Maria Valkonen

Bacterial Exposures and Associations with Atopy and Asthma in Children

Background The increase in prevalence of asthma and atopic diseases in Western countries has been linked to aspects of microbial exposure patterns of people. It remains unclear which microbial aspects contribute to the protective farm effect. Objective The objective of this study was to identify bacterial groups associated with prevalence of asthma and atopy, and to quantify indoor exposure to some of these bacterial groups. Methods A DNA fingerprinting technique, denaturing gradient gel electrophoresis (DGGE), was applied to mattress dust samples of farm children and control children in the context of the GABRIEL Advanced study. Associations between signals in DGGE and atopy, asthma and other allergic health outcomes were analyzed. Quantitative DNA based assays (qPCR) for four bacterial groups were applied on the dust samples to seek quantitative confirmation of associations indicated in DNA fingerprinting. Results Several statistically significant associations between individual bacterial signals and also bacterial diversity in DGGE and health outcomes in children were observed. The majority of these associations showed inverse relationships with atopy, less so with asthma. Also, in a subsequent confirmation study using a quantitative method (qPCR), higher mattress levels of specifically targeted bacterial groups - Mycobacterium spp., Bifidobacteriaceae spp. and two different clusters of Clostridium spp. - were associated with a lower prevalence of atopy. Conclusion DNA fingerprinting proved useful in identifying bacterial signals that were associated with atopy in particular. These findings were quantitatively confirmed for selected bacterial groups with a second method. High correlations between the different bacterial exposures impede a clear attribution of protective effects to one specific bacterial group. More diverse bacterial flora in mattress dust may link to microbial exposure patterns that protect against development of atopic diseases.

Determination of bacterial load in house dust using qPCR, chemical markers and culture

In this study, we developed two novel qPCR-assays for the detection of bacteria in house dust; one that determines the total bacterial amount and another that detects Gram-positive and Gram-negative bacteria separately. The methods were tested in silico and in vitro with microbial strains and vacuum cleaner dust samples, and validated in relation to culture and chemical marker analysis. We also compared the results of these three types of methods (qPCR, culture and chemical marker analysis) in 211 house dust samples from farming and non-farming environments. Microbial concentrations determined by the new qPCR assays (median 7.2 x 10(5) cell equivalents mg(-1)) were about two orders of magnitude higher than concentrations obtained by culture (median 6.7 x 10(3) cfu mg(-1)). The median concentration of muramic acid was 25.67 ng mg(-1) and that of 3-hydroxy fatty acids, expressed as LPS(10-16) was 26.14 pg mg(-1). Correlations between qPCR and chemical markers were moderate, while correlations between culture and qPCR and chemical markers were low to moderate. All the methods used in this study showed that the microbial concentrations are statistically significantly higher (p < 0.001, Mann-Whitney) in farming than non-farming environments.As a conclusion, all tested methods can be used for determining the bacterial load in dust samples, but none of the methods was superior to the others. The results obtained with these methods represent different aspects of bacterial exposure and therefore the results are not expected to be identical with each other.

Endotoxin, extracellular polysaccharides, and β(1-3)-glucan concentrations in dust and their determinants in four European birth cohorts: results from the HITEA project.

UNLABELLED Early-life exposure to microbial agents may play a protective role in asthma and allergies development. Geographical differences in the prevalence of these diseases exist, but the differences in early-life indoor microbial agent levels and their determinants have been hardly studied. We aimed to describe the early-life levels of endotoxin, extracellular polysaccharides (EPS), and β(1-3)-glucans in living room dust of four geographically spread European birth cohorts (LISA in Germany, PIAMA in the Netherlands, INMA in Spain, and LUKAS2 in Finland) and to assess their determinants. A total of 1572 dust samples from living rooms of participants were analyzed for endotoxin, Penicillium/Aspergillus EPS, and β(1-3)-glucans. Information on potential determinants was obtained through questionnaires. Concentrations of endotoxin, EPS, and β(1-3)-glucans were different across cohorts. Concentrations of endotoxin and EPS were respectively lower and higher in INMA than in other cohorts, while glucans were higher in LUKAS2. Season of sampling, dog ownership, dampness, and the number of people living at home were significantly associated with concentrations of at least one microbial agent, with heterogeneity of effect estimates of the determinants across cohorts. In conclusion, both early-life microbial exposure levels and exposure determinants differ across cohorts derived from diverse European countries. PRACTICAL IMPLICATIONS This study adds evidence of variability in the levels of indoor endotoxin, extracellular polysaccharide, and β(1-3)-glucans across four geographically spread European regions. Furthermore, we observed heterogeneity across regions in the effect of exposure determinants. We hypothesize that the variations observed in our study may play a role in the differences in asthma and allergies prevalences across countries.

Predictors of microbial agents in dust and respiratory health in the Ecrhs

BackgroundDampness and mould exposure have been repeatedly associated with respiratory health. However, less is known about the specific agents provoking or arresting health effects in adult populations. We aimed to assess predictors of microbial agents in mattress dust throughout Europe and to investigate associations between microbial exposures, home characteristics and respiratory health.MethodsSeven different fungal and bacterial parameters were assessed in mattress dust from 956 adult ECRHS II participants in addition to interview based home characteristics. Associations between microbial parameters and the asthma score and lung function were examined using mixed negative binomial regression and linear mixed models, respectively.ResultsIndoor dampness and pet keeping were significant predictors for higher microbial agent concentrations in mattress dust. Current mould and condensation in the bedroom were significantly associated with lung function decline and current mould at home was positively associated with the asthma score. Higher concentrations of muramic acid were associated with higher mean ratios of the asthma score (aMR 1.37, 95%CI 1.17-1.61). There was no evidence for any association between fungal and bacterial components and lung function.ConclusionIndoor dampness was associated with microbial levels in mattress dust which in turn was positively associated with asthma symptoms.

Microbial characteristics in homes of asthmatic and non-asthmatic adults in the ECRHS cohort

Microbial exposures in homes of asthmatic adults have been rarely investigated; specificities and implications for respiratory health are not well understood. The objectives of this study were to investigate associations of microbial levels with asthma status, asthma symptoms, bronchial hyperresponsiveness (BHR), and atopy. Mattress dust samples of 199 asthmatics and 198 control subjects from 7 European countries participating in the European Community Respiratory Health Survey II study were analyzed for fungal and bacterial cell wall components and individual taxa. We observed trends for protective associations of higher levels of mostly bacterial markers. Increased levels of muramic acid, a cell wall component predominant in Gram-positive bacteria, tended to be inversely associated with asthma (ORs for different quartiles: II 0.71 [0.39-1.30], III 0.44 [0.23-0.82], and IV 0.60 [0.31-1.18] P for trend .07) and with asthma score (P for trend .06) and with atopy (P for trend .02). These associations were more pronounced in northern Europe. This study among adults across Europe supports a potential protective effect of Gram-positive bacteria in mattress dust and points out that this may be more pronounced in areas where microbial exposure levels are generally lower.

Indoor bacteria and asthma in adults : a multicentre case-control study within ECRHS II

Both protective and adverse effects of indoor microbial exposure on asthma have been reported, but mostly in children. To date, no study in adults has used non-targeted methods for detection of indoor bacteria followed by quantitative confirmation. A cross-sectional study of 198 asthmatic and 199 controls was conducted within the European Community Respiratory Health Survey (ECRHS) II. DNA was extracted from mattress dust for bacterial analysis using denaturing gradient gel electrophoresis (DGGE). Selected bands were sequenced and associations with asthma confirmed with four quantitative PCR (qPCR) assays. 15 out of 37 bands detected with DGGE, which had at least a suggestive association (p<0.25) with asthma, were sequenced. Of the four targeted qPCRs, Clostridium cluster XI confirmed the protective association with asthma. The association was dose dependent (aOR 0.43 (95% CI 0.22–0.84) for the fourth versus first quartile, p for trend 0.009) and independent of other microbial markers. Few significant associations were observed for the three other qPCRs used. In this large international study, the level of Clostridium cluster XI was independently associated with a lower risk of prevalent asthma. Results suggest the importance of environmental bacteria also in adult asthma, but need to be confirmed in future studies. Microbial exposures at home may also protect from asthma in adults, not only in children http://ow.ly/7cnI30hzSox

Indoor microbiota in severely moisture damaged homes and the impact of interventions

BackgroundThe limited understanding of microbial characteristics in moisture-damaged buildings impedes efforts to clarify which adverse health effects in the occupants are associated with the damage and to develop effective building intervention strategies. The objectives of this current study were (i) to characterize fungal and bacterial microbiota in house dust of severely moisture-damaged residences, (ii) to identify microbial taxa associated with moisture damage renovations, and (iii) to test whether the associations between the identified taxa and moisture damage are replicable in another cohort of homes. We applied bacterial 16S rRNA gene and fungal ITS amplicon sequencing complemented with quantitative PCR and chemical-analytical approaches to samples of house dust, and also performed traditional cultivation of bacteria and fungi from building material samples.ResultsActive microbial growth on building materials had significant though small influence on the house dust bacterial and fungal communities. Moisture damage interventions—including actual renovation of damaged homes and cases where families moved to another home—had only a subtle effect on bacterial community structure, seen as shifts in abundance weighted bacterial profiles after intervention. While bacterial and fungal species richness were reduced in homes that were renovated, they were not reduced for families that moved houses. Using different discriminant analysis tools, we were able identify taxa that were significantly reduced in relative abundance during renovation of moisture damage. For bacteria, the majority of candidates belonged to different families within the Actinomycetales order. Results for fungi were overall less consistent. A replication study in approximately 400 homes highlighted some of the identified taxa, confirming associations with observations of moisture damage and mold.ConclusionsThe present study is one of the first studies to analyze changes in microbiota due to moisture damage interventions using high-throughput sequencing. Our results suggest that effects of moisture damage and moisture damage interventions may appear as changes in the abundance of individual, less common, and especially bacterial taxa, rather than in overall community structure.