FUNGAL COLONIZATION OF AIR-CONDITIONING SYSTEMS

Fungi have been implicated as quantitatively the most important bioaerosol component of indoor air associated with contaminated air-conditioning systems. rarely, indoor fungi may cause human infections, but more commonly allergenic responses ranging from pneumonitis to asthma-like symptoms. From all air conditioner filters analyzed, 16 fungal taxa were isolated and identified. Aspergillus fumigatus causes more lethal infections worldwide than any other mold. Air-conditioning filters that adsorb moisture and volatile organics appear to provide suitable substrates for fungal colonization. It is important to stress that fungal colonization of air-conditioning systems should not be ignored, especially in hospital environments.

organ transplantation, or any other medical reason, may lead to the uncontrolled multiplication of fungi and consequent onset of infection, sometimes with fatal effects (P e r d e l i et al., 2006).there are reports that four patients died and 11 contracted the respiratory disease aspergillosis at the Alcala de Hanares Hospital, near Madrid (Spain).the disease was caused by inhaling the spores of Aspergillus fumigatus, which was later detected in the hospital's air-conditioning system (http://www.acr-news.com/news)

Samples studied
Fifteen dust samples were collected from airconditioning filters from school classrooms and offices.Also, five swab samples were collected from an air conditioner in a hospital surgical ward.none of the analyzed filters from classrooms and offices had been removed or cleaned for two years (Fig. 1a), whereas those from the hospital were cleaned once a month on a regular basis.

Direct examination
the tape lift technique used for direct examination allows for immediate determination of the presence of fungal spores and identification of the types of fungi present.direct examinations should only be used to sample visible mold growth in contaminated air-conditioning filters.Samples were collected by pulling the tape of the filter surface with slow steady pressure, holding only the tape edges, after which they were put on slides for light microscopy.

Determination of total spore concentrations in dust
culturable fungal spore concentrations are presented in terms of colony-forming units (cFu)/g of dust.Sub-samples (0.5 g) were taken from each dust sample and suspended in distilled water (0.0425 g/l KH2Po4, 0.25 g/l MgSo4, 0.008 g/l naoH, 0.02% tween 80 detergent).dilution series were prepared and three succesive dilutions were plated in triplicate on malt agar medium (MA) with the antibiotic streptomycin, which was added during the preparation process in order to prevent bacterial growth (P a s a n e n et al., 1997).the plates were incubated on 22 ± 2ºc and read after 72 to 120 hours.Fungal colonies formed on the medium were identified on the basis of both macroscopic and microscopic characteristics of each isolated colony using identification keys (A i n s w o r t h et al., 1973; A r x , 1974; E l l i s and E l l i s , 1997; P i t t , 1979; r a p e r , and F e n n e l 1965).
rESuLtS And dIScuSSIon the direct examination method revealed the presence of different fungal structures: conidia, conidiophores, chlamidospores and mycelia (Fig. 1bf).From all dust samples analyzed from classrooms, offices, and hospital air-conditionning systems, six fungal genera with different numbers of species were recorded: Cladosporium, Penicillium, Aspergillus, Alternaria, Epiccocum, and one Ascomycotyna from the order Sphaeriales (table 1). the fungal colonies isolated from different sources were characterized not only by the presence of different species, but also by their different abundance (Fig. 2a). the genus Aspergillus with five species was the most frequent (tabel 1, Fig. 2b, e, f). the abundance of fungal colonies was much higher in classrooms and offices than in the hospital.Aspergillus and Penicillium species were dominant.Alternaria alternata and Cladosporium cladosporioides were the most frequent dematiaceous fungi (Fig. 2c, d).Fungal growth was quantified by the number of cFus.culturable spore concentrations in 15 dust samples varied from 10 4 to 10 6 cFu/g.In central Finland, culturable and total fungi in dust accumulated in air ducts in single-family houses varied from 10 4 to 10 8 cFu/g (P a s a n e n et al., 1997).
the results of this research confirmed previous findings that air-conditioning systems are highly linked with fungal pollution of indoor air. the diversity and abundance of fungal species isolated from different air-conditioning systems can be attributed to different ways of maintaining the systems themselves.Greater numbers of fungal genera and species with much higher colony abundance were expected and found in samples isolated from the airconditioning systems from classrooms and offices since these systems were not cleaned after installa- tion (Fig. 1a).only two fungal species were found in samples from the hospital units, with very low colony abundance.this result is worrying because one of the two isolated species was Aspergillus versicolor (Fig. 2e).Aspergillus is a large genus of fungal molds, of which only a few species cause human infections, most commonly A. fumigatus.Spores from these species are widespread in the environment, occurring in soil, in dust, and in outdoor and indoor air.Known as aspergillosus, fungal infection with Aspergillus ranges from the benign to the fatal.Healthy individuals usually inhale Aspergillus spores without any untoward effects, but sometimes contract relatively benign infections of the lungs and sinuses.In susceptible compromised patients, however, inhalation leads to multiplication of the fungus in the lungs and subsequent invasive infection that may spread to any organ of the body.dissemination to the brain, gastrointestinal tract, and other organs occurs in up to 30% of cases.the disseminated infection is usually fatal, partly because early diagnosis is difficult and treatment often ineffective (F r e n c h , 2005).carpet dust samples (n = 11) contained A. versicolor in concentrations ranging from <2.5 × 10 1 to 3.6 × 10 5 (median, 3.1 × 10 4 ) cFu/g of dust, and the median proportion of A. versicolor in total culturable fungi was 18%.Based on thin-layer chromatography detection of sterigmatocystin, 49 of 50 A. versicolor isolates (98%) where found to be toxigenic in vitro (E n g e l h a r t et al., 2002).Many Aspergillus species are well known as potential producers of mycotoxins and other volatile harmful compounds, and many of them can cause aspergillosis in humans.Hospitals and other medical facilities are places were patients with damaged immune systems are commonly found.these patients are very receptive to fungal infections.In order to avoid unwanted fungal infections in hospitals, the air-conditioning systems must be subjected to regular maintenance in order to reduce potential fungal pollution.three features of mold biochemistry are of special interest from the standpoint of human health.Molds contain glucan, a compound with inflammatory properties.Spores and mycelial fragments contain allergens (G o r n y et al., 2002).the spores of some species contain low-molecular-weight chemicals that are cytotoxic or have other toxic properties.Some molds, such as A. fumigatus, can cause opportunistic infection in immunocompromised and healthy individuals and severe allergic diseases, such as asthma or cystic fibrosis (Burge, 2000).In our previous investigations, Aspergillus species were recorded in different substrata.Fungal spores can spread from different sources and contaminate air conditioning filters (K a t a r a n o v s k i et al., 2007; ; L j a l j e v i ć and Vu k o j e v i ć , 1997).Микрогљиве, као квантитативно најзначајније биоаеросолне компоненте ваздуха у затвореним просторијама, су чести контаминанти клима уређаја.Оне могу изазвати алергијске реакције типа пнеумонитиса и симптоме сличне астми, а ређе могу бити и изазивачи хуманих инфекција.Из анализираних клима уређаја изоловано је и идентификовано 16 таксона микрогљива.Изо-ловани Aspergillus fumigatus је познат коо изазивач леталних инфекција широм света.Филтери клима уређаја који апсорбују влагу и органске материје су погодан супстрат за колонизацију микрогљивама.Веома је значајно напоменути да се колонизација клима уређаја микрогљивама не сме игнорисати и то нарочито у болничким установама.

Table 1 .
The Proceedings of the 5 th International Conference on Indoor Air Quality Micromycetes isolated from analyzed filter dust of airconditioning systems.
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