditis. In 67% of the patients for whom PCR was positive for cytomegalovirus, in situ hybridization revealed viral DNA in cardiomyocytes. Only 1 of these patients was immunocompromised. In the control group, only human herpesvirus 6 (1 subject) and parvovirus B19 (1 subject) DNA were detected. CONCLUSIONS In this population-based study, cytomegalovirus was found to be the most common specific finding in immunocompetent patients with fatal myocarditis. This may have important clinical implications for the treatment of severe acute myocarditis.