F. Pacheco

Allergenic materials in the house dust of allergy clinic patients

INTRODUCTION Environmental agents including animal, fungal, tree, and weed antigens are known to cause allergic rhinitis and asthma. The following study was performed to measure the antigen concentration of several of these in house dust of children seen in an allergy clinic. Comparisons are made between household allergen levels of children seen for asthma and children seen for other reasons. METHODS Dust samples were solicited from patients in a pediatric allergy specialty clinic and other individuals associated with the clinic. Persons submitting dust were asked to complete a questionnaire describing their house. Samples were extracted, centrifuged, and filtered for sterility. Samples were stored in 50% glycerol at -20 degrees C. Specific antigens for Alternaria, Cladosporium, Aspergillus, Candida, Dermatophagoides farinae, cat, dog, oak, fescue, ragweed, plantain, and cockroach were measured using inhibition assays developed with whole antigen extract. Allergens Der p1, Der f 1, Alt a 1, and Alt a 70 kD were measured using double monoclonal antibody assays. RESULTS Significant concentrations of whole antigen from cat, dog, oak, Alternaria, and Cladosporium were detected. Between 0.1 and 18 microg of Der f1 and Der p1 per gram of dust were also measured. Alt a 1 and Alt a 70 kD levels varied between 3.0 and 1000 U/g of dust. Significant positive correlations were observed in levels of dust mite and Alternaria allergen for patients with an evaluation of asthma. CONCLUSIONS We found measurable levels of fungal antigens (Alternaria, Cladosporium), mite antigens, and animal antigens (dog and cat) in the majority of dust samples in this self-selected set of allergy clinic patients. Specific allergens Alt a 1, Alt a 70kD, and Der p 1 were significantly higher in the homes of asthmatic patients when compared with patients seen for reasons other than asthma. These studies support the hypothesis that fungal allergen exposure is an important component in the pathogenesis of the clinical condition known as asthma.

Comparison of outdoor allergenic particles and allergen levels

INTRODUCTION Spore and pollen counts have been used traditionally to determine aeroallergen exposure. Using a liquid based collector and enzyme immunoassays, we have developed methods for measuring airborne allergen concentrations. In this work we test the hypothesis that airborne allergen concentrations are directly related to spore and pollen counts. METHODS Test samplers used included a high-volume cyclonic liquid impinger (SpinCon) and a standard spore trap (Burkard). Samples were collected on a weekly basis from May to October and were analyzed microscopically for spores and pollen grains. The liquid samples were analyzed by enzyme-linked immunoassay for the presence of allergens from Alternaria, Cladosporium, Aspergillus, oak, fescue, ragweed, and plantain. Specific Alternaria allergens Alt al and GP70 also were measured. RESULTS Pollen counts for the SpinCon and Burkard collectors were similar, though spore counts were lower with the SpinCon. Detectable amounts of three of the seven allergenic species including fescue, ragweed, and Alternaria were present in air samples. Concentrations of pollens were seen in their respective seasons while fungal allergen levels varied throughout the period. Allergen levels generally agreed with particle counts, however peak allergen levels and peak particle counts for individual species did not correlate well. CONCLUSIONS At flow rates of 236 L/min, the SpinCon is comparable to the Burkard for counting airborne pollen and spores. Samples collected by the SpinCon permit quantitative determination of allergen levels in outdoor air. The poor correlation between measured airborne allergen and related particles indicates the potential for significant allergen exposure in the absence of identifiable particles in air.

IgE-reactive proteins from Stachybotrys chartarum.

BACKGROUND Stachybotrys chartarum has been associated with idiopathic pulmonary hemorrhage in infants. This is thought to be mycotoxin-related. There are increasing numbers of reports linking this fungus to the indoor environment of patients with other pulmonary problems, including allergies and asthma. OBJECTIVE Given the potential significance of this fungus as a pulmonary pathogen, this work evaluates the antigenic proteins of S. chartarum as to their molecular size and the prevalence of immunoglobulin (Ig)E and IgG directed against them in the general population. METHODS S. chartarum was isolated from a local home. S. chartarum for extract production was grown on minimum salts and glucose. Plasma from 132 healthy individuals was evaluated for IgE and IgG directed against S. chartarum using direct and inhibition enzyme immunoassay. The number and molecular size of those proteins that were bound by IgE from pooled sera known to contain IgE to S. chartarum were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis immunoblotting. RESULTS Enzyme immunoassay indicated 65 of 132 (49.2%) sera tested contained IgG against S. chartarum and 13 of 139 (9.4%) sera tested contained IgE against S. chartarum. Pooled sera identified two IgE-binding proteins from extracts of S. chartarum spores and mycelia. These proteins are 34 and 52 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis immunoblot. CONCLUSIONS We conclude sensitivity to S. chartarum is potentially much more widespread than previously appreciated. This fungus may impact the asthmatic and allergic population through both immunologic and toxic mechanisms. Its significance in the milieu of allergenic fungi may need to be re-evaluated.

The effect of temperature, relative humidity and rainfall on airborne ragweed pollen concentrations

Major weather parameters have long been known to alter airborne pollen and spore concentrations. The following study was conducted to study the effect of three of these parameters on airborne ragweed pollen concentrations.During the ragweed (RW) season for the years 1997 and 1998, 10 minute pollen collections were taken at least every 4 hours using an Allergenco MK-3 spore trap. Slides were fixed, and counted microscopically at 400X. During this same period, weather parameters were monitored by an Automated Weather Systems recording station located within a few meters of the collector.The ragweed season for this region begins in mid August and ends by mid October. Temperature patterns for the period demonstrated usual daily fluctuations with highs 13 to 35 °C and lows 8 to 24 °C. Relative humidity readings for the period varied between 25 and 100%. Highest RW values were seen after seasonal cooling in September. Daily rainfall for the period varied between 0 and 100 mm. Airborne RW always declined sharply after strong rainfall events (> 10 mm/day). Peak airborne RW concentrations were often associated with the passing of frontal boundaries and the change in wind direction and velocity that accompanies that passing.Factors influencing airborne RW concentrations are multiple and complex, but atmospheric forces have great influence. The passing of major weather fronts and the associated temperature drops, wind disturbances and rainfall are the major factors.

The effect of time and extraction buffers on residual protein and allergen content of extracts derived from four strains of Alternaria

BACKGROUND A series of studies was performed to identify optimal elution conditions for production of desired Alternaria allergens with simultaneous reduction of undesired ones. METHODS Proteins and allergens from four strains of Alternaria extracted for differing time intervals and in different buffers were analyzed by one- and two-dimensional electrophoresis, as well as by immunoglobulin E enzyme-linked immunosorbent assay inhibition and immunoblotting. RESULTS The amount of protein and carbohydrate released varied with each time interval but was consistent between buffers. Extracts from longer time intervals tended to contain more carbohydrate. Electrophoresis of the four strains demonstrated many similar proteins; however, the concentrations of these proteins showed considerable interstrain difference. Comparison of extraction times for single strains by immunoblotting showed that certain allergens are preferentially released during specific time intervals. Some allergens were seen to be most prevalent in a 24-hour extract, whereas others were most prevalent in a 1-hour extract. Two-dimensional electrophoresis resolved bands into discrete spots. The major shared elements of the four strains could be easily identified. The appearance and disappearance of individual protein elements with time was seen. CONCLUSIONS Elution conditions have a significant impact on quantities of specific glycoproteins contained in extracts of Alternaria and must be controlled and optimized when such extracts are produced for allergen purification.

The impact of home cleaning on quality of life for homes with asthmatic children

Treatment with common household bleach containing hypochlorite destroys dust mites and denatures protein allergens. The purpose of this study was to determine if home use of hypochlorite products results in lowered exposure to bacteria, fungi, and protein allergens and improved quality of life (QOL) for asthmatic persons in the home. Asthmatic and nonasthmatic households containing at least three persons (between 2 and 17 years of age) were recruited. Households were supplied one of three sets of cleaning products (regular products, some containing hypochlorite; regular products plus three additional products with dilute hypochlorite; control, no products). Participants were supplied with cleaning instructions and asthma education. The control group was instructed to clean as usual. Participants completed general health and QOL questionnaires. Asthmatic participants completed an additional asthma QOL questionnaire. Families participated in the study for 8 weeks and completed the full set of questions every 2 weeks. Homes were visited at the beginning of the study and twice thereafter at monthly intervals. Samples evaluated were surface bacteria, viable and nonviable airborne spores, and dust antigen content. Reductions in surface bacteria, airborne fungal spores, and dust antigen levels were achieved. Significant improvement in general health parameters was seen for the asthmatic product groups over the control group. Significant improvement in general QOL and asthma-specific QOL was seen in the asthmatic group. Emphasis on cleaning and cleaning education combined with hypochlorite-based cleaning supplies resulted in significantly improved QOL for families with asthmatic children.

Monoclonal Antibody-Based Assay for Alt a1, a Major Alternaria Allergen

BACKGROUND Allergenic materials from Alternaria are significant causes of human disease. One 31-kD glycoprotein, designated Alt a1, has been described previously as being a significant allergen. OBJECTIVES The purpose of this study is to characterize purified Alt a1 and to provide a method for measuring it in laboratory and environmental samples. METHODS Monoclonal antibodies directed to the purified allergen were produced and used to construct a double monoclonal assay for Alt a1. The purified allergen detected by this assay was tested for IgE binding in vitro and in vivo. RESULTS Purified allergen detected by this assay was shown to bind IgE from patients sensitive to Alternaria and gave a positive skin test in 15 of 16 patients skin test reactive to commercial Alternaria extracts. The assay is sensitive to Alt a1 concentrations as low as 0.2 microg/nmL and is very specific for the allergen. The three commercial Alternaria extracts assayed contained measurable quantities of the allergen varying from 12 to 84 microg/mL. CONCLUSIONS Alt a1 is a significant Alternaria allergen. The ability to measure this allergen may assist in standardizing Alternaria extracts and lead to an understanding of the significance of exposure to this allergen.

Low-cost interventions improve indoor air quality and children's health.

Intervention in the home environment to reduce asthma triggers theoretically improves health outcomes for asthmatic children. Practical benefit from application of these interventions has proven difficult. This single-blind study tested the effectiveness of simple low-cost home interventions in improving health scores of children with asthma. Families with at least one asthmatic child were recruited. Initial health examination, health, and home assessments were conducted and targeted interventions were implemented. Interventions included dehumidification, air filtration, furnace servicing, and high-efficiency furnace filters. When present, gross fungal contamination was remediated. Asthma education was provided along with education in healthy home practices. Follow-up assessments were conducted after 6 months. Health surveys were completed at enrollment and follow-up. This study enrolled 219 children with asthma. Home inspections and interventions were conducted in 181 homes and 83 families completed all phases. Reduction in asthma and allergy-related health scores was shown in follow-up health surveys. Health improvements were significant for cough when heating, ventilation, and air conditioning (HVAC) service and dehumidification were used. Breathing problems were significantly improved for dehumidification, HVAC service, and room air cleaners. Total dust allergen load was reduced for the dehumidification group (p < 0.05). Mold spore counts were reduced one order of magnitude in 25% of the homes. Indoor spore counts adjusted for outdoor spore levels were reduced overall (p < 0.01). Simple low-cost interventions directed to producing cleaner indoor air coupled with healthy home education improve the indoor air quality and health in asthmatic children.

Hourly variation of airborne ragweed pollen in Kansas City.

INTRODUCTION Ragweed pollen is one of the major causes of allergic rhinitis in the midwest United States. Previous studies have demonstrated that ragweed pollen production begins after sunrise and airborne pollen levels peak several hours later. Variations in this pattern that may be of clinical importance within a small region and the effect of weather on these variations have not been investigated. METHODS Volumetric pollen collectors were stationed at four sites around the metropolitan area. Ten-minute grab samples were taken at each location every 2 hours for continuous 24-hour periods during the 1997 ragweed season. The downtown location had a weather station that logged meteorological conditions at hourly intervals during this time. Ragweed pollen grains were counted microscopically at 400 x. Uninterrupted data covering at least 20 days during the season were evaluated for each collection site. RESULTS The lowest ragweed pollen counts occur at 6:00 AM and the highest occur at noon for both suburban and urban sites. Rainfall was seen to be the most influential weather-related factor. Significant rainfall events effectively removed pollen grains from the air. CONCLUSIONS Though ragweed pollen emission begins at 6:00 AM, peak pollen exposure occurs at midday. This observation is in agreement with many other studies. The myth that highest ragweed exposure occurs in the early morning hours is not supported by this or previous studies.

Comparison of the Burkard and Allergenco MK-3 volumetric collectors

BACKGROUND The Burkard sampler is a widely used volumetric pollen and spore collector, in part, because it is wind-oriented, it has consistent flow characteristics, and it permits time-discrimination of collected particles. Its main disadvantages are that it is heavy, expensive, and visual counting is very time-consuming. A less-expensive volumetric collector with time discrimination capabilities could permit more widespread particle counting which would enhance our understanding of aerobiology. OBJECTIVE The objective of this study is to compare the collection recoveries of the Burkard sampler with a less-expensive non-wind oriented collector, the Allergenco MK-3, under various wind speeds. METHODS Pollen and spore counts were compared on 20 sampling days during the spring pollen season using a Burkard and Allergenco MK-3 located next to each other on the roof of a 5-story hospital building. A weather station was placed nearby and wind velocity was concurrently measured. RESULTS The median wind velocity was 6 miles/hour with a maximum of 35 miles/hour. The Burkard and Allergenco MK-3 collectors displayed similar collection characteristics at all wind velocities for both pollen and spores. The Burkard gave lower counts than the Allergenco when absolute particle counts were low and similar values at higher absolute counts. CONCLUSIONS Given our data, we conclude that ambient wind speed has no significant effect on collection efficiency at velocities commonly found on the roof of our hospital and that the collection characteristics of the Burkard and Allergenco MK-3 are comparable.