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Personal sampling of small mold spores using slit cassettes
Letters to the Editor Personal sampling of small mold spores using slit cassettes To the Editor: Slit cassettes are routinely used for sampling airborne mold spores because they are inexpensive and easy to use, and analysis is simple. These are key considerations that affect development of public health standards, evaluations of personal exposures, and studies of airborne mold spores’ relationship to fungal diseases.1,2 In recent studies,3,4 slit cassettes were used at low flow rates to assess the applicability of the method for personal sampling of mold spore exposures. There is a popular assumption that smaller mold spores such as Penicillium/Aspergillus (Pen/Asp) are not effectively captured at flow rates <15 L/min (lpm). The studies were designed to test and evaluate this hypothesis. In one of these recent studies,3 slit cassettes were used for personal sampling at 5 lpm to measure exposures while mowing the lawn and during other grounds-keeping tasks. Reference samples were taken for comparison, and the results showed that Pen/Asp mold spores were well represented in both the test and reference samples. In the follow-up study,4 the performance of spore traps and slit cassettes was compared under field conditions outdoors. Sampler positions were fixed on a platform 1.5 m above ground level at a single monitoring station for consistency and repeatability. Ten-minute samples were collected with paired Allergenco D slit cassettes operated at sampling rates of 5 and 15 lpm. Samples of equal duration were also collected with Burkhard (Burkhard, Hertfordshire, UK) and Allergenco MK3 (Environmental Monitoring Systems, Inc, Charleston, SC) spore traps at 10 and 15 lpm, respectively—that is, 4 simultaneous samples were collected side by side. A sequence of five 10-minute samples was collected over a period of approximately 1 hour. Thus, 20 samples were collected per test session with 4 samplers. During 6 sampling sessions, 30 samples were collected with each sampler, totaling 120 samples for the entire study. Samples were collected outdoors over a 3month period from mid July to mid October 2009 to obtain a broad spectrum of airborne fungal spore types and sizes. Air samples were enumerated by transmission light microscopy at 3400 magnification. Total spore counts and the composition of different spore types within each sample were evaluated. Spores were classified according to genera into 19 different types of common environmental molds including Cladosporium, ascospores, basidiospores, and less plentiful spore types such as Alternaria, Epicoccum, Oidium, rusts, smuts, Pen/Asp, and so forth. Unidentified and miscellaneous spore types typically made up 3% or less of the spores and rarely extended beyond 4%. Penicillium and Aspergillus, indistinguishable by microscopy alone, were grouped and counted together as Pen/Asp. The total spore count per cubic meter and mean values were calculated separately for each of the 4 samplers to characterize and compare their performance during individual sampling sessions. Similar statistics were obtained for individual spore types to assess whether the composition of the samples differed. The relative proportion of individual spore types within each sample was determined by calculating percentages, thus allowing for meaningful comparison of their preponderance. The potential for sampler selectivity causing bias, specifically undersampling of smaller spores by operating slit cassettes at low flow rates, was investigated.
TABLE I. Spore count obtained with 4 different slit impactors operated simultaneously Sampling session
1 2 3 4 5 6
Cassette 15 lpm
Cassette 5 lpm
Burkhard 10 lpm
Allergenco 15 lpm
2,881 3,237 10,387 10,819 4,780 949
2,765 3,769 8,181 13,074 4,263 700
3,327 3,833 7,024 12,668 6,898 1,136
3,067 2,412 7,128 10,325 5,613 1,013
FIG 1. Spore composition for 1 of 6 sampling sessions obtained with 4 different slit impactors operated simultaneously.
Slit cassettes operated at the 2 different flow rates showed no systematic or significant differences between paired samples, despite a 3-fold difference in air velocities. Table I presents data for all 4 samplers and sampling sessions, showing clearly that there are no systematic or significant differences in total spore counts measured by any one of these impaction samplers when operated simultaneously side by side (Wilks l 5 0.15). These spore count data show that all 4 samplers captured mold spores equally well, and their performance was similar. Impaction is velocity-dependent; therefore, smaller mold spores with less inertia could conceivably be preferentially affected by reduced velocities.5,6 In practice, this is not the case. Field data indicate that the composition of the samples was unaffected. A compositional analysis found that results were consistent for the 2 slit cassettes operated at very different velocities and also for the Allergenco MK3 and Burkard spore traps, as shown in Fig 1. No such bias was evident. Fig 1 presents data for 1 of the sampling sessions, but all 6 sessions demonstrated the same level of consistency. In this 3dimensional graph, Cladosporium, ascospores, and basidiospores constitute the bulk of the outdoor mold spores, as expected. The remainder is represented by a broad mixture of molds, many at proportions of less than 1%. ‘‘Small’’ spores, namely Pen/Asp, were equally represented in concurrent samples obtained by all 4 methods. The percentage of Pen/Asp in these outdoor samples ranged from 2% to approximately 10% for each test session and were equally captured and detected at 5 lpm and 15 lpm by slit cassettes and by 2 other common spore-trapping methods. 1063
1064 LETTERS TO THE EDITOR
J ALLERGY CLIN IMMUNOL APRIL 2011
Likewise, collection of ascospores, basidiospores, and Cladosporium was consistent between matching samples. The results of field sampling demonstrate strong similarity between all 4 samplers in terms of overall spore capture and composition. No systematic or significant differences in spore counts were identified. Bias caused by undersampling of small spores was not detected at low flow rates using paired slit cassettes. Lower flows did not result in underrepresentative sampling by slit cassettes in terms of either the amount of total spores per cubic meter of air or the amount of small spores such as Pen/Asp, Ganoderma, and other small basidiospores.4 Simple impactors for air sampling of molds such as slit cassettes and spore traps appear to have broad cutoff points that include most common environmental mold spores. Thus, slit cassettes may be used at low flow rates of approximately 5 lpm without systematic bias for evaluating personal exposures3 and for studies of fungal-related disease. This is of practical consequence for occupational and environmental hygienists because these flows are achievable by most air-sampling pumps currently available for exposure assessments. Peter J. Pityn, PhDa James Anderson, MLTb From aOSHTECH, Inc and bLondon Health Sciences, London, Ontario, Canada. E-mail: [email protected]. Supported by OSHTECH, Inc.
Disclosure of potential conflict of interest: The authors have declared that they have no conflict of interest. REFERENCES 1. Portnoy JM, Barnes CS, Kennedy K. Sampling for indoor fungi. J Allergy Clin Immunol 2004;13:189-98. 2. Rao CY, Burge HA, Chang JC. Review of quantitative standards and guidelines for fungi in indoor air. J Air Waste Manage Assoc 1996;46:899-908. 3. Pityn PJ, Anderson JJ. Mold exposures of homeowners during typical lawn maintenance. J Allergy Clin Immunol 2009;123(suppl):S95. 4. Anderson J, Pityn PJ. Slit cassette performance for personal sampling of mold spores at low flow rates. J Allergy Clin Immunol 2010;125(suppl):AB127. 5. Marple VA, Willeke K. Impactor design. Atmos Environ 1976;10:891-6. 6. Cohen JJ, Montan DN. Theoretical considerations, design, and evaluation of a cascade impactor. Am Ind Hygiene Assoc J 1967;28:95-104. Available online December 31, 2010. doi:10.1016/j.jaci.2010.11.016
CD81 T cells are recruited early to allergen exposure sites in atopy patch test reactions in human atopic dermatitis To the Editor: Atopic dermatitis (AD) is a T-cell–mediated chronic inflammatory skin disease associated with cutaneous hyper reactivity to environmental antigens that are innocuous to normal nonatopic individuals. The house dust mites (HDM) Dermatophagoides
FIG 1. A, Summary of the 9 patients included in the study. *Positivity of the APT (according to European Task Force on Atopic Dermatitis consensus) at 9 and 48 hours as well as at 72 hours in the preselection of the study. The severity of the disease is defined as mild (<500 pg/mL) and moderate (>500 pg/mL) according to chemokine (C-C motif) ligand 17 (CCL17) serum level (**normal range, approximately 200 pg/mL). The numbers of CD81 (B) and CD31 (C) T cells/mm2 in skin in control (ctrl) or HDM APT are represented at 9 and 48 hours. D, Comparison between the number of CD81 T cells and eosinophils at 9 and 48 hours in HDM APT.
TABLE I. Spore count obtained with 4 different slit impactors operated simultaneously Sampling session
1 2 3 4 5 6
Cassette 15 lpm
Cassette 5 lpm
Burkhard 10 lpm
Allergenco 15 lpm
2,881 3,237 10,387 10,819 4,780 949
2,765 3,769 8,181 13,074 4,263 700
3,327 3,833 7,024 12,668 6,898 1,136
3,067 2,412 7,128 10,325 5,613 1,013
FIG 1. Spore composition for 1 of 6 sampling sessions obtained with 4 different slit impactors operated simultaneously.
Slit cassettes operated at the 2 different flow rates showed no systematic or significant differences between paired samples, despite a 3-fold difference in air velocities. Table I presents data for all 4 samplers and sampling sessions, showing clearly that there are no systematic or significant differences in total spore counts measured by any one of these impaction samplers when operated simultaneously side by side (Wilks l 5 0.15). These spore count data show that all 4 samplers captured mold spores equally well, and their performance was similar. Impaction is velocity-dependent; therefore, smaller mold spores with less inertia could conceivably be preferentially affected by reduced velocities.5,6 In practice, this is not the case. Field data indicate that the composition of the samples was unaffected. A compositional analysis found that results were consistent for the 2 slit cassettes operated at very different velocities and also for the Allergenco MK3 and Burkard spore traps, as shown in Fig 1. No such bias was evident. Fig 1 presents data for 1 of the sampling sessions, but all 6 sessions demonstrated the same level of consistency. In this 3dimensional graph, Cladosporium, ascospores, and basidiospores constitute the bulk of the outdoor mold spores, as expected. The remainder is represented by a broad mixture of molds, many at proportions of less than 1%. ‘‘Small’’ spores, namely Pen/Asp, were equally represented in concurrent samples obtained by all 4 methods. The percentage of Pen/Asp in these outdoor samples ranged from 2% to approximately 10% for each test session and were equally captured and detected at 5 lpm and 15 lpm by slit cassettes and by 2 other common spore-trapping methods. 1063
1064 LETTERS TO THE EDITOR
J ALLERGY CLIN IMMUNOL APRIL 2011
Likewise, collection of ascospores, basidiospores, and Cladosporium was consistent between matching samples. The results of field sampling demonstrate strong similarity between all 4 samplers in terms of overall spore capture and composition. No systematic or significant differences in spore counts were identified. Bias caused by undersampling of small spores was not detected at low flow rates using paired slit cassettes. Lower flows did not result in underrepresentative sampling by slit cassettes in terms of either the amount of total spores per cubic meter of air or the amount of small spores such as Pen/Asp, Ganoderma, and other small basidiospores.4 Simple impactors for air sampling of molds such as slit cassettes and spore traps appear to have broad cutoff points that include most common environmental mold spores. Thus, slit cassettes may be used at low flow rates of approximately 5 lpm without systematic bias for evaluating personal exposures3 and for studies of fungal-related disease. This is of practical consequence for occupational and environmental hygienists because these flows are achievable by most air-sampling pumps currently available for exposure assessments. Peter J. Pityn, PhDa James Anderson, MLTb From aOSHTECH, Inc and bLondon Health Sciences, London, Ontario, Canada. E-mail: [email protected]. Supported by OSHTECH, Inc.
Disclosure of potential conflict of interest: The authors have declared that they have no conflict of interest. REFERENCES 1. Portnoy JM, Barnes CS, Kennedy K. Sampling for indoor fungi. J Allergy Clin Immunol 2004;13:189-98. 2. Rao CY, Burge HA, Chang JC. Review of quantitative standards and guidelines for fungi in indoor air. J Air Waste Manage Assoc 1996;46:899-908. 3. Pityn PJ, Anderson JJ. Mold exposures of homeowners during typical lawn maintenance. J Allergy Clin Immunol 2009;123(suppl):S95. 4. Anderson J, Pityn PJ. Slit cassette performance for personal sampling of mold spores at low flow rates. J Allergy Clin Immunol 2010;125(suppl):AB127. 5. Marple VA, Willeke K. Impactor design. Atmos Environ 1976;10:891-6. 6. Cohen JJ, Montan DN. Theoretical considerations, design, and evaluation of a cascade impactor. Am Ind Hygiene Assoc J 1967;28:95-104. Available online December 31, 2010. doi:10.1016/j.jaci.2010.11.016
CD81 T cells are recruited early to allergen exposure sites in atopy patch test reactions in human atopic dermatitis To the Editor: Atopic dermatitis (AD) is a T-cell–mediated chronic inflammatory skin disease associated with cutaneous hyper reactivity to environmental antigens that are innocuous to normal nonatopic individuals. The house dust mites (HDM) Dermatophagoides
FIG 1. A, Summary of the 9 patients included in the study. *Positivity of the APT (according to European Task Force on Atopic Dermatitis consensus) at 9 and 48 hours as well as at 72 hours in the preselection of the study. The severity of the disease is defined as mild (<500 pg/mL) and moderate (>500 pg/mL) according to chemokine (C-C motif) ligand 17 (CCL17) serum level (**normal range, approximately 200 pg/mL). The numbers of CD81 (B) and CD31 (C) T cells/mm2 in skin in control (ctrl) or HDM APT are represented at 9 and 48 hours. D, Comparison between the number of CD81 T cells and eosinophils at 9 and 48 hours in HDM APT.