Aeroallergen Monitoring by the National Allergy Bureau: A Review of the Past and a Look Into the Future.

Monitoring aeroallergens has a long history within the American Academy of Allergy, Asthma & Immunology. The Aeroallergen Network of the National Allergy Bureau is composed mainly of members of the American Academy of Allergy, Asthma & Immunology, whose objectives are to enhance the knowledge of aerobiology and its relationship to allergy, increase the number of certified stations, maintain the standardization and quality of aerobiology data, improve the alert and forecast reporting system, and increase ties with other scientific entities inside and outside the United States. The public has a keen interest in pollen counts and pollen forecasts, as do many health professionals in the allergy community. In this review, we explore the past, present, and future of allergen monitoring with a focus on methods used for sampling, the training of those performing the analysis, and emerging technologies in the field. Although the development of automated samplers with machine intelligence offers great promise for meeting the goal of a fully automated system, there is still progress to be made regarding reliability and affordability.

Hurricane María drives increased indoor proliferation of filamentous fungi in San Juan, Puerto Rico: a two-year culture-based approach.

Extensive flooding caused by Hurricane María in Puerto Rico (PR) created favorable conditions for indoor growth of filamentous fungi. These conditions represent a public health concern as contamination by environmental fungi is associated with a higher prevalence of inflammatory respiratory conditions. This work compares culturable fungal spore communities present in homes that sustained water damage after Hurricane María to those present in dry, non-flooded homes. We collected air samples from 50 houses in a neighborhood in San Juan, PR, 12 and 22 months after Hurricane María. Self-reported data was used to classify the homes as flooded, water-damage or dry non-flooded. Fungi abundances, composition and diversity were analyzed by culturing on two media. Our results showed no significant differences in indoor fungal concentrations (CFU/m3) one year after the Hurricane in both culture media studied (MEA and G25N). During the second sampling period fungal levels were 2.7 times higher in previously flooded homes (Median = 758) when compared to dry homes (Median = 283), (p-value < 0.005). Fungal profiles showed enrichment of Aspergillus species inside flooded homes compared to outdoor samples during the first sampling period (FDR-adjusted p-value = 0.05). In contrast, 22 months after the storm, indoor fungal composition consisted primarily of non-sporulated fungi, most likely basidiospores, which are characteristic of the outdoor air in PR. Together, this data highlights that homes that suffered water damage not only have higher indoor proliferation of filamentous fungi, but their indoor fungal populations change over time following the Hurricane. Ultimately, after nearly two years, indoor and outdoor fungal communities converged in this sample of naturally ventilated homes.

Comparison of Atmospheric Fungal Spore Concentrations between Two Main Cities in the Caribbean Basin.

OBJECTIVE: Fungal spores are ubiquitous in the atmosphere worldwide, but their distribution is not homogeneous at different locations. Most studies have compared airborne fungal spores ecology in temperate zones, but less is known about the tropics. METHODS: This study compared, through statistical analysis of archived datasets, the predominant fungal groups, patterns and meteorological variables affecting airborne fungal spore concentrations between two major cities in the Caribbean (Havana and San Juan) during the year 2015. RESULTS: In Havana, the predominant fungal group was Cladosporium while in San Juan were basidiospores. Our data provide evidence of differences and similarities in the monthly distribution of airborne spores in Havana and San Juan, but Cladosporium, ascospores y basidiospores had comparable hourly patterns in both cities and were affected by the same meteorological variables. CONCLUSION: Our study provides additional evidence to help design allergy interventions.

Identification of Immunoglobulin E-Binding Proteins of the Xerophilic Fungus Aspergillus penicillioides Crude Mycelial Mat Extract and Serological Reactivity Assessment in Subjects with Different Allergen Reactivity Profiles.

BACKGROUND: Aspergillus penicillioides is a very common indoor xerophilic fungus and potential causative agent of respiratory conditions. Although people are constantly exposed to A. penicillioides, no proteins with allergenic potential have been described. Therefore, we aim to confirm allergic sensitization to A. penicillioides through reactivity in serological assays and detect immunoglobulin E (IgE)-binding proteins. METHODS: In an indirect ELISA, we compared the serological reactivity to A. penicillioides between subjects with specific IgE (sIgE) (group 1, n = 54) and no sIgE reactivity (group 2, n = 15) against commercial allergens. Correlations and principal component analysis were performed to identify associations between reactivity to commercial allergens and A. penicillioides. IgE-binding proteins in A. penicillioides were visualized using Western blotting (WB) in group 1. The IgE-binding proteins with the highest reactivity were analyzed by mass spectrometry and confirmed by transcript matching. RESULTS: There was no statistical significance (p = 0.1656) between the study groups in serological reactivity. Correlations between reactivity to A. penicillioides, dog epithelia, Aspergillus fumigatus, and Penicillium chrysogenum were observed. WB experiments showed 6 IgE-binding proteins with molecular weights ranging from 45 to 145 kDa. Proteins of 108, 83, and 56 kDa showed higher reactivity. Mass spectrometry analysis of these 3 proteins led to the putative identification of NADP-specific glutamate dehydrogenase and catalase B. This was confirmed with transcriptome analysis. CONCLUSIONS: These results provide evidence of the presence of potential allergenic components in A. penicillioides. Further analysis of the putatively identified proteins should reveal their allergenic potential.

Serological Reactivity and Identification of Immunoglobulin E-Binding Polypeptides of Ganoderma applanatum Crude Spore Cytoplasmic Extract in Puerto Rican Subjects.

BACKGROUND: The allergenic potential of Ganoderma applanatum basidiospores has been demonstrated previously in Puerto Rico. However, basidiomycete allergens are not available for inclusion in allergy diagnostic panels. Therefore, we sought to confirm allergic sensitization to G. applanatum crude spore cytoplasmic extract through reactivity in serological assays and detection of immunoglobulin E (IgE)-binding polypeptides. METHODS: Via an indirect ELISA, serological reactivity was compared between groups of individuals with different allergic profiles. Group 1 (n = 51) consisted of individuals with sIgE to the allergens included in the diagnostic panels; group 2 (n = 14) comprised individuals with no sIgE to the allergens tested; and group 3 (n = 22) included individuals with no allergic history. To visualize IgE-binding polypeptides, group 1 sera were examined via Western blotting (WB). Polypeptide bands with the highest reactivity were analyzed by mass spectrometry (MS) for putative identification. RESULTS: The serological reactivity of group 1 was significantly higher than that of group 3 in an indirect ELISA (p = 0.03). Sixty-five percent of group 1 individuals showed reactivity to polypeptide bands in WB. Bands of 81 and 56 kDa had the highest reactivity proportions among the reactive sera, followed by a 45-kDa band. MS analysis of these 3 polypeptides suggests that they are basidiomycete-derived enzymes with aconitate hydratase, catalase, and enolase functions. CONCLUSIONS: G. applanatum spores have allergenic components recognized by Puerto Rican individuals, which could eventually be considered as markers in cases of fungal allergy and be included in diagnostic allergen panels in Puerto Rico and tropical regions.

Performance of the halogen immunoassay to assess airborne mouse allergen-containing particles in a laboratory animal facility.

Airborne mouse allergen is a risk factor for respiratory diseases. Conventional assessment techniques provide mass-based exposure estimates that may not capture completely the inhalation risk of airborne allergen particles. In contrast to mass-based estimates, the halogen immunoassay (HIA) combines immunoblotting and microscopy to directly assess allergen-containing particles. We evaluated the HIA for the assessment of airborne mouse allergen and compared the results to the enzyme linked immunosorbent assay (ELISA). Particulate matter (PM)(10) and PM(2.5) samples (30 min, 4 l/m) were collected in a mouse facility before, during, and after disturbance of soiled bedding. Concentrations of Mus m 1-positive particles (haloed particles (HPs)) and intensities of the haloes were determined with the HIA. Although HPs/m(3) were positively correlated with mass concentration (statistically significant only with Mus m 1 concentration on PM(10)), replicates of mass concentration showed higher variability than HPs/m(3). After disturbance, most of the HPs were in the PM(2.5) fraction. Mean haloes intensities were similar before, during, and after disturbance. The HIA was able to measure allergen-containing particles with less variability than the ELISA, detected the shift of HPs to smaller particles after disturbance, and may suggests similar halo intensity by particles detected during and after disturbance. Our findings suggest that the HIA can be used to assess indoor concentrations of mouse allergen particles and their morphological characteristics.

Comparison of the interleukin-1ß-inducing potency of allergenic spores from higher fungi (basidiomycetes) in a cryopreserved human whole blood system.

BACKGROUND: Spores from basidiomycete fungi (basidiospores) are highly prevalent in the atmosphere of urban and rural settings. Studies have confirmed their potential to affect human health as allergens. Less is known about their potential to serve as stimuli of the innate immune system and induce proinflammatory reactions. METHODS: In this study, we evaluated the proinflammatory potential of spores from 11 allergenic basidiomycete species (gilled: Pleurotus ostreatus,Oudemansiella radicata,Armillaria tabescens,Coprinus micaceus,Pluteus cervinus, and Chlorophyllum molybdites, and nongilled: Pisolithus arhizus,Merulius tremellosus,Calvatia cyathiformis,Lycoperdon pyriforme, andBoletus bicolor) based on their potency to induce the release of the proinflammatory cytokine interleukin (IL)-1ß in a cryopreserved human whole blood system. In addition, the roles of morphological features of the spores (surface area, shape, and pigmentation) were examined for their role in the IL-1ß-including potency of spores. Peripheral blood from healthy volunteers was collected, pooled, and cryopreserved. After stimulating the cryopreserved pooled blood with 10(6) to 10(3) basidiospores/ml, the concentration of IL-1ß in culture supernatants was determined with ELISA. RESULTS: Basidiospores manifested concentration-dependent IL-1ß-inducing potency, which was more marked among basidiospores from gilled basidiomycetes. At higher concentrations of basidiospores, the IL-1ß-inducing potency could be differentiated in the cryopreserved human whole blood system. Morphological features did not correlate with the IL-1ß-inducing potency of the basidiospores, suggesting that nonmorphological properties modulate the IL-1ß-inducing potency. CONCLUSION: Our data provide evidence of the proinflammatory potential of basidiospores, and the utility of cryopreserved human whole blood as a human-based in vitro system to study the immune reactivity of allergenic basidiospores.

Evidence for the detection of non-endotoxin pyrogens by the whole blood monocyte activation test.

Threats of pyrogenicity were discovered more than a century ago. Measures to determine the safety of parenterals and, more recently, medical devices and cell therapies for human use have been in place for 70 years. Currently, there are three testing possibilities available: the Rabbit Pyrogen Test, the Limulus Amebocyte Lysate test (Bacterial Endotoxin Test), and test systems using human whole blood or human monocytes, called Monocyte Activation Test (MAT). The MAT is based on the human fever reaction and thus most closely reflects the human situation. Unfortunately, regulations and testing guidelines are not fully harmonized, despite formal international validation. Furthermore, data showing that the MAT is capable of covering the totality of possible pyrogens relevant to humans were not included in the MAT validations of the last decade. For this review we collate evidence from published literature, unpublished data of our own, and results from the international validation study to show that there is overwhelming scientific evidence to conclude that the whole blood MAT reliably detects non-endotoxin pyrogens. Therefore, further validation exercises do not seem warranted.

Immunodetection and quantification of airborne (1-3)-ß-D-glucan-carrying particles with the halogen immunoassay.

Fungal cell wall components, such as (1-3)-ß-D-glucan, are known to be capable of activating the innate immune system and pose a respiratory health risk in different environments. Mass-based non-viable techniques commonly used for assessment of fungal exposures could be ß-D-glucan-specific, but are limited to analysis of liquid extracts. The variable solubility of different ß-D-glucans may underestimate ß-D-glucan exposure and long sampling times required for mass-based methods make assessing short-term exposures difficult. In this study, we evaluated the utility of the halogen immunoassay (HIA), an immunoblotting technique previously used for allergens, to immunodetect and quantify ß-D-glucan-carrying particles (BGCPs). The HIA was able to detect BGCPs without background staining when ß-D-glucan standards and air samples collected at a poultry house during short sampling periods were evaluated. The image analysis protocol previously developed by our group for mouse allergen allowed simultaneous immunodetection and quantification of ß-D-glucan-containing particles. Our results suggest that the HIA holds promise for quantifying ß-D-glucan exposures. To our knowledge, this is the first time in which the HIA was used for non-allergenic compounds of microbial or fungal origins.

Cryopreserved human whole blood: a human-based in vitro immunotoxicological system.

The airborne environment poses numerous human health risks, including potential exposure to particulate matter and to fungal bioaerosols with pro-inflammatory capabilities. The utility of animal models in the study of the human health effects of these exposures is often limited. This is due to the necessity for a high-throughput of samples and the high financial costs involved, as well as various associated animal welfare issues. In addition, animal models might not appropriately predict the responses of humans to pro-inflammatory stimuli. Here are discussed the recent applications of a human-based assay, the cryopreserved human whole blood system, to study the human health effects of exposures to particulate matter and to the less-studied fungal spores of higher fungi (phylum Basidiomycota or 'true mushrooms'). Evidence is provided to support the utility of the human-based assay in the fields of environmental health and medical mycology, for the evaluation of common paradigms in environmental health studies, and to highlight the potential human health effects of spores of basidiomycete fungi.

Skin test reactivity of allergic subjects to basidiomycetes' crude extracts in a tropical environment.

Fungal allergies can be detected by the skin prick test with extracts of the organisms, but not all fungi, including the basidiomycetes, are being examined. We determined the level of sensitization to basidiomycetes in allergic subjects and compared their reactivity to commercial extracts commonly used to detect allergies. Crude spore extracts of the basidiomycetes Ganoderma applanatum, Chlorophyllum molybdites, and Pleurotus ostreatus, which are known to release numerous spores, were examined along with commercial extracts on 33 subjects with asthma, allergic or non-allergic rhinitis. Overall, affected subjects showed the highest reactivity to mites (36%), followed by Ganoderma applanatum (30%), grass (27%) Chlorophyllum molybdites (12%) and Pleurotus ostreatus (12%). Allergic rhinitis patients were most reactive to mites (58%), grass (42%), Ganoderma applanatum (25%), Penicillium spp. (25%), and cat (17%). Those with asthma primarily responded to mites (44%), Ganoderma applanatum (44%), grass (33%), and Pleurotus ostreatus (22%). IgE levels correlated with positive basidiomycetes extracts. This finding, coupled with higher reactivity to basidiospores as compared to mitospores, and the similar sensitivities of patients to G. applanatum and mites, suggest that basidiomycetes are important allergen sources in the tropics.

Sensitization to airborne ascospores, basidiospores, and fungal fragments in allergic rhinitis and asthmatic subjects in San Juan, Puerto Rico.

BACKGROUND: Fungal spores are the predominant biological particulate in the atmosphere of Puerto Rico, yet their potential as allergens has not been studied in subjects with respiratory allergies. The purpose of this study was to determine the level of sensitization of subjects with respiratory allergies to these particles. METHODS: Serum samples were drawn from 33 subjects with asthma, allergic rhinitis, or nonallergic rhinitis and 2 controls with different skin prick test reactivity. An MK-3 sampler was used to collect air samples and the reactivity of the sera to fungal particles was detected with a halogen immunoassay. RESULTS: All subjects reacted to at least 1 fungal particle. Thirty-one subjects reacted to ascospores, 29 to basidiospores, 19 to hyphae/fungal fragments, and 12 to mitospores. The median percentage of haloes in allergic rhinitis subjects was 4.82% while asthma or nonallergic rhinitis subjects had values of 1.09 and 0.39%, respectively. Subjects with skin prick tests positive to 3, 2, 1, or no extract had 5.24, 1.09, 1.61, and, 0.57% of haloed particles, respectively. If skin prick tests were positive to basidiomycetes, pollen, animals, or deuteromycetes, the percentages of haloes were 4.72, 4.15, 3.63, and 3.31%, respectively. Of all haloed particles, 46% were unidentified, 25% ascospores, 20% basidiospores, 7% hyphae/fungal fragments, and 2% mitospores. IgE levels and the number of positive skin prick test extracts correlated with the percentage of haloes. CONCLUSION: In tropical environments, sensitization to airborne basidiomycetes, ascomycetes, and fungal fragments seems to be more prevalent than sensitization to mitospores in subjects with active allergies, suggesting a possible role in exacerbations of respiratory allergies.