Occupational exposures to pesticides and other chemicals: a New Zealand motor neuron disease case-control study.

OBJECTIVES: To assess associations between occupational exposures to pesticides and other chemicals and motor neuron disease (MND). METHODS: A population-based case-control study that included 319 MND cases (64% male/36% female) recruited through the New Zealand MND Association complemented with hospital discharge data, and 604 controls identified from the Electoral Roll. For each job held, a questionnaire collected information on 11 exposure categories (dust, fibres, tobacco smoke, fumes, gas, fumigants, oils/solvents, acids/alkalis, pesticides, other chemicals and animals/animal products). ORs were estimated using logistic regression adjusting for age, sex, ethnicity, socioeconomic status, education, smoking, alcohol consumption, physical activities, head/spine injury and other occupational exposures. RESULTS: Two exposure categories were associated with increased MND risks: pesticides (OR 1.70, 95% CI 1.17 to 2.48) and fumigants (OR 3.98, 95% CI 1.81 to 8.76), with risks increasing with longer exposure duration (p<0.01). Associations were also observed for: methyl bromide (OR 5.28, 95% CI 1.63 to 17.15), organochlorine insecticides (OR 3.28, 95% CI 1.18 to 9.07), organophosphate insecticides (OR 3.11, 95% CI 1.40 to 6.94), pyrethroid insecticides (OR 6.38, 95% CI 1.13 to 35.96), inorganic (copper) fungicides (OR 4.66, 95% CI 1.53 to 14.19), petrol/diesel fuel (OR 2.24, 95% CI 1.27 to 3.93) and unspecified solvents (OR 1.91, 95% CI 1.22 to 2.99). In women, exposure to textile fibres (OR 2.49, 95% CI 1.13 to 5.50), disinfectants (OR 9.66, 95% CI 1.29 to 72.44) and cleaning products (OR 3.53, 95% CI 1.64 to 7.59) were also associated with MND; this was not observed in men (OR 0.80, 95% CI 0.44 to 1.48; OR 0.72, 95% CI 0.29 to 1.84; OR 0.57, 95% CI 0.21 to 1.56, respectively). CONCLUSIONS: This study adds to the evidence that pesticides, especially insecticides, fungicides, and fumigants, are risk factors for MND.

Problematic severe asthma in children treated at high altitude: tapering the dose while improving control.

BACKGROUND: Multidisciplinary treatment at high altitude is a possible treatment option for problematic severe asthma (PSA) in children. This management can result in the tapering of inhaled corticosteroids. AIM: Our aim was to analyze the effect of multidisciplinary treatment at high altitude, notably the ability to taper corticosteroids. To get an insight into possible factors influencing tapering, we examined whether demographic variables, disease control and quality of life at treatment entrance could predict the tapering of corticosteroids. METHODS: This prospective open-phase cohort study analyzed the data of 43 children aged 8-17 years referred to a specialized high altitude treatment centre. Lung function (FEV1, FEV1/VC), inflammation (FeNO), medication level, asthma control (ACT) and quality of life [PAQLQ(S)] were evaluated on admission and at discharge. RESULTS: Thirty-two (74%) children fulfilled PSA criteria. Three (7%) children used daily oral steroids. After 72 ± 30 (mean ± SD) days of treatment, the mean dosage of inhaled corticosteroids (ICS) could be significantly reduced from 1315 µg ± 666 budesonide equivalent to 1132 µg ± 514. Oral steroid maintenance therapy could be stopped in all patients. FeNO, asthma control and quality of life improved (p < 0.001) from admission to discharge; FEV1 was in the normal range on both occasions. Apart from ICS levels at entrance, multiple regression analyses did not show any associated factor predicting the reduction of ICS dosage during treatment. CONCLUSION: The results indicate that high altitude treatment may be a treatment option for children with PSA, but it is not possible to predict ICS tapering off from health status variables at treatment entrance.

TBX4 mutations (small patella syndrome) are associated with childhood-onset pulmonary arterial hypertension.

BACKGROUND: Childhood-onset pulmonary arterial hypertension (PAH) is rare and differs from adult-onset disease in clinical presentation, with often unexplained mental retardation and dysmorphic features (MR/DF). Mutations in the major PAH gene, BMPR2, were reported to cause PAH in only 10-16% of childhood-onset patients. We aimed to identify more genes associated with childhood-onset PAH. METHODS: We studied 20 consecutive cases with idiopathic or heritable PAH. In patients with accompanying MR/DF (n=6) array-comparative genomic hybridisation analysis was performed, with the aim of finding common deletion regions containing candidate genes for PAH. Three patients had overlapping deletions of 17q23.2. TBX2 and TBX4 were selected from this area as candidate genes and sequenced in all 20 children. After identifying TBX4 mutations in these children, we subsequently sequenced TBX4 in a cohort of 49 adults with PAH. Because TBX4 mutations are known to cause small patella syndrome (SPS), all patients with newly detected TBX4 mutations were screened for features of SPS. We also screened a third cohort of 23 patients with SPS for PAH. RESULTS: TBX4 mutations (n=3) or TBX4-containing deletions (n=3) were detected in 6 out of 20 children with PAH (30%). All living patients and two parents with TBX4 mutations appeared to have previously unrecognised SPS. In the adult PAH-cohort, one TBX4 mutation (2%) was detected. Screening in the cohort of (predominantly adult) SPS patients revealed no PAH. CONCLUSIONS: These data indicate that TBX4 mutations are associated with childhood-onset PAH, but that the prevalence of PAH in adult TBX4 mutation carriers is low.

Risk factors for symptoms of childhood asthma, allergic rhinoconjunctivitis and eczema in the Pacific: an ISAAC Phase III study.

BACKGROUND AND METHODS: Phase III of the International Study of Asthma and Allergies in Childhood (ISAAC) was conducted in eight Pacific countries, five of which (Samoa, Fiji, Tokelau, French Polynesia and New Caledonia) collected environmental questionnaire (EQ) data. We report the findings of the EQ analyses below. RESULTS: The major factors associated with current wheeze were regular margarine consumption (prevalence odds ratio [POR] 1.19, 95%CI 1.01-1.40), paracetamol use (POR 1.35, 95%CI 1.11-1.64), electric cooking (POR 1.42, 95%CI 1.11-1.80), regular exercise (POR 1.44, 95%CI 1.18-1.75) and maternal smoking (POR 1.16, 95%CI 1.01-1.33). Protective factors included having older siblings (two or more POR 0.69, 95%CI 0.58-0.82; one POR 0.86, 95%CI 0.71-1.05), and being born in the country of the survey (POR 0.74, 95%CI 0.63-0.87). Risk factors for rhinoconjunctivitis included regular consumption of meat, butter, margarine and nuts, regular exercise, regular television viewing, paracetamol use and passive smoking. Eczema was associated with regular meat, pasta and butter consumption, regular television viewing, paracetamol use and passive smoking. CONCLUSIONS: Regular meat and margarine consumption, paracetamol use, electric cooking and passive smoking are risk factors for symptoms of asthma, rhinoconjunctivitis and eczema in the Pacific. However, most of these associations were weak, and account for only a small proportion of cases.

Primary testicular diffuse large B-cell lymphomas have activated B-cell-like subtype characteristics.

Diffuse large B-cell lymphomas (DLBCLs) constitute a heterogeneous group of lymphomas in which germinal centre B-cell-like and activated B-cell-like subtypes can be discerned based on pathology, clinical presentation, and gene expression patterns. Testicular DLBCLs form an immune-privileged site-related subgroup of DLBCLs with an unfavourable prognosis. In the present study, cDNA microarray analysis, immunohistochemistry for CD10, Bcl6 and MUM1, and somatic hypermutation analysis of the immunoglobulin heavy chain gene rearrangements were used to determine the subtype of primary testicular DLBCL. Immunohistochemistry revealed 14/22 testicular DLBCLs with an activated B-cell-like immunophenotype and 8/22 with an ambiguous immunophenotype co-expressing CD10 and high levels of MUM1. cDNA microarray analysis of these 22 and four additional cases showed a uniform activated B-cell-like gene expression pattern for both immunophenotypes. Somatic hypermutation analysis of immunoglobulin heavy chain genes showed a very high mutation load in seven cases tested, but intraclonal heterogeneity was found at low level in only one of these cases. It is concluded that primary testicular DLBCLs have uniform activated B-cell-like subtype characteristics despite a number of cases showing an ambiguous immunophenotype.

The effect of storage on allergen and microbial agent levels in frozen house dust.

BACKGROUND: House dust samples collected for exposure studies are often stored for variable time periods until analysis. However, there is currently no information on the effects of dust storage on the content of biocontaminants. Therefore, associations were analysed between the levels of mite allergens (Der p 1, Der f 1), cat allergen (Fel d 1) and microbial components (endotoxin, beta(1-->3)-glucan) on the one hand and the storage duration of dust samples at -20 degrees C on the other hand. METHODS: Within the framework of a study on the influences of INdoor factors and Genetics on Asthma (INGA), dust samples were collected from living room floors between June 1995 and August 1998 and extracted according to a standardized protocol. The concentrations of Der p 1, Der f 1, Fel d 1 and beta(1-->3)-glucan were determined with specific enzyme immunoassays. Endotoxin content was quantified using a chromogenic kinetic Limulus amoebocyte lysate (LAL) test. All concentrations were expressed per gram of dust RESULTS: Dust samples (n = 1236) were obtained from 655 homes in Hamburg, Hettstedt, Zerbst and Bitterfeld. Storage duration (range 8-298 days) was grouped into four categories ( 120 d). After adjustment for city of residence and season of dust sampling, means ratios comparing categories 2-4 to the first category were not statistically significant for Der p 1, Der f 1, endotoxin and beta(1-->3 glucan). However, Fel d 1 concentrations significantly declined with increased storage times of dust samples. CONCLUSIONS: Storage of house dust at -20 degrees C for up to 10 months has no effect on mite allergen, endotoxin and beta(1-->3)-glucan levels. A potential loss of Fel d 1 during storage of frozen dust samples needs further investigations by repeated measurements of allergen in identical dust samples.

Bioaerosol health effects and exposure assessment: progress and prospects.

Exposures to bioaerosols in the occupational environment are associated with a wide range of health effects with major public health impact, including infectious diseases, acute toxic effects, allergies and cancer. Respiratory symptoms and lung function impairment are the most widely studied and probably among the most important bioaerosol-associated health effects. In addition to these adverse health effects some protective effects of microbial exposure on atopy and atopic conditions has also been suggested. New industrial activities have emerged in recent years in which exposures to bioaerosols can be abundant, e.g. the waste recycling and composting industry, biotechnology industries producing highly purified enzymes and the detergent and food industries that make use of these enzymes. Dose-response relationships have not been established for most biological agents and knowledge about threshold values is sparse. Exposure limits are available for some contaminants, e.g. wood dust, subtilisins (bacterial enzymes) and flour dust. Exposure limits for bacterial endotoxin have been proposed. Risk assessment is seriously hampered by the lack of valid quantitative exposure assessment methods. Traditional culture methods to quantify microbial exposures have proven to be of limited use. Non-culture methods and assessment methods for microbial constituents [e.g. allergens, endotoxin, beta(1-->3)-glucans, fungal extracellular polysaccharides] appear more successful; however, experience with these methods is generally limited. Therefore, more research is needed to establish better exposure assessment tools and validate newly developed methods. Other important areas that require further research include: potential protective effects of microbial exposures on atopy and atopic diseases, inter-individual susceptibility for biological exposures, interactions of bioaerosols with non-biological agents and other potential health effects such as skin and neurological conditions and birth effects.

Farm residence and exposures and the risk of allergic diseases in New Zealand children.

BACKGROUND: Studies in Europe have reported a reduced prevalence of allergy in farmers' children. We aimed to determine if there is a similar reduction in allergy among New Zealand farm children. METHODS: Two hundred and ninety-three children participated (60%) aged 7-10 years, from selected schools in small towns and the surrounding rural area. Skin prick tests (SPT) to eight common allergens were performed. Parents completed questionnaires about allergic and infectious diseases, place of residence, exposure to animals, and diet, and they provided dust from the living-room floor. Endotoxin was measured using an Limulus amoebocyte lysate (LAL) assay and Der p 1 using enzyme-linked immunoassay (ELISA). RESULTS: Current farm abode was found to increase the risk of having symptoms associated with allergy, but not SPT positivity. Independent inverse associations were found for early-life exposures: at least weekly consumption of yoghurt with hayfever (odds ratio (OR) = 0.3, 95% confidence intervals (CI) 0.1-0.7) and allergic rhinitis (OR = 0.3, 95% CI 0.2-0.7); any unpasteurized milk consumption with atopic eczema/dermatitis syndrome (AEDS) (OR = 0.2, 95% CI 0.1-0.8); cats inside or outside with hayfever (OR = 0.4, 95% CI 0.1-1.0) and AEDS (OR = 0.4, 95% CI 0.2-0.8); dogs inside or outside with asthma (OR = 0.4, 95% CI 0.2-0.8); and pigs with SPT positivity (OR = 0.2, 95% CI 0.1-0.9). CONCLUSIONS: Despite finding a protective effect of early-life animal exposures, we found a greater prevalence of allergic disease on farms.

Allergens and endotoxin on mothers' mattresses and total immunoglobulin E in cord blood of neonates.

The current authors examined whether mite and cat allergen and bacterial endotoxin levels in dust of the mothers' mattresses were associated with cord blood immunoglobulin (Ig)E (CB-IgE) levels in newborns. Data from 1,332 term and normal weight neonates, from an ongoing birth cohort study, influences of life-style related factors on the immune system and the development of allergies in childhood (LISA), with complete information on exposure to biocontaminants in mattress dust and CB-IgE were analysed. Two thirds of CB-IgE were undetectable (<0.35 kU x L(-1)). Thus, 0.35 and 0.45 kU x L(-1) (4th quartile) were chosen as cut-offs. Nonparametric smoothing (generalised additive models) showed statistically significant confounder-adjusted associations between elevated CB-IgE levels (> or = 0.45 kU x L(-1)) and log-transformed exposures to cat (linear), mite (inverse u-shaped), and endotoxin (u-shaped). After adjustment for covariables, elevated CB-IgE levels (logistic regression using the 1st-4th quartiles of exposure) were positively associated with high cat-allergen exposure and medium exposure to mite allergen, but were inversely associated with exposure to endotoxin. The associations were similar, but somewhat weaker, when 0.35 kU x L(-1) was used as cut-off. These results, showing an association between prenatal allergen and endotoxin exposures and immunoglobulin E production, suggest that the development of foetal immune responses may be affected.

Upper airway inflammation and respiratory symptoms in domestic waste collectors.

OBJECTIVES: To compare respiratory symptoms and upper airway inflammation in domestic waste collectors and controls, and to find the association between measures of upper airway inflammation on the one hand and exposure concentrations of organic dust or respiratory symptoms on the other hand. METHODS: In a cross sectional study among 47 waste collectors and 15 controls, questionnaire data on respiratory symptoms were collected. Nasal lavage (NAL), to assess upper airway inflammation, was performed before and after a work shift at the beginning and at the end of the working week. In NAL fluid, cells were counted and differentiated and concentrations of interleukin 6 (IL6), IL8, tumour necrosis factor-alpha (TNF alpha), and IL1 beta were measured. In collectors, inhalable dust samples were collected in which bacterial endotoxin and mould beta(1-->3)-glucan were assessed. RESULTS: Prevalence of respiratory symptoms was higher in waste collectors than in controls. Geometric mean exposure concentrations were 0.58 mg/m(3) for dust, 39 EU/m(3) for endotoxin, and 1.3 microg/m(3) for beta(1-->3)-glucan. At the end of the week collectors had higher concentrations of total cells and IL8 in NAL before and after a shift than controls (cells, before 1.9-fold p<0.10, after 3.3-fold p<0.01; IL8, before and after 1.8-fold p<0.05), and after/before work shift ratios of total cells were also higher (2.3-fold p=0.06) in collectors than in controls. Cells in NAL fluid consisted predominantly of neutrophils and epithelial cells, whereas eosinophils and mononuclear cells were rarely found. Exposure to dust and endotoxin was associated with concentrations of IL8 after the shift (p<0.05). Increased concentrations of IL8 (p<0.05) and total cells (p<0.10) after the shift were associated with respiratory symptoms. Concentrations of IL6, TNF alpha, and IL1 beta were not associated with waste collecting, symptoms, or exposure. CONCLUSIONS: Waste collectors show signs of increased upper airway inflammation and respiratory symptoms compared with controls. Exposure to organic dust probably underlies the inflammation mediated by neutrophils that result in respiratory symptoms.

Pets and vermin are associated with high endotoxin levels in house dust.

BACKGROUND: Previous studies have shown that the risk for allergic sensitization is lower in children who grew up on farms and in young adults who were exposed to dogs in early childhood. A higher microbial exposure in general and in particular to endotoxin in early childhood might contribute to this lower risk of atopy. OBJECTIVE: We examined whether the presence of pets or vermin in the home is associated with higher endotoxin concentrations in settled house dust. METHODS: House dust was sampled in a standardized manner on the living room floors of 454 homes of German children aged 5-10 years (participation rate 61%). Endotoxin was assessed with a quantitative kinetic chromogenic Limulus Amebocyte Lysate (LAL) method. Associations between endotoxin levels, pets and vermin are presented as ratios of the crude and confounder adjusted geometric means (means ratios) in the category of study vs. a reference category using multiple linear regression models. RESULTS: Endotoxin concentrations in living room floor dust sampled in homes without pets and vermin were lower (1246 ng per square meter, 1519 ng endotoxin/g dust, n = 157) than those sampled in homes with pets or vermin (2267 ng per square meter, 2200 ng endotoxin/g dust, n = 296). After adjustment for city of residence, season of dust sampling, age of the building and story of the dwelling, means ratios for endotoxin expressed per gram of dust were statistically significantly increased for dog (1.64, 95% CI 1.09-2.46), for cat (1.50, 95% CI 1.03-2.18) and for cockroach (3.01, 95% CI 1.37-6.60), whereas no major statistically significant associations were found for other pets, ants and mice. CONCLUSION: Keeping a dog or a cat in the home is consistent with higher exposure to endotoxin and might therefore contribute to the lower risk of atopy in later life.

Work-related symptoms in sewage treatment workers.

The aim of this study was to investigate health symptoms in sewage treatment workers. A health questionnaire was distributed among 147 sewage treatment workers. Correlating symptoms were clustered using principal component analysis and the association with self-reported exposure was assessed by calculating prevalence odds ratios (OR). Endotoxin was measured in two treatment plants. Personal endotoxin exposure was low (<10 Endotoxin Units/m(3)). Factor analysis of 29 symptoms resulted in four clusters of highly correlating symptoms: 'flu-like symptoms'; 'higher airway symptoms'; 'lower airway symptoms'; and 'neurological symptoms'. These clusters were positively associated with working with sewage, but only significant for 'flu- like symptoms' (OR=5.0; 95%CI=1.4-17.6; p<0.05) and 'neurological symptoms' (OR=4.2; 95%CI=1.5-11.7; p<0.01). Chemical exposure was associated with 'neurological symptoms' (OR=8.4; 95%CI=1.1-65.7; p<0.05). The use of daily washed working clothes was negatively associated with 'flu-like symptoms' (OR=0.3; 95%CI=0.1-0.6; p<0.01). In conclusion, sewage treatment workers develop a large variety of work-related symptoms that are not likely caused by endotoxin exposure only. Good hygienic practice at the workplace may prevent some of these symptoms

Occupational respiratory symptoms in New Zealand hairdressers.

BACKGROUND: A study of occupational respiratory symptoms in hairdressers was carried out in 26 salons in New Zealand. METHODS: A questionnaire was administered to 100 hairdressers and 106 office and shop workers, recording respiratory symptoms, demographic data, and smoking habits. Pulmonary function was measured before each shift. RESULTS: Hairdressers had a higher prevalence of asthma symptoms, diagnosed asthma, and asthma attacks in the previous 12 months, but these differences reduced markedly when adjusted for age, gender and smoking. Hairdressers had a significantly lower pulmonary function, even after adjustment for smoking, but this difference was entirely due to differences between hairdressers and office workers and there were no such differences between hairdressers and shop workers. CONCLUSIONS: The excess symptom prevalence in hairdressers was largely due to the high smoking rates. Mean pulmonary function was lower in hairdressers but this finding requires further study.

Application of the classic Limulus test and the quantitative kinetic chromogenic LAL method for evaluation of endotoxin concentration in indoor air.

The classic (gel-clot procedure) Limulus test (CLT) and the quantitative kinetic chromogenic LAL method (KQCL) used for the evaluation of bacterial endotoxin concentration in the indoor air of dwellings were compared. The scientific procedure included analyses of 40 air samples supplemented by the analysis of 20 sample duplicates (selected at random) which were taken during the fall season from 10 flats located in 3 towns of the Upper Silesian region (southern Poland). The particulate aerosol probes were sampled by Harvard impactor and Casella sampler. The same samples were analyzed in the Netherlands using the quantitative kinetic chromogenic LAL method, and in Poland using the classic Limulus test. Comparison of both methods revealed that the quantitative kinetic chromogenic LAL method was more precise, with better reproducibility (the coefficient of variation between analyses of the main probe and its duplicate was over two times smaller in the KQCL method than in the CLT method), fully automated in the phase of analysis and data reading, and faster and more effective than the classic Limulus test. Nevertheless, on the basis of the obtained results, the usefulness of the classic Limulus method for assessment of the degree of pollution of indoor air with bacterial endotoxin seems to be confirmed as in the majority of examined samples (21 out 40) the results obtained by both methods were of the same order of magnitude, and in the remaining 19 samples did exceed one order of magnitude. Thus, the data received by means of the classic Limulus test may be regarded as acceptable.

Longitudinal changes in inflammatory markers in nasal lavage of cotton workers. Relation to endotoxin exposure and lung function changes.

OBJECTIVES: The aim of this study was to evaluate time-related changes in inflammatory markers in nasal lavage (NAL) in relation to endotoxin exposure and lung function changes. METHODS: Endotoxin in cotton dust is well known to elicit proinflammatory mediators involved in acute or chronic respiratory inflammation due to cotton dust. We applied nasal lavage as well as lung function and exposure measurements in a small group (n = 11) of cotton workers during 6 weeks of observation (after 2 weeks free of exposure). Several inflammatory markers in NAL, such as interleukin 8 (IL-8), IL-6, and down-stream mediators, including soluble tumor necrosis factor receptor 75 (sTNF-R75) and soluble intercellular adhesion molecule 1 (ICAM-1), cell counts, marker of epithelial response (uric acid), and marker of plasma exudation (albumin) were measured. RESULTS: At the exposure levels examined in this study [cotton dust, geometric mean (GM) = 1.10 mg/m3; endotoxin, GM = 2,869 EU/m3], several inflammatory markers in NAL such as IL-8, sTNF-R75, and albumin tended to increase through the measurement period with increasing airborne endotoxin concentration (in six workers who had paired measurements in weeks 1 and 6). Although no statistical significance was reached, airborne endotoxin exposure and all inflammatory markers in NAL were relatively higher in cotton workers with a 6-week lung function decrease. CONCLUSION: The data suggest that inflammatory markers in NAL are associated with repeated airborne endotoxin exposure.

Loss of the plasma membrane-bound protein Gas1p in Saccharomyces cerevisiae results in the release of beta1,3-glucan into the medium and induces a compensation mechanism to ensure cell wall integrity.

Deletion of GAS1/GGP1/CWH52 results in a lower beta-glucan content of the cell wall and swollen, more spherical cells (L. Popolo, M. Vai, E. Gatti, S. Porello, P. Bonfante, R. Balestrini, and L. Alberghina, J. Bacteriol. 175:1879-1885, 1993; A. F. J. Ram, S. S. C. Brekelmans, L. J. W. M. Oehlen, and F. M. Klis, FEBS Lett. 358:165-170, 1995). We show here that gas1delta cells release beta1,3-glucan into the medium. Western analysis of the medium proteins with beta1,3-glucan- and beta1,6-glucan-specific antibodies showed further that at least some of the released beta1,3-glucan was linked to protein as part of a beta1,3-glucan-beta1,6-glucan-protein complex. These data indicate that Gas1p might play a role in the retention of beta1,3-glucan and/or beta-glucosylated proteins. Interestingly, the defective incorporation of beta1,3-glucan in the cell wall was accompanied by an increase in chitin and mannan content in the cell wall, an enhanced expression of cell wall protein 1 (Cwp1p), and an increase in beta1,3-glucan synthase activity, probably caused by the induced expression of Fks2p. It is proposed that the cell wall weakening caused by the loss of Gas1p induces a set of compensatory reactions to ensure cell integrity.

Measurement of beta(1-->3)-glucans in occupational and home environments with an inhibition enzyme immunoassay.

beta (1-->3)-Glucans are known for their potent ability to induce nonspecific inflammatory reactions and are believed to play a role in bioaerosol-induced respiratory symptoms. An inhibition enzyme immunoassay (EIA) was developed for the quantitation of beta (1-->3)-glucans in dust samples from occupational and residential environments. Immunospecific rabbit antibodies were produced by immunization with bovine serum albumin-conjugated laminarin [beta (1-->3)-glucan] and affinity chromatography on epoxy-Sepharose-coupled beta (1-->3)-glucans. The laminarin-based calibration curve in the inhibition EIA ranged from approximately 40 to 3,000 ng/ml (15 to 85% inhibition). Another beta (1-->3)-glucan (curdlan) showed a similar inhibition curve but was three to five times less reactive on a weight basis. Pustulan, presumed to be a beta (1-->6)-glucan, showed a parallel dose-response curve at concentrations 10 times higher than that of laminarin. Control experiments with NaIO4 and beta (1-->3)-glucanase treatment to destroy beta (1-->6)- and beta (1-->3)-glucan structures, respectively, indicate that the immunoreactivity of pustulan in the assay was due to beta (1-->3)-glucan and not to beta (1-->6)-glucan structures. Other polysaccharides, such as mannan and alpha (1-->6)-glucan, did not react in the inhibition EIA. Beta (1-->3)-Glucan extraction of dust samples in water (with mild detergent) was performed by heat treatment (120 degrees C) because aqueous extracts obtained at room temperature did not contain detectable beta (1-->3)-glucan levels. The assay was shown to detect heat-extractable beta (1-->3)-glucan in dust samples collected in a variety of occupational and environmental settings. On the basis of duplicate analyses of dust samples, a coefficient of variation of approximately 25% was calculated. It was concluded that the new inhibition EIA offers a useful method for indoor beta (1-->3)-glucan exposure assessment.

Retention of Saccharomyces cerevisiae cell wall proteins through a phosphodiester-linked beta-1,3-/beta-1,6-glucan heteropolymer.

Yeast cell wall proteins, including Cwp1p and alpha-agglutinin, could be released by treating the cell wall with either beta-1,3-or beta-1,6-glucanases, indicating that both polymers are involved in anchoring cell wall proteins. It was shown immunologically that both beta-1,3- and beta-1,6-glucan were linked to yeast cell wall proteins, including Cwp1p and alpha-agglutinin. It was further shown that beta-1,3-glucan was linked to the wall protein through a beta-1,6-glucan moiety. The beta-1,6-glucan moiety could be removed from Cwp1p and other cell wall proteins by cleaving phosphodiester bridges either enzymatically using phosphodiesterases or chemically using ice-cold aqueous hydrofluoric acid. These observations are consistent with the notion that cell wall proteins in Saccharomyces cerevisiae are linked to a beta-1,3-/beta-1,6-glucan heteropolymer through a phosphodiester linkage and that this polymer is responsible for anchoring cell wall proteins. It is proposed that this polymer is identical to the alkali-soluble beta-1,3-/beta-1,6-glucan heteropolymer characterized by Fleet and Manners (1976, 1977).

Enzyme immunoassays for total and allergen specific IgE in population studies.

OBJECTIVE: Extensive IgE serology in occupational or environmental health studies is often hampered by a lack of technical facilities and finance. The use in population studies of relatively simple and inexpensive enzyme immunoassays (EIAs) was therefore evaluated for the assessment of total serum immunoglobulin E (IgE), and of specific IgE reactions with various common (house dust mites, grass and birch pollen, and cat) or occupational (fungal alpha-amylase and rat urinary protein) allergens. METHODS: Total IgE was measured with a sandwich EIA, calibrated with commercially available IgE standards. Reproducibility was studied by testing pooled normal human serum samples in each of a large series of test plates. A panel of 156 children's serum samples with known IgE values was used to compare the assay with other total IgE assays. A previously developed EIA for anti-yeast IgE was adapted for the measurement of IgE reacting with various common and occupational allergens. Binding of IgE to microwells coated with commercially available allergen extracts, or allergen preparations from our own laboratory, was measured with a monoclonal anti-human IgE antibody and subsequent incubations with biotinylated rabbit anti-mouse Ig and avidin-peroxidase. Panels of serum samples from school children (n = 116), bakery workers (n = 126), and laboratory animal workers (n = 52) were used to study sensitivity and specificity, with reference to skin prick tests as the standard, and to compare the EIAs with commercially available test kits. RESULTS: The detection limit of the EIA for total IgE was 0.5-1 kU/l for undiluted serum samples, and the coefficient of variation between assays was less than 15% at serum concentrations between 1 and 150 kU/l. Results obtained with the panel of 156 children's serum samples were strongly correlated (r2 = 0.86) with IgE concentrations measured previously by radioimmunoassay. The results of the EIA for various occupational allergens correlated very well, both qualitatively and quantitatively, with the results of commercial test kits. Sensitivity and specificity of the EIA results as a predictor of skin prick test reactivity towards common allergens (house dust mite, grass pollen, birch pollen, and cat) were remarkably high (> 80%-90%) in the series of 116 children's serum samples. In a population of bakery workers the specificity of the EIAs was also very high (> 90%). The sensitivity was notably lower (30%-70%) in this adult population, which is, however, in agreement with results reported for conventional IgE tests. CONCLUSION: As the costs were estimated to be at least five to 10-fold lower than those of commercial test kits, the EIAs for total and specific IgE may be very useful tools in epidemiological studies of atopic respiratory or other disorders.

Influence of various dust sampling and extraction methods on the measurement of airborne endotoxin.

The influence of various filter types and extraction conditions on the quantitation of airborne endotoxin with the Limulus amebocyte lysate test was studied by using airborne dusts sampled in a potato processing plant. Samples were collected with an apparatus designed to provide parallel samples. Data from the parallel-sampling experiment were statistically evaluated by using analysis of variance. In addition, the influence of storage conditions on the detectable endotoxin concentration was investigated by using commercially available lipopolysaccharides (LPS) and endotoxin-containing house dust extracts. The endotoxin extraction efficiency of 0.05% Tween 20 in pyrogen-free water was seven times higher than that of pyrogen-free water only. Two-times-greater amounts of endotoxin were extracted from glass fiber, Teflon, and polycarbonate filters than from cellulose ester filters. The temperature and shaking intensity during extraction were not related to the extraction efficiency. Repeated freeze (-20 degrees C)-and-thaw cycles with commercial LPS reconstituted in pyrogen-free water had a dramatic effect on the detectable endotoxin level. A 25% loss in endotoxin activity per freeze-thaw cycle was observed. Storage of LPS samples for a period of 1 year at 7 degrees C had no effect on the endotoxin level. House dust extracts showed a decrease of about 20% in the endotoxin level after they had been frozen and thawed for a second time. The use of different container materials (borosilicate glass, "soft" glass, and polypropylene) did not result in different endotoxin levels. This study indicates that the assessment of endotoxin exposure may differ considerably between groups when different sampling, extraction, and storage procedures are employed.