Immunofluorescence versus xTAG multiplex PCR for the detection of respiratory picornavirus infections in children

BACKGROUND: Polymerase chain reaction (PCR) is a sensitive tool for detection of respiratory picornaviruses. However, the clinical relevance of picornavirus detection by PCR is unclear. Immunofluorescence (IF), widely used to detect other respiratory viruses, has recently been introduced as a promising detection method for respiratory picornaviruses. OBJECTIVES: To compare the clinical manifestations of respiratory picornavirus infections detected by IF with those of respiratory picornavirus infections detected by xTAG multiplex PCR in hospitalized children. STUDY DESIGN: During a 1-year period, nasopharyngeal aspirates (NPA) from all children hospitalized due to an acute respiratory infection were prospectively analyzed by IF. All respiratory picornavirus positive IF samples and 100 IF negative samples were further tested with xTAG multiplex PCR. After exclusion of children with co-morbidities and viral co-infections, monoinfections with respiratory picornaviruses were detected in 108 NPA of 108 otherwise healthy children by IF and/or PCR. We compared group 1 children (IF and PCR positive, n=84) with group 2 children (IF negative and PCR positive, n=24) with regard to clinical manifestations of the infection. RESULTS: Wheezy bronchitis was diagnosed more often in group 1 than in group 2 (71% vs. 46%, p=0.028). In contrast, group 2 patients were diagnosed more frequently with pneumonia (17% vs. 6%, p=0.014) accompanied by higher levels of C-reactive protein (46mg/l vs. 11mg/l, p=0.009). CONCLUSIONS: Picornavirus detection by IF in children with acute respiratory infection is associated with the clinical presentation of wheezy bronchitis. The finding of a more frequent diagnosis of pneumonia in picornavirus PCR positive but IF negative children warrants further investigation.

Imatinib mesylate selectively impairs expansion of memory cytotoxic T cells without affecting the control of primary viral infections

Imatinib mesylate (imatinib) is a potent inhibitor of defined tyrosine kinases (TKs) and is effective in the treatment of malignancies characterized by constitutive activation of these TKs such as chronic myeloid leukemia and gastrointestinal stromal tumors. TKs also play an important role in T-cell receptor (TCR) signal transduction. Inhibitory as well as stimulating effects of imatinib on T cells and dendritic cells have been described. Here, we analyzed the effects of imatinib treatment on antiviral immune responses in vivo. Primary cytotoxic T-cell (CTL) responses were not impaired in imatinib-treated mice after infection with lymphocytic choriomeningitis virus (LCMV) or after immunization with a tumor cell line expressing LCMV glycoprotein (LCMV-GP). Similarly, neutralizing antibody responses to vesicular stomatitis virus (VSV) were not affected. In contrast, secondary expansion of LCMV-specific memory CTLs was reduced in vitro and in vivo, resulting in impaired protection against reinfection. In addition, imatinib treatment delayed the onset of diabetes in a CTL-induced diabetes model. In summary, imatinib treatment in vivo selectively inhibits the expansion of antigen-experienced memory CTLs without affecting primary T- or B-cell responses. Therefore, imatinib may be efficacious in the suppression of CTL-mediated immunopathology in autoimmune diseases without the risk of acquiring viral infections.

Oral purified bacterial extracts in acute respiratory tract infections in childhood: a systematic quantitative review

BACKGROUND: Recurrent acute respiratory tract infections (ARTI) are a common problem in childhood. Some evidence suggests a benefit regarding the prevention of ARTI in children treated with the immunomodulator OM-85 BV (Bronchovaxom). METHODS: We summarised the evidence on the effectiveness of the immunomodulator OM-85 BV in the prevention of ARTI in children. We searched randomised comparisons of oral purified bacterial extracts against inactive controls in children with respiratory tract diseases in nine electronic databases and reference lists of included studies. We extracted salient features of each study, calculated relative risks (RR) or weighted mean differences (WMD) and performed meta-analyses using random-effects models. RESULTS: Thirteen studies (2,721 patients) of low to moderate quality tested OM-85 BV. Patients and outcomes differed substantially, which impeded pooling results of more than two trials. Two studies (240 patients) reporting on the number of patients with less than three infections over 6 month of follow-up in children not in day care showed a trend for benefit RR 0.82 (95% CI, 0.65-1.02). One out of two studies examining the number of children not in day care without infections over 4-6 month reported a significant RR of 0.42 (95% CI, 0.21-0.82) whereas the smaller, second study did not [RR 0.92 (95% CI, 0.58-1.46)]. Two studies reporting the number of antibiotic courses indicated a benefit for the intervention arm [WMD 2.0 (95% CI, 1.7-2.3)]. Two out of the three studies showed a reduction of length of episodes of 4-6 days whereas a third study showed no difference between the two groups. CONCLUSION: Evidence in favour of OM-85 BV in the prevention of ARTI in children is weak. There is a trend for fewer and shorter infections and a reduction of antibiotic use.

Clinical Characteristics of Children with Viral Single- and Co-Infections and a Petechial Rash

BACKGROUND:: Children with petechial rash are more likely to undergo invasive diagnostics, to be treated with antibiotics for potential bacterial infection and to be hospitalized. However, viruses have also been associated with petechial rash. Nonetheless, a systematic analysis of viral infections with modern available techniques as quantitative real time polymerase chain reaction (q-PCR) in the context of petechial rash is lacking. The purpose of this pediatric study was to prospectively uncover viral pathogens that may promote the emergence of petechiae and to analyze the correlation with the clinical characteristics and course. METHODS:: We conducted a prospective study in children (0 to 18 years) presenting with petechiae and signs or symptoms of infection at the emergency department between November 2009 and March 2012. In nasopharyngeal aspirates the following viruses were analyzed by q-PCR: Cytomegalovirus, Epstein-Barr virus, parvovirus B19, Influenza A and B, parainfluenza viruses, human respiratory syncytial virus A and B, human metapneumovirus, rhinovirus, enterovirus, adenovirus, human coronavirus OC43, 229E, NL63 and human bocavirus. RESULTS:: A viral pathogen was identified in 67% of the analyzed 58 cases with petechial rash. Virus positive patients showed a significantly higher incidence of lower respiratory tract infections. Forty-one percent were viral co-infections, which were significantly younger than virus negative patients, had a higher leukocyte count and were hospitalized for a longer time. CONCLUSIONS:: A petechial rash is frequently associated viral single- and co-infections and can rapidly be identified via q-PCR.

A prospective study of the impact of air pollution on respiratory symptoms and infections in infants

Rationale: There is increasing evidence that short-term exposure to air pollution has a detrimental effect on respiratory health, but data from healthy populations, particularly infants, are scarce. Objectives: To assess the association of air pollution with frequency and severity of respiratory symptoms and infections measured weekly in healthy infants. Methods: In a prospective birth cohort of 366 infants of unselected mothers, respiratory health was assessed weekly by telephone interviews during the first year of life (19,106 total observations). Daily mean levels of particulate matter (PM10), nitrogen dioxide (NO2), and ozone (O3) were obtained from local monitoring stations. We determined the association of the preceding week's pollutant levels with symptom scores and respiratory tract infections using a generalized additive mixed model with an autoregressive component. In addition, we assessed whether neonatal lung function influences this association and whether duration of infectious episodes differed between weeks with normal PM10 and weeks with elevated levels. Measurements and Main Results: We found a significant association between air pollution and respiratory symptoms, particularly in the week after respiratory tract infections (risk ratio, 1.13 [1.02-1.24] per 10 μg/m(3) PM10 levels) and in infants with premorbid lung function. During times of elevated PM10 (>33.3 μg/m(3)), duration of respiratory tract infections increased by 20% (95% confidence interval, 2-42%). Conclusions: Exposure to even moderate levels of air pollution was associated with increased respiratory symptoms in healthy infants. Particularly in infants with premorbid lung function and inflammation, air pollution contributed to longer duration of infectious episodes with a potentially large socioeconomic impact.

Rhinovirus infections in infancy and early childhood

Rhinovirus (RV) infections occur early and recurrently in life, imposing a significant burden of disease on infants and young children. They are the most frequent causative agents of both upper and lower respiratory tract infections in this age group and are associated with a broad variety of clinical outcomes, ranging from asymptomatic infections to severe respiratory disease requiring hospitalisation. In addition to their impact on short-term morbidity, RVs are also debated as important pathogens in the development of recurrent wheeze and/or asthma. Several studies in infants at high-risk for atopy and asthma and in hospitalised children have demonstrated that recurrent wheezing illnesses induced by RVs early in life are a risk factor for the development of asthma later in childhood. However, underlying mechanisms are poorly understood. The question whether RVs are directly involved in the development of childhood wheeze and asthma, or whether symptomatic RV infections only represent a proxy for infants prone to develop obstructive lung diseases, is still open. In this review we provide an overview on the role of RVs as important disease-causing agents from infancy to early childhood and discuss their contribution to the subsequent development of childhood wheeze and/or asthma.

[How the bovine viral diarrhea virus outwits the immune system]

The interaction of bovine viral diarrhea virus (BVD virus) with its host has several unique features, most notably the capacity to infect its host either transiently or persistently. The transient infection stimulates an antiviral immune reaction similar to that seen in other transient viral infections. In contrast, being associated with immunotolerance specific for the infecting BVD viral strain, the persistent infection differs fundamentally from other persistent infections like those caused by lentiviruses. Whereas the latter are characterized by complex viral evasion of the host's adaptive immune response by mechanisms such as antigenic drift and interference with presentation of T cell epitopes, BVD virus avoids the immune response altogether by inducing both humoral and cellular immune tolerance. This is made possible by invasion of the fetus at an early stage of development. In addition to adaptive immunity, BVD virus also manipulates key elements of the host's innate immune response. The non-cytopathic biotype of BVD virus, which is capable of persistently infecting its host, fails to induce type I interferon. In addition, persistently infected cells are resistant to the induction of apoptosis by double-stranded RNA and do not produce interferon when treated with this pathogen-associated molecular pattern (PAMP) that signals viral infection. Moreover, when treated with interferon, cells persistently infected with non-cytopathic BVD virus do not clear the virus. Surprisingly, however, despite this lack of effect on persistent infection, interferon readily induces an antiviral state in these cells, as shown by the protection against infection by unrelated viruses. Overall, BVD virus manipulates the host's interferon defense in a manner that optimises its chances of maintaining the persistent infection as well as decreasing the risks that heterologous viral infections may carry for the host. Thus, since not all potential host cells are infected in animals persistently infected with BVD virus, heterologous viruses replicating in cells uninfected with BVD virus will still trigger production of interferon. Interferon produced by such cells will curtail the replication of heterologous viruses only, be that in cells already infected with BVD virus, or in cells in which the heterologous virus may replicate alone. From an evolutionary viewpoint, this strategy clearly enhances the chances of transmission of BVD virus to new hosts, as it attenuates the negative effects that a global immunosuppression would have on the survival of persistently infected animals.

The prevalence of penicillin-non-susceptible Streptococcus pneumoniae among children aged < 5 years correlates with the biannual epidemic activity of respiratory syncytial virus

This study investigated whether the epidemiology of penicillin-non-susceptible pneumococci (PNSP) colonising small children correlated with the biannual epidemic activity of respiratory syncytial virus (RSV). Colonisation rates and the prevalence of PNSP among paediatric outpatients aged < 5 years was analysed between January 1998 and September 2003 using an established national surveillance network. Resistance trends were investigated using time-series analysis to assess the correlation with the biannual pattern of RSV infections and national sales of oral paediatric formulations of antibiotics and antibiotic prescriptions to children aged < 5 years for acute respiratory tract infections. PNSP rates exhibited a biannual cycle in phase with the biannual seasonal RSV epidemics (p < 0.05). Resistance rates were higher during the winter seasons of 1998-1999 (20.1%), 2000-2001 (16.0%) and 2002-2003 (19.1%), compared with the winter seasons of 1997-1998 (8.2%), 1999-2000 (11.6%) and 2001-2002 (9.5%). Antibiotic sales and prescriptions showed regular peaks during each winter, with no significant correlation with the biannual pattern of RSV activity and seasonal trends of PNSP. RSV is an important determinant of the spread of PNSP and must be considered in strategies aimed at antimicrobial resistance control.

Snoring in preschool children: prevalence, severity and risk factors

Epidemiological data on snoring from preschool children are scarce, although habitual snoring (snoring on almost all nights) has been associated with poor long-term outcomes. In a population survey of 6,811 children aged 1-4 yrs (from Leicestershire, UK) the present authors determined prevalence, severity and risk factors for snoring, especially habitual snoring. In 59.7% of the children, parents reported snoring in the previous 12 months, including 7.9% with habitual snoring and 0.9% with habitual snoring and sleep disturbance. Prevalence of habitual snoring increased with age from 6.6% in 1-yr-olds to 13.0% in 4-yr-olds. Habitual snoring was associated with: one and both parents smoking (adjusted odds ratio (OR) 1.46 and 2.09, respectively); road traffic (OR 1.23); single parent (OR 1.60); and in White but not South Asian children, socioeconomic deprivation (OR 1.25 and 2.03 for middle and upper thirds of Townsend score, respectively). Respiratory tract symptoms related to atopic disorders and to respiratory infections were strongly associated with snoring; however, body mass index was not. In conclusion, habitual snoring is common in preschool children with one-third of cases attributable to avoidable risk factors. The strong association with atopic disorders, viral infections and environmental exposures suggests a complex aetiology, based on a general vulnerability of the respiratory tract.

Swiss recommendations for the management of varicella zoster virus infections

Infections with varicella zoster virus (VZV) are common viral infections associated with significant morbidity. Diagnosis and management are complex, particularly in immunocompromised patients and during pregnancy. The present recommendations have been established by a multidisciplinary panel of specialists and endorsed by numerous Swiss medical societies involved in the medical care of such patients (Appendix). The aim was to improve the care of affected patients and to reduce complications.

How useful is the ultrastructural study of the cilia of the respiratory tract in the diagnosis of an immotile cilia syndrome?

The immotile cilia syndrome (ICS) comprises a range of congenital defects of the ciliary apparatus most probably transmitted by autosomal recessive inheritance. Because cilia occur mainly in the respiratory and genital tract, the clinical symptoms of ICS are most commonly chronic sinusitis, bronchitis, bronchiectasis and male sterility. The syndrome can be associated with a situs inversus and is then called Kartagener's syndrome. We studied the ciliary ultrastructure in airway biopsies of 5 patients suffering from chronic upper and lower respiratory tract infections. With the single exception of one female patient with confirmed ICS diagnosis (Kartagener's syndrome) the etiology of the recurrent infections was unknown. The following ciliary defects were observed: missing dynein arms, radial spoke defects, missing nexin links, microtubular transpositions, compound cilia, supernumerary, absent, or incomplete microtubules, lack of ciliary orientation and various abnormal patterns of microtubular arrangement. In no instance did a patient show only a single anomaly; defects were always combined. Missing dynein arms, radial spoke defects and microtubular transpositions have frequently been described as lesions specific for ICS. Whenever these lesions were found simultaneously in both the respiratory and genital tracts, their genetic origin cannot be doubted. In our confirmed ICS patient the outer dynein arms were not missing but were reduced in number and length in a large number of cilia. The biopsy was, however, obtained from the heavily infected maxillary sinus and it is known that inflammation can lead to a loss of dynein arms. In the light of our investigations and of a review of the published cases of ciliary anomalies, it is concluded that none of the above defects in itself is specific for ICS. They may all occur as secondary lesions or sporadically as varieties in otherwise healthy subjects. It therefore appears questionable whether ICS can be diagnosed from the ciliary ultrastructure of a single airway biopsy. Assessment of ICS cannot be based simply on the ultrastructural demonstration of a particular ciliary defect, but necessitates additional considerations particularly regarding the origin of the biopsy, the sampling procedures and quantitation of defects. It appears necessary to investigate samples from different parts of the airways and quantitatively analyze the prominent lesions.

Quantitative multiparameter assays to measure the effect of adjuvants on human antigen-specific CD8 T-cell responses

Large numbers and functionally competent T cells are required to protect from diseases for which antibody-based vaccines have consistently failed (1), which is the case for many chronic viral infections and solid tumors. Therefore, therapeutic vaccines aim at the induction of strong antigen-specific T-cell responses. Novel adjuvants have considerably improved the capacity of synthetic vaccines to activate T cells, but more research is necessary to identify optimal compositions of potent vaccine formulations. Consequently, there is a great need to develop accurate methods for the efficient identification of antigen-specific T cells and the assessment of their functional characteristics directly ex vivo. In this regard, hundreds of clinical vaccination trials have been implemented during the last 15 years, and monitoring techniques become more and more standardized.

Absence of MyD88 results in enhanced TLR3-dependent phosphorylation of IRF3 and increased IFN-(beta) and RANTES production

Toll-like receptors are a group of pattern-recognition receptors that play a crucial role in "danger" recognition and induction of the innate immune response against bacterial and viral infections. TLR3 has emerged as a key sensor of viral dsRNA, resulting in the induction of the anti-viral molecule, IFN- . Thus, a clearer understanding of the biological processes that modulate TLR3 signaling is essential. Previous studies have shown that the TLR adaptor, Mal/TIRAP, an activator of TLR4, inhibits TLR3-mediated IFN- induction through a mechanism involving IRF7. In this study, we sought to investigate whether the TLR adaptor, MyD88, an activator of all TLRs except TLR3, has the ability to modulate TLR3 signaling. Although MyD88 does not significantly affect TLR3 ligand-induced TNF- induction, MyD88 negatively regulates TLR3-, but not TLR4-, mediated IFN- and RANTES production; this process is mechanistically distinct from that employed by Mal/TIRAP. We show that MyD88 inhibits IKK -, but not TBK1-, induced activation of IRF3. In doing so, MyD88 curtails TLR3 ligand-induced IFN- induction. The present study shows that while MyD88 activates all TLRs except TLR3, MyD88 also functions as a negative regulator of TLR3. Thus, MyD88 is essential in restricting TLR3 signaling, thereby protecting the host from unwanted immunopathologies associated with the excessive production of IFN- . Our study offers a new role for MyD88 in restricting TLR3 signaling through a hitherto unknown mechanism whereby MyD88 specifically impairs IKK -mediated induction of IRF3 and concomitant IFN- and RANTES production.