Airborne Aspergillus fumigatus levels outside and within a large clinical center

Airborne Aspergilhs fumigatus levels outside and within a large clinical center William Ann Arbor,

‘Il. Solomon,

M.D., Harriet

P. Burge, Ph.D., and Jean R. Boise, B.S.

Mich.

Most considerations of Aspergillus fumigatus prevalence have implied that patterns of occurrence observed within London hospitals are generally applicable. Since prevalence data are almost nonexistent elsewhere, this assumption remains untested. To provide a comparison relevant to North America, we have monitored thermotolerant fungi outside as well as at two sites within the University Hospital, Ann Arbor, Michigan, during one year. Collections were made with paired Andersen samplers and malt agar for 30- to 40-min periods in a hallway adjacent to 6W, a general medical ward (47 days), and 2W, a lower level service and supply area (40 days); in addition, IO-min outdoor samples (44 days) were taken on an unobstructed hospital rooftop (out). Recoveries were analyzed after 3 and 7 days of 3P C aerobic incubation. Virtually complete suppression of Cladosporium form species at 37” left a mycofora with A. fumigatus, A. niger, Paecilomyces spp., Mucor spp., and yeast/bacteria predominating. Although the proportions of samples yielding A. fumigatus were 76% for 6W, 57% for 2W, and 56% (out), levels exceeded 40 isolatesim3 only twice and were over 10 isolates/m3 on only 10 of 131 total samples. For 6W, 2W and out, respectively, means were 4.78, I .97, and 6.25 isolateslm3; medians were I .20, I .05 and I .75/m” without annual trends indoors and with only a limited outdoor summer increase. Our data fail entirely to show the fall-winter abundance observed in the London report and suggest substantially lower indoor exposure levels of A. fumigatus than those noted in London.

Bronchial carriage of aspergilli, especially Aspergillus fumigatus, is now a well-accepted source of pulmonary infiltrates and eosinophilia among asthmatic subjects.’ Although this syndrome is observed widely, its regional prevalence has appeared to vary markedly,2-4 with North American rates often well below those reported from Great Britain and Western Europe. Local differences in clinical recognition and data processing have been invoked to explain these disparities; however, regional variations in human exposure to the fungus have not been excluded. In fact, relatively few reports of volumetric data systematically describing the prevalence of this organism in free air have been published.5-g In the most recent From the Section of Allergy, Department of Internal Medicine, University of Michigan Medical School. Supportedby ResearchGrant AI 10181from the National Institute of Allergy and Infectious Diseases,National Institute of Health, United StatesPublic Health Service. Received for publication Jan. 2 I, 1978. Accepted for publication April 14, 1978. Reprint requests to: William R. Solomon, M.D., Department of Internal Medicine, Section of Allergy, D3138 N. Outpatient Building, University of Michigan Medical School, Ann Arbor, Mich. 48 109. Vol. 62, No. 1, pp. 56-60

studies authors have implied that the seasonal patterns and levels of prevalence described by Noble and ClaytonlO are obtained generally in temperate regions. However, since that report was based upon indoor recoveries (in one large British hospital during a single year), such broad assumptions are not easily justified. The present study was begun to extend this limited data base and to increase its relevance for North American clinicians. In addition, we have striven to generate results to compare with those previously published by British workers. MATERIALS

AND METHODS

During the period from April, 1975, through May, 1976, regular collections of viable thermotolerant fungi were made with the use of paired, upright, Andersen samplers,” model 010 1, (2000 Inc., Atlanta, Ga.) at two points within the University Hospital, Ann Arbor, Michigan. One of these locations, 6 West (6W), was a hallway which gave accessto semiprivate moms occupied by patients of a general medical service, as well as to a service elevator and clothing lockers of hospital personnel. The corridor chosen as the second sampling point, 2 West (2W), served diverse support activities including repair shops and uniform serOOSI-674917810162-0056$00.50/001978

The C.V. Mosby

Co.

VOLUME NUMBER

Airborne Aspergillus

62 1

I. Consistency of collection ratios for specific taxa at 37” C

TABLE

(i.e., percent

Consistency

6 West Taxon

or type

Arthrospores* Aureobasidium Aspergillus (total) A. cervinus A. javipes A. flavus A. fumigatus A. glaucus A. niger A. sylvaticus A. terreus A. ustus A. versicolor Chaetomium Coniothyrium Monilia Mucor

Mycotypha Paecilomyces Penicillium Sporotrichum Streptomyces Trichoderma YeasVbacteriat Other $

(93)

of plates providing

of recovery

2nd level (73)

4 92 1 16 74 1 65 1

recoveries)

85 1 1 10 62 49

6 West 2 West

57

and frequency

Frequency Outside (44)

levels

fumigatus

ratios

2 West Outdoor

x 100

6 West Outdoor 36

II 2 62

108

137

148

52

100 160 119

119

142

20

133

245

325

1 3 2

20 3 45 17 1

80 29

100

2 3 1 3 15

5 16

133

94

125

46 27

11 16

98 63

418 169

409 169

1 5 78 32

39 29

103 91

200 110

205 100

Numbers in parenthesesare total samples. *Includes types commonly assignedto Geotrichum and conidial stagesof some basidiomycetes. tDetermined macroscopically; no attempt was made to distinguish these diverse types. #Includes nonsporulating tilamentous fungi. vices besides leading onto the main hospital loading dock. Both areas were heavily travelled and neither was specially ventilated. Carefully timed sampling periods of 30- to 60-min duration were used so that recoveries were derived from 1.7- to 3.4-m3 pottions of air. In addition, after June 1975, collections were obtained atop the Kresge Medical Research Bldg. adjacent to the hospital with the use of an Andersen sampler wind-oriented on the vane of a Burkard spore trap (Burkard Mfg. Co., Ltd.). This rooftop site (out), 50’ above grade, was completely open to the south, west, and north, although a low elevator housing was present to the east. Outdoor collections were of IO- to 20-min duration, corresponding to 0.28 to 0.56 m3 of sampled air. Initially, either 6 West or 2 West as well as the outdoor station was studied on any given morning; during the last 8 mo, however, sampling was carried out at all three points on specific dates. Each site was studied at approximate 8-day intervals resulting in a total of 47 samples for 6W, 40 for 2W, and 44 outside. All collections were made between 0830 and 1130 A.M. For much of the study, integrated samples were obtained with a single, flat-bottomed Petri

plate (designed for the Andersen sampler) positioned beneath the sixth stage12; an additional plate was used below stage 3 for indoor collections during 1976. Recoveries were made on malt extract agar (malt extract, 32 gm; yeast extract, 1 gm; agar, 16 gm; distilled water, to 1 L). Following exposure, plates were incubated at 37” in high humidity for 7 days. Colonies were identified macroscopically or at scanning power where possible. Form species of aspergilli and additional taxa were identified from slide cultures with standard references.‘“~ I4

RESULTS Incubation of samples at 37” produced almost complete suppression of familiar dematiaceous fungi leaving a thermotolerant mycoflora in which A. fumigatus was the best-represented filamentous form. Table I lists the taxa recovered overall and indicates the regularity of isolation (i.e., percent of total samples yielding recoveries) for each of the three study sites as well as comparative capture ratios. Of the

58

Solomon,

Burge, and Boise

TABLE II. Mean, median,

J. ALLERGY

and peak levels for the 5 most common Concentration

6 West

types recovered

in culture

CLIN.

IMMUNOL. JULY 1979

at 37” C

(isolates/m3)

2 West

Outdoor

Taxon and type

Mean

Median

Peak

Mean

Median

Peak

Mean

Median

Peak

A. jiimigatus A. niger Mucor spp. Paecilomycesspp.

5.2 1.3 0.3

Yeast/bacteria

7.9

1.2 0.9 0 0.3 4.3

123.0 10.6 2.9 28.8 48.3

2.0 2.4 0.2 2.7 7.2

1.2 0.9 0 0.6 5.6

8.2 21.8 4.1 38.3 30.5

6.3 1.0 0.5 0.5 3.1

3.5 0 0 0 0

67.0 7.1 3.5 7.1 35.0

2.0

most prevalent types, Aspergillus jlavus, A. niger, Paecilomyces spp., and yeast/bacteria each was taken two or more times as frequently from indoor samples as at the rooftop site. Differences in rates of recovery between 6W and 2W were less than 2-fold,* with total Penicillium showing the highest (namely, 1.6) 6W/2W ratio. All taxa listed in Table I except Mycotypha have been recovered locally during several previous years of outdoor sampling. Prevalence data for the five most abundantly recovered types are shown in Table II. The 6W and 2W values listed are derived from mean recoveries of 47 and 40 sampling occasions, respectively, by paired samplers. Where stage 3 and stage 6 plates were available, their individual recoveries were summed for this purpose. Spores/m3 exceed 50 (A. jiimigatus 123/M3 (6W), 67/M3 [out]) on only two occasions, levels remaining below 10 isolates/m3 for the 5 taxa. Mean levels of A. fumigatus did not differ substantially among the three study sites, although 2W tended to provide the sparsest recoveries of this form species. An analysis of paired 6W and outside data points for specific days using the Wilcoxon signed rank test supported the null hypothesis, i.e., that the members of these pairs come from populations having the same median values (dp] > 0.1). Fig. 1, which displays all data for A. fumigatus, illustrates these features and shows, in addition, the lack of seasonal prevalence trends for indoor levels and the very slight increase in outdoor levels during the summer months. Results of 6W exposures are shown in Fig. 2 against a logarithmic plot of indoor A. fumigatus levels reported by Noble and Clayton1o from London to further emphasize the contrasting seasonal patterns implicit in these two sets of data. For the most prevalent thermotolerant types, Table III shows the average distribution of recoveries between the third and sixth stage culture plates for 32 collection periods. With this crude two-stage parti-

*For taxa recovered on at least 10% of samples at one site.

tion, significant differences in deposition were evident for each of the categories studied. Over 80% of Aspergillus niger and yeast/bacteria isolations occurred at stage 3, while a clear majority of A. fumigatus and Paecilomyces spp. colonies appeared on the sixth stage plates. DISCUSSION Our results indicate low levels of A. fumigatus within and about the University Hospital throughout the year. This finding contrasts with indoor data of Noble and Clayton which showed a definite increase lasting approximately 3 mo with peak levels of this organism over 1,000 times higher than those in Ann Arbor. Because the findings of Noble and Clayton ( 1963) are invoked so widely and are based on indoor, nosocomial exposures, every effort was made to provide a comparable sampling sequence as part of the present investigation. In each of the studies, relatively large sample volumes were possible, assuring that days totally without A. fumigatus recoveries would be minimal. However, one can assume also that prevailing particle levels were somewhat underestimated at both sampling stations because of inconsistent spore viability and potentially suboptimal growth conditions. The validity of studying integrated collections, secured on single, sixth stage Andersen plates has been demonstrated by direct comparison with 6-plate arrays. I2 Overloading, the major risk confronting this modified use of the Andersen sampler, was virtually never encountered in the present study. Our data describing the partition of recoveries between third and sixth stage plates suggest that a majority of A. fumigatus propagules have the dimensions of single spores, confirming a previous report. lo Paecilomyces spp. recoveries also are typical of single spore units. By contrast, the predominant capture of A. niger and yeast/bacteria at stage 3 suggests airborne spore aggregates and/or association of viably units with debris or “rafts,” including epidermal scales.‘5

VOLUME NUMBER

Aspergillus

Airborne

62 1

i I ! I I I

APRIL

MAY

JUNE

JULY

AUG

SEPT

OCT

DEC

NOV

;

fumigatus

Asperqillus fumigotus Isobtes -6W . . . . . . . . . . .pw

levels

59

/ M3

0-----Outside

JAN

FEB

MARCH

APRIL

MAY

1975 ‘I976

FIG. 1. Aspergihs span.

Each

point

fumigatus is derived

lo.000.0 r

recoveries at sites outside and from a single collection period.

Asper$Ilus

within

the University

Hospital

during

a 14-mo

fumigatus

-

6 West El Noble 8 Cloyton (1963)

Mar 6pr MO; Jun FIG. 2. Aspergillus repotted logarithmic

July

Aug Sept Dct Nov Dee Jon Feb Mbr Apr hiay

fumigatus

by Noble and ordinate.

prevalence (o---o) at the 6 West site compared with Clayton,10 indicated by the stippled area. Both sets of data

The occurrence of A. fumigatus in a large majority of our study samples accords well with the reported regularity of occurrence of this organism in other areas.5-8* “3 l6 Our data also emphasize the utility of 37” incubation for suppressing growth of ubiquitous dark-spored hyphomycetes-especially Cludosporium form species. Despite these favorable isolation conditions, A. fumigatus recoveries in the University Hospital in Ann Arbor suggested airborne levels substantially below those noted by Noble and Claytonlo

indoor levels previously have been adapted to a

and lacking the strong seasonal trends evident in their report and subsequent data.16 Concentrations recently observed at Cardiff by Mullins and co-workers” approximated our own results, although a winter peak of prevalence also was evident in the Welsh data. There are, a priori, no bases for assuming that the levels associated with the University Hospital are characteristic of other points in our region. However, winter volumetric collections previously obtained in over 150 local homes” as well as university libraries

60 Solomon, Burge, and Boise

J. ALLERGY

CLIN. IMMUNOL. JULY

TABLE III. Differential recoveries at third and sixth Andersen sampler stages for 4 microbial categories

Taxon

Yeast/bacteria A. fumigatus A. niger Paecilomyces

spp

Total recoveries

3rd stage

6th stage

813 123 142 80

83 16 87 30

17 84 13 70

Y. recovery

Significance level

0.01 0.01

0.01 0.05

and animal care facilities9 have produced comparably low recoveries of A. fumigatus. Similar reservations must arise in attempting to apply the data of Noble and Clayton to increasingly remote enclosed spaces and free air stations. The possibility that A. fumigatus levels encountered by these workers were derived from discrete intramural sources is not easily dismissed. Examples of contamination of hospital ventilating systems by thermotolerant aspergilli have been suggested1*-21 and respiratory shedding of viable fungus particles by an apparently well individual22 documented. Where internal heating components are colonized, local thermal and aerodynamic factors might be expected to lead to spore prevalence peaks during colder months. Despite these uncertainties, the observations presented here reinforce the view that differences in A. fumigatus prevalence may, in fact, exist among temperate regions. Several reports have suggested a selective abundance of this species in areas enjoying maritime conditions rather than continental climates.16’ 23 In addition, the 8% rate of A. fumigatus isolation from initial cultures of asthmatic sputa, reported by Pepys and associates26 implies substantial exposure (although carriage by other respiratory disease populations2’ must be studied for purposes of comparison). Differences in the intensity and consistency of specific exposure may be reflected directly in the rates of overt Aspergillus-induced illness among susceptible groups. The development of a synoptic picture of A. fumigutus prevalence, now technically feasible, appears to merit a concerted, global effort. REFERENCES I. Pepys, J.: Hypersensitivity diseases of the lungs due to fungi and organic dusts, Monographs in Allergy, vol. 4, Basel, 1969, S. Karger AG. 2. Slavin, R. G.: Allergic bronchopulmonary aspergillosis-a North American rarity, Am. J. Med. 47~306, 1969. 3. Hoehne, J. H.: Allergic bronchopulmonary aspergillosis is not rare, Chest 63~177, 1973. 4. Novey, H. S.: Incidence of certain allergic lung disease in California, Western J. Med. 122~491, 1975.

1978

5. Mullins, J., Harvey, R., and Seaton, A.: Sources and incidence of airborne Aspergillusfumigarus (Fres.), Clin. Allergy 6:209, 1976. 6. Evans, H. C.: Thermophilous fungi isolated from the air, Trans. Br. Mycol. Sot. 59:516, 1972. 7. Hudson, H. J.: Aspergilli in the air spora at Cambridge, Trans. Br. Mycol. Sot. 52~153, 1969. 8. Hudson, H. J.: Thermophilous and thermotolerant fungi in the air spora at Cambridge, Trans. Br. Mycol. Sot. 60:596, 1973. 9. Solomon, W. R., and Burge, H. P.: Aspergillus fumigatus levels in and out of doors in urban air, J. ALLERGY CLIN. IMMUN~L. 55~90, 1975. IO. Noble, W. C., and Clayton, Y. M.: Fungi in the air of hospital wards, J. Gen. Microbial. 32397, 1963. I 1. Andersen, A. A. : New sampler for the collection, sizing and enumeration of viable airborne particles, J. Bacterial. 76~47 I, 1958. 12. Solomon, W. R., and Gilliam, J. A.: A simplified application of the Andersen sampler to the study of airborne fungus particles, J. ALLERGY 45: 1, 1970. 13. Barnett, H. L., and Hunter, B. B.: Illustrated genera of imperfect fungi, ed. 3, Minneapolis, 1972, Burgess Publ. Co. 14. Raper, K. B., and Fennell, D. I.: The Genus Aspergillus, Baltimore, 1965. The Williams & Wilkins Co. 15. Davies, R. R., and Noble, W. C.: Dispersal of bacteria on desquamated skin, Lancet 2: 1295, 1962. 16. Mallea, M., Murray, I. G., Segretain, G., Philpot, C. M., Charpin, H., Gueho, E., and Charpin, J. : Census of Aspergil/us colonies in the air comparison between London, Paris, Lyon, Marseilles, Acta Allergol. 27:273, 1972. 17. Solomon, W. R.: A volumetric study of winter fungus prevalence in the air of midwestern homes, J. ALLERGY CLIN. IMMUNOL.5246, 1976. 18. Aisner, J., Schimpff, S. C., Bennett, J. E., Young, V. M., and Wiernik, P. H.: Aspergillus infections in cancer patients. Association with fireproofing materials in a new hospital, J. A. M. A. 235:411, 1976. 19. Rose, H. D.: Mechanical control of hospital ventilation and Aspergillus infections, Am. Rev. Resp. Dis. 103306, 1972. 20. Rosen, P. P., and Stemberg, S. S.: Decreased frequency of aspergillosis and mucormycosis, N. Engl. J. Med. 295: 13 19, 1976. 21. Gage, A. A., Dean, D. C., Schimert, G., and Minsley, N.: Aspergihs infection after cardiac surgery, Arch. Surg. 101:384, 1974. 22. Staib, F.: Aspergillus fimigatus in der Ausatmungsluft eines Arztes, Dtsch. Med. Wochenschr. 99: 1804, 1974. 23. Colen, J., and Van Arsdel, P. P.: Molds of allergenic significance in the Puget Sound area, Ann. Allergy 19: 1399, 1961. 24. Kramer, C. L., Pady, S. M., and Rogerson, C. T.: Kansas aeromycology. V. Penicillium and Aspergillus, Mycologia 52:545, 1960. 25. Pady, S. M., and Kapica, L.: Fungi in air masses over Montreal during I950 and I95 1, Can. J. Bot. 34: I, 1956. 26. Pepys, J., Riddell, R. M., Citron, K. M., Clayton, Y. M., and Short, E. I.: Clinical and immunological significance of Aspergillus fumigatus in the sputum, Am. Rev. Respir. Dis. 80~167, 1959. 27. Louridas, G.: Bronchopulmonary aspergillosis. An epidemiological study in a hospital population, Respiration 33:281, 1976.