Background: Cockroach is one of the most important sources of indoor allergens and can lead to IgE sensitization and development of rhinitis and asthma. Objective: We sought to perform a cockroach allergen component analysis to determine the allergens and antibody levels and patterns of sensitization associated with asthma and rhinitis. Methods: Antibody (IgE, IgG, and IgG(4)) levels to total cockroach and 8 cockroach allergens were determined in 2 groups of cockroach-sensitized 10-year-old children with (n = 19) or without (n = 28) asthma and rhinitis. Allergen-specific antibody levels were measured in streptavidin ImmunoCAPs loaded with each of the recombinant allergens from groups 1, 2, 4, 5, 6, 7, 9, and 11, and total cockroach-specific IgE levels were measured with the i6 ImmunoCAP. Results: IgE antibody levels to cockroach allergens and extract, but not IgG or IgG(4) antibody levels, differed between subjects with and without asthma and rhinitis. Specifically, recognition of more cockroach allergens with higher allergen-specific IgE levels was associated with disease. Variable patterns of sensitization with no immunodominant allergens were found in both groups. There was a good correlation between the sum of allergen-specific IgE and total cockroach IgE levels (r = 0.86, P < .001). Conclusions: Component analysis of 8 cockroach allergens revealed significant differences in IgE reactivity associated with the presence of asthma and rhinitis. Allergen-specific IgE titers and sensitization profiles were associated with asthma and rhinitis.

A new and potentially more pathogenic group of human rhinovirus (HRV), group C (HRVC), has recently been discovered. We hypothesised that HRVC would be present in children with acute asthma and cause more severe attacks than other viruses or HRV groups. Children with acute asthma (n = 128; age 2–16 yrs) were recruited on presentation to an emergency department. Asthma exacerbation severity was assessed, and respiratory viruses and HRV strains were identified in a nasal aspirate. The majority of the children studied had moderate-to-severe asthma (85.2%) and 98.9% were admitted to hospital. HRV was detected in 87.5% and other respiratory viruses in 14.8% of children, most of whom also had HRV. HRVC was present in the majority of children with acute asthma (59.4%) and associated with more severe asthma. Children with HRVC (n = 76) had higher asthma severity scores than children whose HRV infection was HRVA or HRVB only (n = 34; p = 0.018), and all other children (n = 50; p = 0.016). Of the 19 children with a non-HRV virus, 13 had HRV co-infections, seven of these being HRVC. HRVC accounts for the majority of asthma attacks in children presenting to hospital and causes more severe attacks than previously known HRV groups and other viruses.

Rationale: Most asthma exacerbations are initiated by viral upper respiratory illnesses. It is unclear whether human rhinovirus (HRV)–induced exacerbations are associated with greater viral replication and neutrophilic inflammation compared with HRV colds.Objectives: To evaluate viral strain and load in a prospective asthma cohort during a natural cold.Methods: Adults were enrolled at the first sign of a cold, with daily monitoring of symptoms, medication use, and peak expiratory flow rate until resolution. Serial nasal lavage and induced sputum samples were assessed for viral copy number and inflammatory cell counts.Measurements and Main Results: A total of 52 persons with asthma and 14 control subjects without atopy or asthma were studied for over 10 weeks per subject on average; 25 participants developed an asthma exacerbation. Detection of HRVs in the preceding 5 days was the most common attributable exposure related to exacerbation. Compared with other infections, those by a minor group A HRV were 4.4-fold more likely to cause exacerbation (P = 0.038). Overall, sputum neutrophils and the burden of rhinovirus in the lower airway were similar in control subjects without atopy and the asthma group. However, among HRV-infected participants with asthma, exacerbations were associated with greater sputum neutrophil counts (P = 0.005).Conclusions: HRV infection is a frequent cause of exacerbations in adults with asthma and a cold, and there may be group-specific differences in severity of these events. The absence of large differences in viral burden among groups suggests differential lower airway sensitization to the effects of neutrophilic inflammation in the patients having exacerbations.

RATIONALE: Accumulating evidence suggests a role for regulatory T cells in balancing inflammatory responses. The role of T regulatory cell subsets in the expression of atopic eczema (AE) in childhood is not well characterized. METHODS: PBMCs were obtained from genetically at-risk children enrolled in the Childhood Origins of ASThma (COAST) cohort at ages 8-9. Analyses included flow cytometry for T regulatory cell (Treg) markers (CD25, CD127 and Foxp3), PHA-induced cytokine levels, and intracellular cytokine staining. RESULTS: Distinct subsets of Foxp3+ cells exist, based on levels of Foxp3 and CD25 expression. Upon stimulation, there is an increase in %Foxp3+/CD4+ cells in children with (5.22% vs 11.5%, p=0.004) and without (5.75% vs 8.94%, p=0.03) AE. It is the CD25intermediateFoxp3intermediate T cells that appear to upregulate Foxp3. In children with AE, Foxp3+ cells have increased mean fluorescent intensity of Foxp3 (490 vs 1090, p=0.01) with stimulation. Interestingly, the main source IL-10 in stimulated PBMC was CD14+ monocytic cells (50.0% CD14+/IL-10+ cells vs 18.5% Foxp3+/IL-10+ cells), and children with AE had increased IL-10 expression in the CD14+ population (20.72% IL-10+/CD14+ cells with AE vs 10.72% IL-10+/CD14+ cells without AE p=0.08). CONCLUSIONS: There is heterogeneity in Foxp3 expressing T cells and a differential response of these cells occurs with stimulation in children with and without AE. Increased IL-10 expression in the monocytic population may suggest an important role for antigen presenting cells in regulatory T cell responses in AE.

RATIONALE: Previously, mitogen-induced interferon (IFN) responses at birth were shown to be inversely correlated with illness frequency. Here we more comprehensively evaluate whether variations in immunologic responses at birth to multiple innate and mitogen stimuli are associated with the risk of acquiring respiratory illnesses during early childhood. METHODS: Cord blood mononuclear cells were obtained at birth from 285 children enrolled in the Childhood Origins of ASThma (COAST) study. After stimulating them with multiple mitogen and innate stimuli (including PHA, LPS, PMA, RV, RSV, and SAC), IFN-γ, IL-6, IL-10, IL-12, IFN-α, and TNF-α were measured. Cytokine levels were correlated to the frequency of moderate to severe respiratory illnesses (MSIs) during the first 3 years of life. A Spearman correlation was utilized for cytokine/stimuli pairs for which ≥75% of patient samples had detectable results; a Wilcoxon rank-sum test was used below this threshold. RESULTS: Of all stimuli/cytokine pairs analyzed, the most consistent finding was with IFNs. IFN-γ levels in cord blood were lower in children who developed more respiratory illnesses for the following stimulants: PMA (r = -0.16, p = 0.008), PHA (r = -0.12, p = 0.05), RSV (detectable vs. undetectable: 5.03 vs. 5.93 MSIs, p = 0.07), and LPS (detectable vs. undetectable: 3.83 vs. 6.2 MSIs, p = 0.1). Similarly, LPS-induced IFN-α levels were lower in children who developed more respiratory illnesses (r = -0.23, p = 0.02). CONCLUSIONS: Greater interferon responses to multiple innate and mitogen stimuli at birth appear to be protective against the development of frequent respiratory illnesses during early childhood.

RATIONALE: Accumulating evidence suggests a role for regulatory T cells in balancing inflammatory responses. The role of T regulatory cell subsets in the persistance vs remission of atopic dermatitis (AD) in childhood is not well characterized.METHODS: PBMCs were obtained from genetically at-risk children enrolled in the Childhood Origins of ASThma (COAST) cohort. Analyses included PHA-induced cytokine levels at age 5, flow cytometry for T regulatory cell (Treg) markers (CD25 and Foxp3) and intracellular cytokine staining. AD was assessed by at yearly intervals beginning at age 1 year.RESULTS: Of the 285 infants enrolled in the COAST study, 65 (23%) developed physician-diagnosed AD during infancy. Persistence of disease at age 5 yrs was associated with increased PHA-induced IL-10 from peripheral blood mononuclear cells (1088 vs 831 pg/mL, p = 0.01). On average, 2.14% of CD4+ lymphocytes are IL-10+ within this subset, 23.4% are Foxp3 + IL-10+ and 2.3% are IFN-γ + IL-10+; additionally, 7.09% of monocytes produce IL-10. At age 6, AD persistence was associated with decreased CD4 + CD25high T cells (1.55% vs 4.77%, p = 0.01), but no significant difference in CD4 + Foxp3 + T cells (4.3% vs 4.7%).CONCLUSIONS: Persistent AD was associated with increased IL-10, which appears to originate from monocytes and a small subset of lymphocytes suggestive of a predominately Tr1-type regulatory T cell population. These data suggest that blood CD25highTreg cells are quantitatively lower and are not the major cell source for the increased IL-10 in persistent AD. Ongoing studies are exploring the role of Treg cell subsets in the underlying disease pathogenesis. RATIONALE: Accumulating evidence suggests a role for regulatory T cells in balancing inflammatory responses. The role of T regulatory cell subsets in the persistance vs remission of atopic dermatitis (AD) in childhood is not well characterized. METHODS: PBMCs were obtained from genetically at-risk children enrolled in the Childhood Origins of ASThma (COAST) cohort. Analyses included PHA-induced cytokine levels at age 5, flow cytometry for T regulatory cell (Treg) markers (CD25 and Foxp3) and intracellular cytokine staining. AD was assessed by at yearly intervals beginning at age 1 year. RESULTS: Of the 285 infants enrolled in the COAST study, 65 (23%) developed physician-diagnosed AD during infancy. Persistence of disease at age 5 yrs was associated with increased PHA-induced IL-10 from peripheral blood mononuclear cells (1088 vs 831 pg/mL, p = 0.01). On average, 2.14% of CD4+ lymphocytes are IL-10+ within this subset, 23.4% are Foxp3 + IL-10+ and 2.3% are IFN-γ + IL-10+; additionally, 7.09% of monocytes produce IL-10. At age 6, AD persistence was associated with decreased CD4 + CD25high T cells (1.55% vs 4.77%, p = 0.01), but no significant difference in CD4 + Foxp3 + T cells (4.3% vs 4.7%). CONCLUSIONS: Persistent AD was associated with increased IL-10, which appears to originate from monocytes and a small subset of lymphocytes suggestive of a predominately Tr1-type regulatory T cell population. These data suggest that blood CD25highTreg cells are quantitatively lower and are not the major cell source for the increased IL-10 in persistent AD. Ongoing studies are exploring the role of Treg cell subsets in the underlying disease pathogenesis.

To better understand the viral aetiology of recurrent and prolonged illnesses, nasal secretions were prospectively collected from 285 infants at increased risk of developing asthma. Of these, 27 infants had recurrent (at least five) moderate-to-severe respiratory illnesses (MSIs). The viral aetiology of the 150 MSIs and 86 scheduled visits was analysed by molecular diagnostics. The demographic and clinical data were compared with infants who had 0-4 MSIs. Frequently ill infants had higher exposure to other children and more wheezing illnesses than less symptomatic children. Viruses were detected in 136 (91%) out of 150 MSIs, 14 (67%) out of 21 mild illnesses and 29 (45%) out of 65 asymptomatic visits. Human rhinovirus was the most common aetiological agent (61, 43 and 35% in MSIs, mild illnesses and asymptomatic visits, respectively). Mixed viral infections were generally associated with more severe illnesses (27, 0 and 5%, respectively). Among the 27 frequently ill infants, only eight (5.3%) out of 150 MSIs were prolonged (> or =2 weeks duration). Considering all samples, detection of the same virus strain > or =2 weeks apart was unusual (5.3% of all 244 positive findings). Human rhinovirus infections occur early, pervasively and repetitively in these high-risk infants. Infants with prolonged or recurrent respiratory illnesses most often have a series of infections rather than persistent infection with one virus strain.

RATIONALE: Accumulating evidence suggests a role for CD4+CD25+ T regulatory cells in balancing chronic inflammatory responses. The role of CD4+CD25+ T regulatory cells in atopic dermatitis (AD) and how it relates to further atopic disease is not well characterized. METHODS: PBMCs were obtained from children enrolled in the Childhood Origins of ASThma (COAST) cohort: a genetically at-risk population of infants. Analyses included PHA-induced cytokine levels, CD4+CD25+ T cells (% of CD4 cells) by flow cytometry and quantitation of FOXP3 mRNA in CD4+ T cells by real-time QPCR. RESULTS: Of the 285 infants enrolled in the COAST study, 65 (23%) developed physician-diagnosed AD during infancy. Persistence of disease at age 5 yrs was associated with increased PHA-induced IL-10 from peripheral blood (1088 vs 831 pg/mL, p=0.01). Preliminary data (65/255) suggest that AD at age 6 may be associated with increased FOXP3 expression (normalized relative expression, 35 vs 9, p=0.04). However, an increase in CD4+CD25high T cells is preliminarily associated with remission of disease at age 6 (n=13, 5.4% vs 3.3%, p=0.01). CONCLUSION: Persistent AD was associated with increased IL-10 and FOXP3 expression, while remission was associated with higher numbers of CD4+CD25high cells. These data suggest a link between CD4+CD25high cells that is not related to cell quantity, but rather cell function. The higher levels of IL-10 and FOXP3 may be a compensatory response to chronic inflammation, or alternately, expression of IL-10 and FOXP3 may not be completely associated with CD25high T cells.

RATIONALE: Rhinovirus (RV) illnesses in infancy are a risk factor for recurrent wheezing and asthma. To better understand this relationship, we analyzed viral etiology and RV serotypes from nasal secretions obtained throughout infancy in children who experienced recurrent respiratory illnesses. METHODS: Nasal wash samples were obtained from 26 infants during 151 moderate to severe illnesses and 80 scheduled visits between Mar/99-Mar/01, and analyzed using multiplex PCR and traditional culture techniques. Rhinoviruses were sequenced and compared to known serotypes (85% completed). RESULTS: Virus was detected in 138/151 (91%) illnesses and 29/62 (47%) well visits. During illnesses, RV was the most common virus (64%), followed by RSV (15%), adenovirus (11%), parainfluenza virus (9%), coronavirus (9%), influenza virus (8%), metapneumovirus (7%) enteroviruses (3%), and multiple viruses (28%). RV infections occurred earliest (mean age at first infection 4 months; before RSV infections in 69% of cases), were most frequent (mean 3.6 episodes compared to <1.0 episodes for other viruses) and had similar severity as other infections. At well visits, RV was most often detected (35%) compared to other viruses (50 different RV strains were found over 24 mo, and only half of the strains corresponded with known serotypes. Persistent (duration >2 wks) RV (n = 3) or other virus (n = 7) infections, and mixed RV infections (n = 3) were rare. CONCLUSIONS: RV infections occur early, pervasively, and repetitively in these high risk infants. Children with persistent symptoms most often have a series of infections rather than prolonged infection with one virus.

RATIONALE: Radioallergosorbent tests (RAST) and skin prick testing are both commonly used to test for allergic sensitization. However, information about how the results of the two tests agree is limited, particularly for preschool children. METHODS: Children enrolled in the Childhood Origins of ASThma (COAST) project underwent both a peripheral blood draw and percutaneous skin testing at age 5 years. Plasma allergen-specific IgE results were evaluated by fluoroenzyme immunoassay (Unicap® 100, Pharmacia Diagnostics) for the following antigens: D. farinae, D. pteronyssinus, Alternaria alternata, dog epithelium, cat dander, grass, birch, and ragweed. Skin prick testing was performed utilizing the Multi-Test® II method (Lincoln Diagnostics, Inc.). Agreement between the two tests was evaluated through calculation of a correlation coefficient and kappa statistic. RESULTS: The prevalence of skin prick test positivity was higher than the prevalence of RAST positivity for cat dander (23% vs. 14%, p=0.0004), dog epithelium (19% vs. 14%, p=0.08), D. farinae (20% vs. 15%, p=0.04), D. pteronyssinus (19% vs. 15%, p=0.09), and grass (12% vs. 8%, p=0.07). For other allergens, the prevalences of skin prick test and RAST positivity were comparable; agreement between the tests was substantial for Alternaria (kappa=0.71), moderate for birch (kappa=0.56) and fair for ragweed (kappa=0.39). CONCLUSIONS: In preschool children, for most allergens, positive results to skin prick testing occurred more frequently than positive RAST results, which could reflect dependence of the skin test response on immune-mediated mechanisms in addition to IgE antibody.

RATIONALE: Atopic dermatitis (AD) is the earliest clinical expression of the atopic march, a progression that may lead to the development of childhood asthma. Identification of risk factors for the expression and persistence of AD may lead to improved primary and secondary prevention strategies. METHODS: Identification of genetic and environmental factors associated with the persistence or remission of AD in children enrolled in the Childhood Origin of ASThma (COAST) cohort: a genetically at-risk population of infants based on parental histories of asthma and/or allergies. RESULTS: Of the 285 infants enrolled in the COAST study, 65 (23%) developed physician-diagnosed AD during infancy. Of these, 35 (54%) went into remission by age 5, while 30 (46%) had persistence of their disease. Remission was significantly associated with larger increases in PHA-induced IFN-γ responses from cord blood (83 vs 38 pg/mL, p=0.006), while persistence was associated with wheezing at age 5 yrs (43% vs 18%, p=0.05) and increased PHA-induced IL-10 responses from peripheral blood at age 5 yrs (1088 vs 831 pg/mL, p=0.01). Factors that were similar in both groups included breastfeeding, smoke exposure, presence of siblings, daycare attendance, pet exposure, wheezing illnesses during years 1 or 3, and PHA-induced IL-5 and IL-13 responses from either cord blood or peripheral blood. CONCLUSIONS: In this genetically at-risk population of children, persistent expression of AD into early childhood was associated with differences in immune response profiles at birth and at age 5 yrs, and the presence of wheeze at age 5 yrs.