J.K. Sercombe

Airborne fungal fragments and allergenicity

Exposure to fungi, particularly in water damaged indoor environments, has been thought to exacerbate a number of adverse health effects, ranging from subjective symptoms such as fatigue, cognitive difficulties or memory loss to more definable diseases such as allergy, asthma and hypersensitivity pneumonitis. Understanding the role of fungal exposure in these environments has been limited by methodological difficulties in enumerating and identifying various fungal components in environmental samples. Consequently, data on personal exposure and sensitization to fungal allergens are mainly based on the assessment of a few select and easily identifiable species. The contribution of other airborne spores, hyphae and fungal fragments to exposure and allergic sensitization are poorly characterized. There is increased interest in the role of aerosolized fungal fragments following reports that the combination of hyphal fragments and spore counts improved the association with asthma severity. These fragments are particles derived from any intracellular or extracellular fungal structure and are categorized as either submicron particles or larger fungal fragments. In vitro studies have shown that submicron particles of several fungal species are aerosolized in much higher concentrations (300-500 times) than spores, and that respiratory deposition models suggest that such fragments of Stachybotrys chartarum may be deposited in 230-250 fold higher numbers than spores. The practical implications of these models are yet to be clarified for human exposure assessments and clinical disease. We have developed innovative immunodetection techniques to determine the extent to which larger fungal fragments, including hyphae and fractured conidia, function as aeroallergen sources. These techniques were based on the Halogen Immunoassay (HIA), an immunostaining technique that detects antigens associated with individual airborne particles >1 microm, with human serum immunoglobulin E (IgE). Our studies demonstrated that the numbers of total airborne hyphae were often significantly higher in concentration than conidia of individual allergenic genera. Approximately 25% of all hyphal fragments expressed detectable allergen and the resultant localization of IgE immunostaining was heterogeneous among the hyphae. Furthermore, conidia of ten genera that were previously uncharacterized could be identified as sources of allergens. These findings highlight the contribution of larger fungal fragments as aeroallergen sources and present a new paradigm of fungal exposure. Direct evidence of the associations between fungal fragments and building-related disease is lacking and in order to gain a better understanding, it will be necessary to develop diagnostic reagents and detection methods, particularly for submicron particles. Assays using monoclonal antibodies enable the measurement of individual antigens but interpretation can be confounded by cross-reactivity between fungal species. The recent development of species-specific monoclonal antibodies, used in combination with a fluorescent-confocal HIA technique should, for the first time, enable the speciation of morphologically indiscernible fungal fragments. The application of this novel method will help to characterize the contribution of fungal fragments to adverse health effects due to fungi and provide patient-specific exposure and sensitization profiles.

The reduction of rhinitis symptoms by nasal filters during natural exposure to ragweed and grass pollen

Background:  Prototype nasal filters were developed to collect inhaled pollen. This study evaluated the efficacy of the filters for prevention of rhinitis symptoms during acute outdoor pollen exposure.

The nasal air sampler: a device for sampling inhaled aeroallergens

OBJECTIVE The object is to design, develop, and test a personal aerosol sampling device consisting of impaction samplers worn just inside the nostrils, driven by the wearers respiration. The device provides a novel and unique measure of individual exposure to aeroallergens. It was conceived as an integral part of an allergen diagnostic system, in which collected aerosols are immunostained with monoclonal antibodies or the patients IgE and associated particles positively identified using techniques of image analysis. METHODS Each sampler comprises a slot impactor with a detachable impaction plate covered with either a specially developed medical adhesive or a protein-binding membrane. Sampler performance has been validated by rig tests of aerodynamic resistance and collection efficiency of different sized particles at various flow rates. There have also been field trials with human subjects which show that the sampler can be comfortably worn for periods of up to 4 hours. This is sufficient to gather a representative sample of inhaled allergens in most environments. RESULTS The sampler collects an increasing proportion of particles in the inhalable range at and above 5 microm. This includes most bioaerosols of interest to allergists. Sampler prototypes have been built by CNC mill and stereolithography. Batches of samplers have been molded in biocompatible materials for field and clinical trials. CONCLUSIONS The device successfully collects aeroallergens from a patients own respiration. While developed specifically as a vehicle for the allergen diagnostic system, it can be adapted for studies of other aspects of air quality or for prophylactic use.

Four methods of sampling for dust mite allergen: differences in ‘dust’

Background: Measurement of exposure to the dust mite allergen Der p 1 is important in asthma research and is potentially useful in managing asthma. As no single measure can capture all characteristics of an exposure, it is important to recognize differences in the available methods of measuring exposure to Der p 1.

Effect of Floorcovering Construction on Content and Vertical Distribution of House Dust Mite Allergen, Der p I

Domestic floorcoverings often contain appreciable quantities of particulate pollutants, such as house dust mite allergen, Der p I. Exposure to Der p I is a risk factor for the development of mite sensitization and asthma. We investigated whether carpet construction was related to the Der p I content in normal use, and its vertical distribution. We hoped to inform development of methods to remove such material. Along with one hard flooring surface, a range of carpets with differing pile conformations (loop vs. cut), pile heights, yarn twists, and pile densities were placed in houses for 13 months. The carpets were later sectioned to allow profiling of Der p I throughout the pile strata using monoclonal antibody, enzyme-linked immunosorbent assay. Not surprisingly, significantly lower amounts of Der p I were found on hard flooring than any of the carpets, which all contained similar amounts of mite allergen. For all carpets, the Der p I concentration per unit area was found to be inversely related to the distance from the carpet backing. We conclude that carpet construction is not a good predictor of Der p I content in the home. We also suggest that, as carpet construction likely influences ease of disturbance of material within the pile, methods to remove or denature particulate pollutants such as Der p I will be most effective if they are able to target the bulk of allergenic material, found toward the base of the pile.

Evaluation of home allergen sampling devices

Background:  Simple, inexpensive methods of sampling from allergen reservoirs are necessary for large‐scale studies or low‐cost householder‐operated allergen measurement.

Domestic exposure to fungal allergenic particles determined by halogen immunoassay using subject's serum versus particles carrying three non-fungal allergens determined by allergen-specific HIA.

UNLABELLED Studies that estimate indoor aeroallergen exposure typically measure a pre-selected limited range of allergens. In this study, inhalable aeroallergen particles were quantified using the halogen immunoassay (HIA) to determine the contribution of fungal and non-fungal aeroallergens to total allergen exposure. Bioaerosols from 39 homes of fungal-allergic subjects were sampled using inhalable fraction samplers and immunostained by HIA using resident subjects immunoglobulin E (IgE) to detect allergen-laden particles. Fungal aerosols as well as particles carrying mite, cat, and cockroach allergens were identified and enumerated by HIA. Reservoir dust-mite (Der p 1), cat (Fel d 1), and cockroach (Bla g 1) allergen concentrations were quantified by ELISA. Fungal particles that bound subjects IgE in the HIA were 1.7 (bedroom)- and 1.4 (living room)-fold more concentrated than Der p 1, Fel d 1, and Bla g 1 allergen particles combined. Predominant fungal conidia that bound IgE were derived from common environmental genera including Cladosporium and other fungi that produce amerospores. Airborne mite, cat, and cockroach allergen particle counts were not associated with reservoir concentrations determined by ELISA. This study demonstrates that inhalable fungal aerosols are the predominant aeroallergen sources in Sydney homes and should be considered in future exposure assessments. PRACTICAL IMPLICATIONS Indoor allergen exposure assessment studies have primarily focused on a limited range of allergen sources in samples derived from reservoir dust samples. Using an innovative immunodiagnostic approach, this study demonstrates that fungal bioaerosols are the dominant source of aeroallergen exposure in the domestic environment, providing unique insight into domestic aeroallergen exposure.