SEARCH

SEARCH BY CITATION

Keywords:

  • aerobiological data;
  • alternaria;
  • fungal spore;
  • mold allergy;
  • portable volumetric sampler

Abstract

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Sampling method
  5. Statistical data elaboration
  6. Results
  7. Discussion
  8. References

Background: Alternaria tenuis (Alt) is one of the main allergens in pediatric age. In temperate climates, airborne Alt spores are detectable from May to November with peaks in late summer and autumn. Sensitized children display symptoms even in the absence of airborne Alt spores. Alt spore concentration, as well as pollen, is usually detected by fixed devices located on the roof of a building at a height of 10–20 m. The aim of the current study is to find out whether ground-level (50 cm) Alt spore concentrations are different from those at roof-top level, even during low-concentration periods.

Methods:  Alt samples were taken simultaneously using a Hirst fixed volumetric collector (FVC) placed on a 15 m-high roof and by a portable volumetric collector (PVC). Firstly, the results of FVC and PVC, both placed on the roof-top, were compared to verify the correlation coefficient of the two samplers. Subsequently, the PVC was placed 50 cm above the ground in a courtyard (30 samplings) and in private green areas (50 samplings). The results were compared by statistical analysis (Student's t-test or K–S test).

Results:  The values of the 20 samples taken jointly in summer time (FVC 195 ± 134 spores/m3; PVC = 134 ± 131 spores/m3) showed a good correlation between the two samplers (r = 0.850; P < 0.01), with a correction factor equal to 1.177.

  • 1
    Thirty samples obtained in summer and winter when the PVC was positioned in an enclosed courtyard directly below the FVC showed no significant difference (PVC, 181 ± 194 spores/m3; FVC, 152 ± 145 spores/m3; P = 0.221).
  • 2
    Fifty samples taken by PVC placed in private green areas in a low-concentration period, showed significantly higher concentrations than by FVC: PVC, 531 ± 925 spores/m3; FVC, 25 ± 51 spores/m3 (K–S test: P < 0.0001). In particular, 33 samples taken in winter when Alt counts by FVC were <10 spores/m3 still demonstrated highly significant differences: PVC, 398 ± 961 spores/m3; FVC, 2.0 ± 2 spores/m3 (K–S test: P < 0.0001).

Conclusion:  Our results lead to the conclusion that Alt spore concentration is significantly higher at ground level in the presence of vegetation, even when the spore concentration is very low (<10 spores/m3). These results further suggest that the individual's exposure to Alt, especially in the case of children, is underestimated by samples taken at roof-top level by FVC.

Alternaria tenuis (Alt) is a ubiquitous mycete, highly resistant to adverse climatic conditions and develops readily even when humidity is minimal. Wind and dry weather seem to favor spore dispersion. In temperate climates, airborne Alt spores are detectable everywhere from May to November with peaks in late summer and autumn, especially in grain-growing areas (1, 2).

Fungi are among the most recently recognized allergens; their role in producing respiratory allergy (3) and asthma has long been established. Alt is considered one of the most important allergens in some climates with a high prevalence of sensitization during childhood (4–8). Moreover, correlation between high spore concentration and outbreaks of asthma is well documented (8–10). Sensitized patients have a high incidence of asthma (11–13) and they could also experience acute respiratory failure (14, 15).

Patients are rarely monosensitized to Alt, usually showing polysensitization to many allergens (4). Children sensitized only to Alt and Ragweed frequently display symptoms even when airborne mold spores are not detectable, or hardly so, and pollens are absent.

The aerobiological monitoring sites are usually placed on the roof-tops of high buildings. Natural exposure, especially among children, occurs closer to the ground. Differences in pollen concentrations at different heights have already been observed. Some taxa are more abundant at low levels, depending above all on the source and the size of pollen grains (16–19). Only one study, conducted in northern Europe in a climate different from ours, has also studied the distribution of spores. The study reports a greater abundance of Alt spores at ground level (with significant intercorrelation with roof-top spore counts) and a longer Alt spore period, but it did not find a significant spore count between October and April, whether at ground or roof-top level (20).

The aim of the current study is to compare the Alt spore concentrations in proximity of the ground and at high level, during both high- and low-concentration periods. We also set out to verify whether the presence or absence of vegetation can influence Alt spore values.

Material and methods

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Sampling method
  5. Statistical data elaboration
  6. Results
  7. Discussion
  8. References

As past studies have demonstrated a good correlation between a Hirst-type fixed volumetric collector (FVC) and a portable volumetric collector (PVC), we used the latter sampler to detect Alt spores close to the ground (21). The study was carried out over the period May 2000 to January 2002 in Modena, Italy, a town located in the Po Valley with a humid temperate-continental climate. The Regional Agency for Prevention and the Environment has performed aerobiologic pollen research over the last 18 years and recorded Alt spore concentrations as well. As in other temperate regions worldwide, Alt spore concentrations present seasonal variations; Alt spores are detectable from May to November, with the highest concentration being observed during summer (Fig. 1).

image

Figure 1. Trend of weekly average Alt concentrations (spores/m3) over 6 years (1995–2000) collected by the Regional Agency of Prevention and Environment of Modena (Hirst-type collector VPPS 200, Lanzoni, Bologna, Italy). On the x-axis, not all dates are reported. The highest concentrations are observed in summer and at the beginning of autumn.

Download figure to PowerPoint

We used a wind-oriented Hirst-type FVC (VPPS 2000, Lanzoni, Bologna, Italy) placed on the 15 m-high roof of the Regional Agency for Prevention and the Environment of Modena. The PVC was a PAR TRAP FA 52 sampler (Coppa, Biella, Italy). This portable pollen trap has technical characteristics comparable to those of the Hirst sampler, with constant 10 l/min suction flow and 6000 r.p.m. turbine. It is compact and small (18 × 9 × 4 cm). It is equipped with a single-use sampling chamber suitable for either personal or environmental aerobiologic and microbiologic samples (19).

Each sample was carried out simultaneously by FVC and PVC for 8 consecutive hours (from 8 am to 4 pm). In accordance with the official method for sampling and counting pollen grains as well as mold spores, the tapes from both samplers (PVC and FVC) were collected and fixed on slides with glycerinated gelatin stained with basic fuchsin. They were analyzed under a light microscope (Zeiss Axioskop, 400× magnification). The values obtained from the calculations revealed Alt spore concentration as the number of spores/cubic meter (spores/m3) of aspirated air.

Some samples were obtained during periods considered to be of low spore concentration on the basis of local data obtained over a 6-year period (1995–2000) (Fig. 1).

Sampling method

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Sampling method
  5. Statistical data elaboration
  6. Results
  7. Discussion
  8. References

With the aim of verifying the correlation between PVC and FVC, 20 samples were first taken during summer with the PVC mounted on the FVC. The PVC was fixed above the louvre of the FVC with the inlets facing in the same direction. In this way, the position and exposure to environmental parameters (including air currents) of both samplers was identical. Subsequently, two series of samples were performed with the PVC 50 cm from the ground in two different environments:

  • 1
    The PCV was placed in an enclosed courtyard, beneath the FVC. Thirty tests were performed, 20 in summer time (Alt season) and 10 in winter time (period of concentration: <10 spores/m3).
  • 2
    Fifty tests were performed in private green areas in the months when Alt concentration was lower than 10% of the annual average; of these, 33 measurements were taken in winter time, when the Alt spore count detected by FVC was less than 10 spores/m3.

Statistical data elaboration

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Sampling method
  5. Statistical data elaboration
  6. Results
  7. Discussion
  8. References

All data are reported in spores/m3. Results are expressed as means ± standard deviation (SD). For the comparison of data collected with collectors in the same position (PVC mounted on the FVC), we used Student's t-test to assess differences between means, Pearson's correlation coefficient to verify the concordance between collectors and linear regression analysis to calculate a conversion factor between collectors. The sensitivity of the two collectors was assumed to be equal for a regression analysis that was performed setting the intercept value of the regression model equal to 0, to obtain indication of the ratio between the two values. In the latter case, in order to establish the ratio between the PVC and FVC readings, the intercept value of the regression model was set at 0 based on the assumption that the two collectors were of equal sensitivity.

For the comparison of data collected in different environments, Student's t-test or the K–S nonparametric test were used, according to the distribution of data. The level of statistical significance was set at P < 0.05.

Results

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Sampling method
  5. Statistical data elaboration
  6. Results
  7. Discussion
  8. References

The mean (±SD) values of the 20 samplings of the two-paired collectors were 195 ± 134 spores/m3 in the case of PVC and 134 ±131 spores/m3 in the case of FVC (Fig. 2). The difference between the two values was not significant (P = 0.155). The Pearson correlation coefficient was 0.850, with a highly significant correlation (P < 0.01). The regression coefficient was 0.817, with the intercept value of 78.193 and the R2 value of 0.723. The multiplication ratio (conversion factor) between FVC and PVC (calculated for Alt spores) was 1.177. The residuals in both cases follow the normal curve.

image

Figure 2. Comparison between PVC and FVC samples, when the PVC was positioned close to the FVC at a height of 15 m (20 samples; on the x-axis, not all dates are reported). The findings of the two instruments are closely correlated (correlation coefficient = 0.85); the PVC–FVC conversion factor is 1.177.

Download figure to PowerPoint

The two series of samplings with the PVC 50 cm from the ground provided the following data:

  • 1
    Thirty samples collected in an enclosed courtyard below the FVC: the respective mean spore concentration values for PVC and FVC were 181 ± 194 and 152 ± 145 spores/m3 (P = 0.221). Twenty samples taken in summer time showed an average concentration for PVC and FVC, respectively, of 271 ± 180 and 228 ± 118 spores/m3 (P = 0.207). The mean values of the 10 measurements performed in winter time (when Alt spore concentration is less than 10 spores/m3) for PVC and FVC, respectively, were 2.0 ± 3.0 and 0.9 ± 1.0 spores/m3 (P = 0.313) (Fig. 3).
  • 2
    Fifty samples taken in green areas in low-concentration periods showed mean Alt concentrations for PVC and FVC, respectively, of 531 ± 925 and 25 ± 51 spores/m3 (P < 0.0001), with a ground-to-roof ratio of 21.24. In this group of samples, 33 measurements taken by PVC in winter time, when the Alt spore count detected by FVC was less than 10 spores/m3 (average concentration, 2.0 ± 2.0), showed a mean concentration of 398 ± 961 spores/m3 (P < 0.0001) (Fig. 4 and Table 1).
image

Figure 3. Comparison between PVC and FVC samples when the PVC was positioned in a walled courtyard below the FVC (30 samples; on the x-axis not all dates are reported). The values detected by the two instruments are not significantly different (P = 0.221).

Download figure to PowerPoint

image

Figure 4. Comparison between PVC and FVC samples when the PVC was positioned in green areas during a low-concentration period (Alt concentration <10% annual average) (50 samples; on the x-axis, not all dates are reported). The difference is highly significant (P < 0.0001).

Download figure to PowerPoint

Table 1.  PVC samples in green areas when FVC detected <10 spores/m3 (33 samplings). The difference is highly significant (P < 0.0001)
Day*Spore concentration†
 PVCFVC
  1. * Day/month/year.

  2. † In spores/m3.

01/02/01360
10/02/0160
26/02/0110
06/03/0100
19/03/01102
19/04/0100
08/11/01235
15/11/0124444
17/11/012938
20/11/01301
22/11/01101
25/11/0116256
26/11/012433
02/12/0119765
03/12/0146502
05/12/014252
06/12/01124
07/12/012105
08/12/01762
12/12/01103
13/12/013495
19/12/012402
24/12/01931
28/12/01250
30/12/0170
04/01/0231
05/01/0221
07/01/021821
12/01/02591
13/01/02181
14/01/02161
15/01/02320
17/01/02120

Discussion

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Sampling method
  5. Statistical data elaboration
  6. Results
  7. Discussion
  8. References

The comparison between FVC and PVC samplers positioned in the same place confirmed a highly significant correlation (P < 0.01), as previously demonstrated for certain pollens (21). In our work, the computation of a conversion factor allows for the exact comparison of the two sampling methods.

Aerobiologic data represent an important reference element for allergists and pulmonologists in interpreting the causes of symptoms and deciding on the preventive/therapeutic measures to be taken with allergic patients.

Some surveys have underlined the fact that sampling height influences pollen count (17–19). Indeed, it stands to reason that the number of mold allergens growing on dead vegetation and rotting leaves on the ground may well be underestimated by samplings taken by an FVC usually placed at roof level. This phenomenon is more probable in the case of Alt spores, which are one of the largest spores in common outdoor fungi.

Only one previous study has researched both pollen and spores; the data reported in that study would suggest that Alt spores are more abundant at ground level during the Alt season and that outside this period, Alt spores are absent both at roof and ground level (20). In our study, on the other hand, we demonstrate the presence of high Alt concentrations in gardens in the winter months as well. This difference is probably due to the fact that the previous study was conducted in northern Europe, where the ground is covered by snow and ice for many months of the year, whereas our study was carried out in a temperate region of light snowfalls and short-lived rigid winter temperatures.

In our study, Alt spore concentration close to the ground was greatly influenced by the type of environment examined. In green areas, even if in a low-concentration period, the difference between Alt spore concentrations at 15 and 50 cm was highly significant. In the walled courtyard, on the other hand, there was no significant difference in concentration, regardless of the period during which collection took place.

The general opinion is that mycete sporulation occurs in mild months, but we observed that it takes place in winter time, too. For instance, in this season, there were four readings of over 1000 spores/m3 in the green areas, when the aerobiologic data showed only 2 spores/m3, even the highest value near the ground (4650 spores/m3) actually occurred in December (Fig. 4 and Table 1).

It is well known that Alt survives even at extreme temperatures by protecting itself in rotting leaves and other biomaterial. In summer and autumn, under certain weather conditions (mild temperature, a dry period following a humid period, increase in light), Alt may flourish (1, 2), and even in winter, some spot weather changes can trigger spore release. We did not perform sampling in green areas during the Alt season because to demonstrate high exposure during a well-known risk period would have no clinical relevance.

In the present study, we observed great variability in spore concentration in just a few days. This phenomenon is well known and may depend only in part on different garden environments. Many aerobiologic studies have shown that fungal spore concentrations can suddenly change from one day to another.

Our findings confirm the need for aerobiologic measurements that better reflect an individual's actual exposure, so as to gain further insight into the relationship between exposure and allergic symptoms.

The study cited above (18) shows that, at ground level, the increase in the concentrations of some pollens and spores at the beginning of the Alt season occurs earlier, while the corresponding decrease at the end of the season occurs later; as a consequence, the exposure period is longer than is generally supposed. And since airborne allergens can be present in the air for many days before the detection and after the disappearance of airborne spores, as demonstrated by the radioimmunoassay (RIA) method, the exposure period is even longer (10, 22). However, a prolonged Alt season cannot account for the perennial pattern of allergic symptoms of many sensitized patients. Exposure may also occur within home, although studies of indoor environments using volumetric collector samplers, or even dish cultures, are not conclusive (23–26).

In our experience, perennial allergic symptoms in sensitized individuals can be explained by the exposure to high Alt concentrations near to the soil. We demonstrate, in fact, that even during the lowest Alt period, people can be exposed to this allergen in green areas. This is a particularly important consideration in the case of children, who most often play close to the ground. In this environment, individual exposure is constant throughout the year, with the possible exception of brief periods of intense cold when snow and ice cover the ground. The parents of mold-sensitized children should be advised against the risk of allergen exposure when the children play in green areas in both hot and cold periods of the year and especially in the warmest hours of the day (1).

Our study underlines the fact that airborne particle concentrations, of Alt spores in particular, as detected by conventional samplings, do not reflect the actual levels of exposure. In future, the microenvironment in which Alt-allergy sufferers live should be monitored using a mobile personal collector located in the most frequented areas or, preferably, worn on the person.

References

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Sampling method
  5. Statistical data elaboration
  6. Results
  7. Discussion
  8. References
  • 1
    Rotem J. The genus Alternaria: biology, epidemiology and pathogenecity. The American Phytopathological Society, 1994.
  • 2
    Solomon WR. Common pollen and fungus allergens. In: BiermanCV, PearlmanDS, editors. Allergic disease of infancy, childhood and adolescence. Allergic disease of infancy, childhood and adolescence. Philadelphia, PA: WB Saunders Company, 1980: 219225.
  • 3
    Johansson SGO, O'B Hourihane J, Bousquet J, Bruijnzeel-Koomen C, Dreborg S, Haahtela T et al. A revised nomenclature for allergy. An EAACI position statement from the EAACI nomenclature task force. Allergy, 2001;56: 813824.
  • 4
    D'Amato G, Chatzigeorgiou G, Corsico R, Gioulekas D, Jager L, Jager S et al. Evaluation of the prevalence of skin prick test positivity to Alternaria and Cladosporium in patients with suspected respiratory allergy. An European multicenter study promoted by the Subcommittee on Aerobiology and Environmental Aspect of Inhalant Allergens of the European Academy of Allergology and Clinical Immunology. Allergy 1997;52: 711716.
  • 5
    Corsico R, Cinti B, Feliziani V, Gallesio MT, Liccardi G, Loreti A et al. Prevalence of sensitization to Alternaria in allergic patients in Italy. Ann Allergy Asthma Immunol 1998;80: 7176.
  • 6
    Halonen M, Stern DA, Wright AL, Taussig LM, Martinez FD. Alternaria as a major allergen for asthma in children raised in a desert environment. Am J Respir Crit Care Med 1997;155: 13561361.
  • 7
    Kauffman HF, Tomee JFC, Van Der Werf TS, De Monchy JGR, Koëter GK. Review of fungus-induced asthmatic reactions. Am J Respir Crit Care Med 1995;151: 21092116.
  • 8
    Downs SH, Mitakakis TZ, Marks GB, Car NG, Belousova EG, Leüppi JD et al. Clinical importance of Alternaria exposure in children. Am J Respir Crit Care Med 2001;3: 455459.
  • 9
    Targonski PV, Persky VW, Ramakrishnan V. Effect of enviromental moulds on risk of death from asthma during the pollen season. J Allergy Clin Immunol 1995;95: 955961.
  • 10
    O'Hollaren M, Yunginger JW, Offord DJ, Somers MJ, O'Connell EJ, Ballard DJ. Exposure to aeroallergens as a possible precipitating factor in respitatory arrest in young patients with asthma. N Engl J Med 1991;324: 359363.
  • 11
    Tariq SM, Matthews SM, Stevens M, Hakim EA. Sensitization to Alternaria and Cladosporum by the age of 4 years. Clin Exp Allergy 1996;26: 794798.
  • 12
    Peat JK, Tovey E, Mellis CM, Leeder SR, Woolcock AJ. Importance of house dust mite and Alternaria allergens in childhood asthma: an epidemiological study in two climatic regions of Australia. Clin Exp Allergy 1993;23: 812820.
  • 13
    Perzanowski MS, Sporik R, Squillace SP, Gelber LE, Call RM, Carter M et al. Association of sensitization to Alternaria allergens with asthma among school-age children. J Allergy Clin Immunol 1998;101: 626632.
  • 14
    Neukirch C, Henry C, Leynaert B, Liard R, Bousquet J, Neukirch F. Is sensitization to Alternaria alternata a risk factor for severe asthma?A population-based study. J Allergy Clin Immunol 1999;103: 709711.
  • 15
    Black PN, Udy AA, Brodie SM. Sensitivity to fungal allergens is a risk factor for life-threatening asthma. Allergy 2000;55: 501504.
  • 16
    Gregory PH. The air-spora near the earth's surface. In: GregoryPH, editor. The microbiology of the atmosphere, 2nd edn. New York: Hill L, 1973: 108121.
  • 17
    Alcazar P, Galan C, Carinanos P, Dominguez-Vilches E. Effects of sampling height and climatic condition in aeribiological studies. Invest Allergol Clin Immunol 1999;9: 253261.
  • 18
    Fiorina A, Mincarini M, Sivori M, Brichetto L, Scordamaglia A, Canonica GW. Aeropollinic sampling at three different heights by personal volumetric collector (Partrap FA 52). Allergy 1999;54: 13091315.
  • 19
    Thibaudon M, Sulmont G. The influence of the height position of two captors at Amiens. Allergie Immunol 2002;24: 169171.
  • 20
    Rantio-Lehtimaki A, Koivikko A, Kupias R, Makinen Y, Pohjola A. Significance of sampling height of airborne particles for aerobiological information. Allergy 1991;46: 6876.
  • 21
    Fiorina A, Scordamaglia M, Mincarini L, Fregonese L, Canonica GW. Aerobiologic particle sampling by a new personal collector (Partrap FA52) in comparison to the Hirst (Burkard) sampler. Allergy 1997;52: 10261030.
  • 22
    Barnes C, Schreiber K, Pacheco F, Landuyt J, Hu F, Portnoy J. Comparison of outdoor allergenic particles and allergen levels. Ann Allergy Asthma Immunol 2000;84: 4754.
  • 23
    Ren P, Jankun TM, Belanger K, Bracken MB, Leaderer BP. The relation between fungal propagules in indoor air and home characteristics. Allergy 2001;56: 419424.
  • 24
    Verhoeff AP, Van Wijnen JH, Brunekreef B, Fischer P, Van Reenen-Hoekstra ES, Samson RA. Presence of viable mould propagules in indoor air in relation to house damp and outdoor air. Allergy 1992;47: 8391.
  • 25
    Verhoeff AP, Van Wijnen JH, Brunekreef B, Fischer P, Van Reenen-Hoekstra ES, Samson RA. Fungal propagules in house dust II. Relation with residential characteristic and respiratory symptoms. Allergy 1994;49: 540547.
  • 26
    Katz Y, Verleger H, Barr J, Rachmiel M, Kivitis S, Kuttin ES. Indoor survey of moulds and prevalence of mould atopy in Israel. Clin Exp Allergy 1999;29: 186192.