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Abstract Populations of green turtles (Chelonia mydas), a megaherbivore that consumes seagrasses via cultivation grazing, are recovering worldwide. Information on plant‐mediated effects on herbivore foraging behavior is critical to understanding plant–herbivore interactions and sustainability of grazing as ecosystems continue to change. In a Caribbean seagrass ecosystem, we use stationary cameras and benthic surveys to evaluate the effects of seagrass morphology and leaf nitrogen content on green turtle grazing behavior. Thalassia testudinum leaf morphology has significant effects on forage intake (in milligrams of dry mass [DM] per minute) for green turtles, whereas leaf nitrogen content has no effect. Intake increases in grazed areas with shorter leaves and higher leaf biomass concentration (in milligrams of DM per cubic centimeter), indicating more efficient foraging under these conditions. Bite rate (in bites per minute) increases in grazed areas with short leaves, a result of reduced search time. Bite size (in milligrams of DM per bite) increases in grazed areas with short but dense canopies, because a turtle crops more shoots with each bite. Increased foraging efficiency and reduced search time in grazed areas with high biomass concentrations collectively maximize intake. Ingested leaves are shorter than the mean height of all available leaves in grazed areas, indicating herbivore selection for shorter leaves. Our estimate for daily intake is 86.1 g DM day−1 33‐kg turtle−1. Our study provides a novel contribution on the effects of plant‐level cues on the grazing behavior of a marine megaherbivore, and how cultivation grazing behavior optimizes the green turtle foraging strategy by maximizing foraging efficiency and intake.

A fundamental problem in biological sciences is understanding how macromolecular machines work and how the structural changes of a molecule are connected to its function. Time-resolved techniques are vital in this regard and essential for understanding the structural dynamics of biomolecules. Time-resolved small- and wide-angle X-ray solution scattering has the capability to provide a multitude of information about the kinetics and global structural changes of molecules under their physiological conditions. However, standard protocols for such time-resolved measurements often require significant amounts of sample, which frequently render time-resolved measurements impossible. A cytometry-type sheath co-flow cell, developed at the BioCARS 14-ID beamline at the Advanced Photon Source, USA, allows time-resolved pump-probe X-ray solution scattering measurements to be conducted with sample consumption reduced by more than ten times compared with standard sample cells and protocols. The comparative capabilities of the standard and co-flow experimental setups were demonstrated by studying time-resolved signals in photoactive yellow protein., (open access.)

Objective: To determine the potential longer-term effects of maternal antenatal respiratory syncytial virus (RSV) vaccination, we examined the association between cord-blood RSV-neutralizing antibodies (RSV-NA) and RSV infections in the first 2 years of life, RSV-NA at 3 years, and respiratory health to age 5 years., Methods: Two community-based Australian birth cohorts were combined. For children with at least one atopic parent, paired serum RSV-NA levels were compared in cord blood and at age 3 years. Weekly nasal swabs were collected in one cohort and during acute respiratory infections (ARI) in the other. Wheeze history up to age 5 years and physician-diagnosed asthma at 5 years was collected by parent report., Results: In 264 children, each log 10 increase of cord-blood RSV-NA level was associated with 37% decreased risk (adjusted incidence-rate-ratio [aIRR] 0.63; 95% confidence interval [CI]: 0.40-1.01) of RSV-ARI and 49% decreased risk (aIRR 0.51; 95% CI: 0.25-1.02) of RSV acute lower respiratory infections (ALRI) at 12-24 months of age. However, higher cord-blood RSV-NA was associated with increased risk of all-cause ALRI (aIRR 1.29; 95% CI: 0.99-1.69), wheeze-associated ALRI (aIRR 1.75; 95% CI: 1.08-2.82), and severe ALRI (aIRR 2.76; 95% CI: 1.63-4.70) at age 6-<12 months. Cord-blood RSV-NA was not associated with RSV-ARI in the first 6-months, RSV-NA levels at 3 years, or wheeze or asthma at 5 years., Conclusions: Higher levels of cord-blood RSV-NA did not protect against RSV infections during the first 6-months-of-life, time-to-first RSV-ARI, or wheeze or asthma in the first 5 years of life. Additional strategies to control RSV-related illness in childhood are needed., (© 2021 Wiley Periodicals LLC.)

A facility for performing serial crystallography measurements has been developed at the Australian synchrotron. This facility incorporates a purpose built high viscous injector, Lipidico, as part of the macromolecular crystallography (MX2) beamline to measure large numbers of small crystals at room temperature. The goal of this technique is to enable crystals to be grown/transferred to glass syringes to be used directly in the injector for serial crystallography data collection. The advantages of this injector include the ability to respond rapidly to changes in the flow rate without interruption of the stream. Several limitations for this high viscosity injector (HVI) exist which include a restriction on the allowed sample viscosities to >10 Pa.s. Stream stability can also potentially be an issue depending on the specific properties of the sample. A detailed protocol for how to set up samples and operate the injector for serial crystallography measurements at the Australian synchrotron is presented here. The method demonstrates preparation of the sample, including the transfer of lysozyme crystals into a high viscous media (silicone grease), and the operation of the injector for data collection at MX2.

Toe flexor strength is a pivotal biomechanical contributor for effecting balance and gait. However, there are limited reports that evaluate measurement accuracy and repeatability of this important attribute. Dynamometers are designed to measure force which can be used to derive joint torque if the perpendicular distance to the joint axis is known. However, an accurate and reliable measurement method to assess the ability of the toe flexor muscles to produce torque, is lacking. Here we describe a new device and method, designed to quantify the toe flexor torque developed at the metatarsal phalangeal joint. We evaluate measurement bias and the ability of the instrument to consistently measure what it is supposed to measure (Interclass Correlation Coefficient). Results suggest that our device is an accurate tool for measuring angle and torque with a small (0.10° and 0.07 Nm, respectively) bias. When tested for reliability and repeatability in measuring toe flexor torque (n = 10), our device showed high interclass correlation (ICC = 0.99), small bias (-1.13 Nm) and small repeatability coefficient (CR = 3.9). We suggest mean bias and CR to be reported for future measurement methods and our protocol used as standard approach to measure maximal toe flexor torque., (Copyright © 2019 IPEM. Published by Elsevier Ltd. All rights reserved.)

Repeated cycles of infection-associated lower airway inflammation drive the pathogenesis of persistent wheezing disease in children. In this study, the occurrence of acute respiratory tract illnesses (ARIs) and the nasopharyngeal microbiome (NPM) were characterized in 244 infants through their first five years of life. Through this analysis, we demonstrate that >80% of infectious events involve viral pathogens, but are accompanied by a shift in the NPM toward dominance by a small range of pathogenic bacterial genera. Unexpectedly, this change frequently precedes the detection of viral pathogens and acute symptoms. Colonization of illness-associated bacteria coupled with early allergic sensitization is associated with persistent wheeze in school-aged children, which is the hallmark of the asthma phenotype. In contrast, these bacterial genera are associated with "transient wheeze" that resolves after age 3 years in non-sensitized children. Thus, to complement early allergic sensitization, monitoring NPM composition may enable early detection and intervention in high-risk children., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: Vitamin D (25(OH)D) deficiency has been implicated as a possible risk factor for asthma development, but studies at selected time points measuring 25(OH)D levels during childhood have yielded conflicting findings. Prospective studies tracking 25(OH)D levels during the initiation phase of asthma in early childhood have not been reported., Objective: We sought to elucidate relationships between 25(OH)D levels from birth to age 10 years and susceptibility to allergic sensitization, respiratory tract infections, and asthma., Methods: Asthma-, allergy-, and respiratory tract infection-associated phenotypes (including pathogen identification) were characterized in a high-risk birth cohort. Plasma 25(OH)D concentrations were quantified at birth and at clinical follow-ups at the ages of 0.5, 1, 2, 3, 4, 5, and 10 years, and relationships with clinical outcomes were examined., Results: Cross-sectional analyses demonstrated inverse associations between 25(OH)D concentrations and the risk for concurrent sensitization at age 0.5, 2, and 3 years, and mixed-effects regression demonstrated inverse longitudinal associations of 25(OH)D levels with both sensitization and eczema. Multivariate regression modeling suggested that the number of 25(OH)D-deficient follow-ups was positively associated with risk for asthma/wheeze, eczema, and sensitization at 10 years; adjustment for sensitization (particularly by 2 years) in the asthma/wheeze models reduced 25(OH)D associations with these latter outcomes. 25(OH)D levels were also inversely associated with early nasopharyngeal colonization with Streptococcus species and age of first febrile lower respiratory illness, both of which are known asthma risk factors., Conclusion: 25(OH)D deficiency in early childhood is associated with increased risk for persistent asthma, potentially through modulating susceptibility to early allergic sensitization, upper respiratory tract colonization with bacterial pathogens, or both. These relationships are only evident if 25(OH)D status is monitored prospectively and longitudinally., (Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Radiation damage is a major limitation to synchrotron small-angle X-ray scattering analysis of biomacromolecules. Flowing the sample during exposure helps to reduce the problem, but its effectiveness in the laminar-flow regime is limited by slow flow velocity at the walls of sample cells. To overcome this limitation, the coflow method was developed, where the sample flows through the centre of its cell surrounded by a flow of matched buffer. The method permits an order-of-magnitude increase of X-ray incident flux before sample damage, improves measurement statistics and maintains low sample concentration limits. The method also efficiently handles sample volumes of a few microlitres, can increase sample throughput, is intrinsically resistant to capillary fouling by sample and is suited to static samples and size-exclusion chromatography applications. The method unlocks further potential of third-generation synchrotron beamlines to facilitate new and challenging applications in solution scattering.

Background: Although most children with asthma and rhinitis are sensitized to aeroallergens, only a minority of sensitized children are symptomatic, implying the underlying operation of efficient anti-inflammatory control mechanisms., Objective: We sought to identify endogenous control mechanisms that attenuate expression of IgE-associated responsiveness to aeroallergens in sensitized children., Methods: In 3 independent population samples we analyzed relationships between aeroallergen-specific IgE and corresponding allergen-specific IgG (sIgG) and associated immunophenotypes in atopic children and susceptibility to asthma and rhinitis, focusing on responses to house dust mite and grass., Results: Among mite-sensitized children across all populations and at different ages, house dust mite-specific IgG/IgE ratios (but not IgG4/IgE ratios) were significantly lower in children with asthma compared with ratios in those without asthma and lowest among the most severely symptomatic. This finding was mirrored by relationships between rhinitis and antibody responses to grass. Depending on age/allergen specificity, 20% to 40% of children with allergen-specific IgE (sIgE) of 0.35 kU/L or greater had negative skin test responses, and these children also expressed the high sIgG/sIgE immunophenotype. sIgG1 from these children inhibited allergen-induced IgE-dependent basophil activation in a dose-dependent fashion. Profiling of aeroallergen-specific CD4(+) TH memory responses revealed positive associations between sIgG/sIgE ratios and IL-10-dependent gene signatures and significantly higher IL-10/TH2 cytokine (protein) ratios among nonsymptomatic children., Conclusion: In addition to its role in blocking TH2 effector activation in the late-phase allergic response, IL-10 is a known IgG1 switch factor. We posit that its production during allergen-induced memory responses contributes significantly to attenuation of inflammation through promoting IgG1-mediated damping of the FcεRI-dependent acute-phase reaction. sIgG1/sIgE balance might represent a readily accessible therapeutic target for asthma/rhinitis control., (Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

The nasopharynx (NP) is a reservoir for microbes associated with acute respiratory infections (ARIs). Lung inflammation resulting from ARIs during infancy is linked to asthma development. We examined the NP microbiome during the critical first year of life in a prospective cohort of 234 children, capturing both the viral and bacterial communities and documenting all incidents of ARIs. Most infants were initially colonized with Staphylococcus or Corynebacterium before stable colonization with Alloiococcus or Moraxella. Transient incursions of Streptococcus, Moraxella, or Haemophilus marked virus-associated ARIs. Our data identify the NP microbiome as a determinant for infection spread to the lower airways, severity of accompanying inflammatory symptoms, and risk for future asthma development. Early asymptomatic colonization with Streptococcus was a strong asthma predictor, and antibiotic usage disrupted asymptomatic colonization patterns. In the absence of effective anti-viral therapies, targeting pathogenic bacteria within the NP microbiome could represent a prophylactic approach to asthma., (Copyright © 2015 Elsevier Inc. All rights reserved.)

It is uncertain whether the vitamin D status of pregnant women influences bone mass of their children. Cohort studies have yielded conflicting results; none have examined offspring at skeletal maturity. This longitudinal, prospective study investigated the association between maternal vitamin D status and peak bone mass of offspring in 341 mother and offspring pairs in the Western Australian Pregnancy Cohort (Raine) Study. Maternal serum samples collected at 18 weeks gestation were assayed for 25-hydroxyvitamin D (25OHD). Outcomes were total body bone mineral content (BMC) and bone mineral density (BMD) measured by dual-energy X-ray absorptiometry in offspring at 20 years of age. The mean (± SD) maternal serum 25OHD concentration was 57.2 ± 19.2 nmol/L; 132 women (38.7%) were vitamin D-deficient (25OHD <50 nmol/L). After adjustment for season of sample collection, maternal factors, and offspring factors (sex, birth weight, and age, height, lean mass, and fat mass at 20 years), maternal 25OHD concentration was positively associated with total body BMC and BMD in offspring, with a mean difference of 19.2 (95% confidence interval [CI], 5.6-32.7) g for BMC and 4.6 (95% CI, 0.1-9.1) mg/cm(2) for BMD per 10.0 nmol/L of maternal 25OHD. Maternal vitamin D deficiency was associated with 2.7% lower total body BMC (mean ± SE) (2846 ± 20 versus 2924 ± 16 g, p = 0.004) and 1.7% lower total body BMD (1053 ± 7 versus 1071 ± 5 mg/cm(2) , p = 0.043) in the offspring. We conclude that vitamin D deficiency in pregnant women is associated with lower peak bone mass in their children. This may increase fracture risk in the offspring in later life., (© 2014 American Society for Bone and Mineral Research.)

Epidemiologic associations between viral lower respiratory infections (LRIs) and asthma in later childhood are well known. However, the question of whether such infections cause asthma or unmask asthma in a susceptible host has still not been settled. Most early evidence centered on the role of the respiratory syncytial virus; however, recent studies highlight a potential role for human rhinovirus as a risk factor for asthma. The links between early-life viral LRI and subsequent asthma are generally via wheeze; however, the presence of wheeze does not give any information about why the child is wheezing. Wheeze in early life is, at best, a fuzzy phenotype and not specific for subsequent asthma. The risk of asthma after viral LRI is increased in the presence of allergic sensitization in early life and if the infection is more severe. Atopy-associated mechanisms also appear to be involved in viral-induced acute exacerbations of asthma, especially in prolonging symptomatology after the virus has been cleared from the lungs. Breaking the nexus between viral respiratory infections and asthma may be possible with interventions designed to inhibit atopy-related effectors mechanisms from participating in the host response to respiratory viral infections., (Copyright (c) 2010 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.)

Background: Atopy and asthma are commonly initiated during early life, and there is increasing interest in the development of preventive treatments for at-risk children. However, effective methods for assessing the level of risk in individual children are lacking., Objective: We sought to identify clinical and laboratory biomarkers in 2-year-olds that are predictive of the risk for persistent atopy and wheeze at age 5 years., Methods: We prospectively studied 198 atopic family history-positive children to age 5 years. Clinical and laboratory assessments related to asthma history and atopy status were undertaken annually; episodes of acute respiratory illness were assessed and classified throughout and graded by severity., Results: Aeroallergen-specific IgE titers cycled continuously within the low range in nonatopic subjects. Atopic subjects displayed similar cycling in infancy but eventually locked into a stable pattern of upwardly trending antibody production and T(H)2-polarized cellular immunity. The latter was associated with stable expression of IL-4 receptor in allergen-specific T(H)2 memory responses, which was absent from responses during infancy. Risk for persistent wheeze was strongly linked to early sensitization and in turn to early infection. Integration of these data by means of logistic regression revealed that attaining mite-specific IgE titers of greater than 0.20 kU/L by age 2 years was associated with a 12.7% risk of persistent wheeze, increasing progressively to an 87.2% risk with increasing numbers of severe lower respiratory tract illnesses experienced., Conclusion: The risk for development of persistent wheeze in children can be quantified by means of integration of measures related to early sensitization and early infections. Follow-up studies along similar lines in larger unselected populations to refine this approach are warranted.

Background: It has been proposed that immune dysfunction during early childhood plays an important role in asthma pathogenesis. However, it is not known specifically whether changes in dendritic cells (DCs) during infancy antedate the development of respiratory tract infections, asthma, and related clinical phenotypes., Objectives: We sought to assess the association between the level of blood DCs during the first year and the subsequent development of respiratory tract infections, wheezing, and allergic sensitization., Methods: A community-based cohort of children with a family history of atopy was followed to age 5 years. Children were monitored intensively for respiratory tract infections. History of wheeze and asthma was collected annually, atopy was documented at 5 years, and flow cytometry was used to identify DC subsets in blood samples collected when children were well., Results: Levels of plasmacytoid DCs (pDCs) during infancy were inversely correlated with symptoms of lower respiratory tract infections, parent-reported wheezing, and the cumulative rate of physician-diagnosed asthma up to age 5 years. These relationships were independent of atopy, as determined by allergy skin test results and total and specific IgE levels. In contrast, levels of myeloid DCs were not associated with respiratory tract infections, asthma, or wheezing but were associated with total IgE levels at age 5 years., Conclusion: In children with a family history of atopy, relative deficiency of circulating pDCs during infancy appears to be a risk factor for more frequent and more severe respiratory tract infections, wheezing, and a diagnosis of asthma. Infants with higher numbers of pDCs are protected against these outcomes.

Rationale: Respiratory infections in early life are associated with risk for wheezing bronchiolitis, especially in children at high risk of atopy. The underlying mechanisms are unknown, but are suspected to involve imbalance(s) in host defense responses against pathogens stemming from functional immaturity of the immune system in this age group., Objectives: To assess the contribution of eosinophil-trophic IL-5, and the potent antiinflammatory cytokine IL-10, to risk for infection in early life., Measurements and Main Results: We prospectively monitored a cohort of 198 high-risk children to age 5 years, recording every acute respiratory infection episode and classifying them by severity. We measured cord blood T-cell capacity to produce IL-10 and IL-5, and related these functions to subsequent infection history. IL-10 and IL-5 were associated, respectively, with resistance versus susceptibility to infections. The greatest contrasting effects of these two cytokines were seen when they were considered in combination by generating IL-10/IL-5 response ratios for each subject. The low IL-10/high IL-5 T-cell response phenotype was strongly associated with susceptibility to all grades of acute respiratory infection, relative to the more resistant high IL-10/low IL-5 phenotype., Conclusions: Excessive production of IL-5 by T cells at birth is associated with heightened risk for subsequent severe respiratory infections, and this risk is attenuated by concomitant IL-10 production. The underlying mechanisms may involve IL-10-mediated feedback inhibition of IL-5-dependent eosinophil-induced inflammation, which is a common feature of host antiviral responses in early life.

Atopy is characterized by eosinophilic inflammation associated with recruitment of eosinophil/basophil (Eo/B) progenitors. We have previously shown that Eo/B progenitor phenotypes are altered in cord blood (CB) in infants at high risk of atopy/asthma, and respond to maternal dietary intervention during pregnancy. As respiratory tract viral infections have been shown to induce wheeze in infancy, we investigated the relationship between CB progenitor function and phenotype and acute respiratory illness (ARI), specifically wheeze and fever. CB from 39 high-risk infants was studied by flow cytometry for CD34(+) progenitor phenotype and by ex vivo Eo/B-colony forming unit (CFU) responses to cytokine stimulation in relation to ARI in the first year of life. A consistent relationship was observed between increased numbers of granulocyte/macrophage (GM)-colony-stimulating factor (CSF)- and IL-3-responsive Eo/B-CFU in CB and the frequency/characteristics of ARI during infancy. Comparable associations were found between ARI and CB IL-3R(+) and GM-CSFR(+)CD34(+) cell numbers. Conversely, a reciprocal decrease in the proportion of CB IL-5R(+) cells was found in relation to the clinical outcomes. The elevation of IL-3/GM-CSF-responsive Eo/B progenitors in high-risk infants in relation to ARI outcomes suggests a mechanism for the increased severity of inflammatory responses in these subjects following viral infection.

The aim of this study was to investigate the relationship between air pollution and respiratory symptoms in young children. A total of 263 children at high risk of developing asthma or atopy were recruited antenatally and all respiratory symptoms experienced by the children were recorded by their parents for five years. Daily pollutant concentrations and meteorological data (ambient temperature and humidity) were collected from network monitoring sites. Logistic regression models investigating relationships between individual air pollutants and respiratory symptoms showed significant associations between Ozone (O3) (1 h and 8 h) concentrations and raised body temperature (lag 0); Carbon monoxide (CO) (8 h) and wheeze/rattle and runny/blocked nose (lag 5 and additive exposure over 5 days); Nitrogen dioxide (NO2) (24 h) concentrations and cough (lag 0 and additive exposure over 5 days) and PM2.5 and visibility (BSP) (1 h) with cough (lag 0). These associations were observed even though air pollutant concentrations were below national standards throughout the study period.

Background: The timing of allergen sensitization is controversial, with conflicting evidence suggesting transplacental priming versus exclusively postnatal priming. Resolution of this question is important in relation to rational design of allergy prevention strategies, particularly the issue of allergen avoidance during pregnancy., Objective: To elucidate the kinetics of sensitization in high-risk children during their first 2 years of life., Methods: We prospectively studied house dust mite (HDM)-specific IgE and IgG(4) antibody production and associated T-cell immunity in a cohort of 200 high-risk infants. Parallel antibody studies tracked responses against a broader panel of inhalant and dietary allergens including peanut., Results: HDM-induced T(H)2 responses in PBMC from 6 months onward, particularly IL-4 and IL-5, correlated increasingly strongly with sensitization outcomes at 2 years, and a contrasting negative relationship was observed with IFN-gamma response capacity. HDM-induced T-cell responses in cord blood, although common, were unrelated to subsequent sensitization. Transient HDM-IgE (and IgG(4)) production frequently peaked at 6 or 12 months before returning to baseline, which suggests the onset of protective tolerance. This finding contrasted with progressively increasing HDM-IgE titers in children sensitized by 2 years of age. Comparably contrasting patterns were observed in peanut-specific responses in sensitized versus nonsensitized children., Conclusion: Priming of T(H)2 responses associated with persistent HDM-IgE production occurs entirely postnatally, as HDM reactivity in cord blood seems nonspecific and is unrelated to subsequent development of allergen-specific T(H)2 memory or IgE., Clinical Implications: These findings question the scientific basis for existing recommendations for allergen avoidance by high-risk women during pregnancy.

Background: The relationship between atopy and bronchial allergy in young children is not completely understood., Objective: To examine the association between response to bronchial allergen challenge, immune markers of atopy and other clinical characteristics in 5- to 6-year-old children., Methods: Children with positive skin test (SPT) to aeroallergen, together with a proportion of SPT negative children (as controls), were recruited from a birth cohort of 198 children at high risk of developing atopic disease and underwent allergen challenge., Results: Thirty-seven children (26 atopic and 11 SPT negative), median age 74.5 months, were challenged: 31 with house dust mite and six with grass allergen. Only atopic children responded to challenge: n = 12/26 (46%). Wheal size [odds ratio (OR) 2.5 (1.2-5.3), P = 0.01], allergen-specific immunoglobulin E (IgE) [OR 3.4 (1.23-9.61), P = 0.02], total IgE [OR 8.6 (1.1-68.7), P = 0.04], current wheeze [OR 12 (1.7-81.7), P = 0.006] and persistent eczema [OR 11.0 (1.7-68.3), P = 0.006] emerged as the strongest independent predictors of response to allergen challenge. Prediction of response to allergen challenge was significantly improved when immune markers of atopy, and in particular wheal size, were combined with clinical characteristics., Conclusion: The relationship between atopy and bronchial allergy is quantitative at this age. There may be potential to create more powerful indicators of the presence of respiratory allergy in young children when immunological markers of atopy are considered quantitatively and when combined with clinical history of coexistent allergic disease.

Background: Gene-environment interactions play central roles in controlling postnatal maturation of immune function, but their effects on infant vaccine responses are unknown. Genetic variants associated with atopy and the environmental factor of exposure to parental smoking (PS) of tobacco independently alter immune responses., Objective: We sought to investigate the hypothesis that genetic variants associated with atopy and their interaction with PS influence infant vaccine responsiveness., Methods: In 200 infants with parental atopic history, relationships were sought between polymorphisms in the IL-4, IL-4 receptor alpha (IL-4Ralpha), and IL-13 genes; PS; and immune responses to diphtheria/tetanus vaccination., Results: Analyses stratified by PS unmasked negative associations between atopic alleles of these genes and vaccine outcomes. The most consistent involved the IL-4Ralpha 551 QR/QQ genotypes, which were associated with reduced IgG levels (P = .02) and T-cell responses (IFN-gamma, P = .002; IL-10, P = .01; 1L-13, P = .01; IL-5, P = .06) to tetanus toxoid and parallel reductions in polyclonal T-cell responses and innate immune responses in PS-exposed infants., Conclusion: PS potentiates suppressive effects of variants in immune response genes in children. These effects are not observed in the absence of this exposure. Ultimately, this finding might have implications for infant vaccination in countries with high smoking rates. It might also have broader implications in relation to environmental toxicology because it demonstrates specific mechanisms through which the developing immune system might be differentially sensitive to low-level toxicant exposures., Clinical Implications: PS interacts with genes associated with atopy to impair vaccine responses. These interactions might have vaccine design and public health implications.

Interferon-gamma (IFNgamma) gene expression is tightly regulated in early life, and exaggerated negative control of IFNgamma production in CD4(+) T cells has been associated with risk for subsequent development of atopy. Recent studies have demonstrated hypermethylation of CpG sites in the IFNgamma promoter in neonates, a mechanism which in mice leads to strong suppression of IFNgamma gene transcription. In the present study, the methylation status of six CpG sites in the proximal promoter of the human IFNgamma gene was determined by bisulphite sequencing. Cell populations studied were Th1 or Th2 polarized cell lines derived from neonatal and adult CD4(+)/CD45RA(+) T cells, CD4(+) and CD8(+) naive T cells from cord blood of children followed to outcome age 2 for assessment of atopy status, and CD4(+) and CD8(+) naive T cells from 6 yr old and adult atopics and controls. We demonstrate that in vitro differentiation of CD4(+) T cells down the Th1 pathway (but not the Th2 pathway) is accompanied by progressive demethylation of CpG sites in the IFNgamma promoter, which is most marked in neonatal cells. Atopy development by age 2 was not associated with variations in methylation patterns in cord blood T cells. However, IFNgamma promoter methylation was reduced in CD8(+) T cells from atopic children in the age range in which hyperproduction of IFNgamma as recently been identified as a common feature of the atopic phenotype. The findings demonstrate the potency of IFNgamma promoter methylation as a mechanism for control of human IFNgamma gene expression, particularly during early life. Differential regulation of IFNgamma promoter methylation in T cells may be an important contributory factor in atopy development in childhood, and this possibility warrants further detailed investigation.

Purpose: Continuing professional development is an integral component of modern medical practice, yet traditional educational methods are impractical for many Primary Care Physicians. Web-based programs may fulfill the requirements of busy practitioners who have difficulty attending formal education sessions., Methods: We piloted the use of a learning management system to deliver asthma education materials to Primary Care Physicians in both Australia and Italy in their native languages. Each group of Physicians accessed an education module which contained content pages, self-tests, a quiz and a survey. Details of how the Physicians used the system, their preferences and performance on the assessment were monitored., Results: The learning management system was well received by both Italian and Australian Physicians. Thirty-eight (18 Australian, 20 Italian) Physicians used the system. Participants visited an average of 8.8 pages, with a mean time per hit of 2.9 min. Formative assessment was undertaken by 63.2% and summative assessment by 68.4% of participants. There were no substantial differences in performance between Physicians from both countries. Italian physicians tended to use the system after hours whereas Australian Physicians appear to do so between patient visits., Conclusions: Simple web-based systems are suitable for delivering educational materials to Primary Care Physicians in a manner likely to be used.

The capacity of the immune system in infants to develop stable T-cell memory in response to vaccination is attenuated, and the mechanism(s) underlying this developmental deficiency in humans is poorly understood. The present study focuses on the capacity for expression of in vitro recall responses to tetanus and diphtheria antigens in lymphocytes from 12-month-old infants vaccinated during the first 6 months of life. We demonstrate that supplementation of infant lymphocytes with "matured" dendritic cells (DC) cultured from autologous CD14+ precursors unmasks previously covert cellular immunity in the form of Th2-skewed cytokine production. Supplementation of adult lymphocytes with comparable prematured autologous DC also boosted vaccine-specific T-cell memory expression, but in contrast to the case for the infants, these cytokine responses were heavily Th1 skewed. Compared to adults, infants had significantly fewer circulating myeloid DC (P < 0.0001) and plasmacytoid DC (P < 0.0001) as a proportion of peripheral blood mononuclear cells. These findings suggest that deficiencies in the numbers of antigen-presenting cells and their functional competence at 12 months of age limit the capacity to express effector memory responses and are potentially a key factor in reduced vaccine responsiveness in infants.

Background: High genetic risk (HR) of atopy among unstratified populations of infants is associated with attenuated IFN-gamma responses. However, the role of IFN-gamma in progression from HR status to active disease is less clear., Objective: To identify immune function markers in neonates with HR that are associated with positive atopic outcomes at 2 years., Methods: Cord blood mononuclear cells (CBMCs) were collected from 175 children with HR and cryopreserved. The children were assessed for atopy by skin prick at 0.5 and 2 years. CBMCs were thawed and stimulated with allergens and mitogens PHA and staphylococcal enterotoxin B (SEB), and cytokine responses were determined., Results: No correlations were observed between allergen-specific CBMC responses and atopic outcomes. In contrast, sensitization was strongly associated with polyclonal IFN-gamma responses to both PHA (P=.002) and SEB (P=.005), and also with SEB-induced IL-5 (P =.05), IL-10 (P =.02), and IL-13 (P =.01). Logistic regression analysis identified elevated PHA-induced IFN-gamma and SEB-induced IL-13 responses as the strongest independent predictors of atopy development. Cell separation studies confirmed CD8+ T cells as the source of approximately 90% of IFN-gamma production., Conclusions: IFN-gamma produced by CD8+ T cells may synergize with T(H)2 cytokines in driving atopy development in children with HR.

Background: Mononuclear cells from children with active atopic dermatitis (AD) have been reported to be hyper-responsive to certain microbial stimuli, in particular staphylococcal enterotoxin B (SEB). However, it is not known whether this responsiveness is acquired during disease development, or is inherent. We investigated this question in a cohort of children at high risk of atopy followed prospectively from birth to age 3 years. We asked whether their cord blood mononuclear cell (CBMC) cytokine responses to SEB, to an unrelated microbial stimulus purified protein derivative (PPD), or to common allergens, were predictive of risk for subsequent AD development during infancy., Methods: Children at high risk of developing atopy were randomly selected from an ongoing prospective cohort. Cord blood was collected at birth. The children were seen at 6 months, 1, 2 and 3 years and examined for the development of AD. IFN-gamma, IL-5, IL-10 and IL-13 production by CBMC cultured in the presence of SEB, PPD, PHA, house dust mite (HDM) allergen, ovalbumin (OVA) and cat allergen was determined., Results: SEB-induced IL-5 production by CBMC was elevated in children who developed AD at 6 months (P = 0.01) and 2 years (P = 0.009). PPD-induced IL-5 responses were also elevated in CBMC from children who developed AD at 6 months, 2 years and 3 years (P = 0.05, P = 0.06 and P = 0.06, respectively), as were PPD-induced IL-10 responses (P = 0.05 at 1 years, P = 0.007 at 2 years, P = 0.003 at 3 years) and corresponding IFN-gamma responses (P = 0.05 at 6 months, P = 0.003 at 2 years, P = 0.0004 at 3 years). Increased IL-10 responses to HDM allergen were also observed throughout the observation period in CBMC from children who developed AD., Conclusion: Children who develop infantile AD appear to have a predisposition to respond to SEB in a Th2-dominant manner involving selective stimulation of IL-5 production. The increased IL-10 and IFN-gamma induced in response to PPD by children with AD may point to additional intrinsic differences in responses to microbial stimuli between those at high vs. those at low risk for AD, which merit more detailed investigations.

The normally acidic mucosal surface pH of 6.24 +/- 0.02(30) in rat proximal jejunum in vivo is effectively neutralised by 30 min exposure to heat-stable Escherichia coli (STa) enterotoxin (14 micrograms/ml) to 6.80 +/- 0.07 (n = 5) or to a forskolin/theophylline combination (1 mM:20 mM) to 7.10 +/- 0.07(7) while perfusion with Krebs-phosphate buffer alone without glucose left the mucosal surface pH unchanged at a pH of 6.21 +/- 0.02(9). Forskolin alone had no effect, and 20 mM theophylline moderately elevated the surface pH to 6.52 +/- 0.03(5). Theophylline, forskolin and their combination all elevated cAMP levels per mg tissue DNA above control values while STa enterotoxin was without effect. In contrast, all agents elevated cGMP levels per mg tissue DNA above control levels. These findings indicate that surface pH is only moderately affected by changes in cAMP levels and is affected to a much greater extent by altered cGMP levels.