Detection of Airborne Stachybotrys chartarum Macrocyclic Trichothecene Mycotoxins in the Indoor Environment

The existence of airborne mycotoxins in mold-contaminated buildings has long been hypothesized to be a potential occupant health risk. However, little work has been done to demonstrate the presence of these compounds in such environments. The presence of airborne macrocyclic trichothecene mycotoxins in indoor environments with known Stachybotrys chartarum contamination was therefore investigated. In seven buildings, air was collected using a high-volume liquid impaction bioaerosol sampler (SpinCon PAS 450-10) under static or disturbed conditions. An additional building was sampled using an Andersen GPS-1 PUF sampler modified to separate and collect particulates smaller than conidia. Four control buildings (i.e., no detectable S. chartarum growth or history of water damage) and outdoor air were also tested. Samples were analyzed using a macrocyclic trichothecene-specific enzyme-linked immunosorbent assay (ELISA). ELISA specificity was tested using phosphate-buffered saline extracts of the fungal genera Aspergillus, Chaetomium, Cladosporium, Fusarium, Memnoniella, Penicillium, Rhizopus, and Trichoderma, five Stachybotrys strains, and the indoor air allergens Can f 1, Der p 1, and Fel d 1. For test buildings, the results showed that detectable toxin concentrations increased with the sampling time and short periods of air disturbance. Trichothecene values ranged from <10 to >1,300 pg/m³ of sampled air. The control environments demonstrated statistically significantly (P < 0.001) lower levels of airborne trichothecenes. ELISA specificity experiments demonstrated a high specificity for the trichothecene-producing strain of S. chartarum. Our data indicate that airborne macrocyclic trichothecenes can exist in Stachybotrys-contaminated buildings, and this should be taken into consideration in future indoor air quality investigations.     

Rapid method for the determination of ochratoxin A in urine by immunoaffinity column clean-up and high-performance liquid chromatography

Buildings with poor indoor air quality (IAQ) frequently have many areas with surface fungal contamination. Studies have demonstrated that certain fungal genera (e.g., Cladosporium, Penicillium, and Stachybotrys) are able to grow on building materials such as wallpaper, drywall, and ceiling tiles, particularly after water damage has occurred. Due to the increasing awareness of sick building syndrome (SBS), it has become essential to identify building materials that prevent the interior growth of fungi. The objective of this study was to identify building materials that would not support the growth of certain fungal genera, regardless of whether an external food source was made available. The growth of three fungal genera (Cladosporium, Penicillium, and Stachybotrys) was evaluated on cellulose-containing ceiling tile (CCT) and inorganic ceiling tile (ICT). Both types of ceiling tile were exposed to environmental conditions which can occur inside a building. Our results show that ICT did not support the growth of these three fungal genera while CCT did. Our data demonstrate that ICT could serve as an ideal replacement for CCT. This revised version was published online in June 2006 with corrections to the Cover Date.     

Mycotoxins as harmful indoor air contaminants

Fungal metabolites (mycotoxins) that pose a health hazard to humans and animals have long been known to be associated with mold-contaminated food and feed. In recent times, concerns have been raised about exposures to mycotoxin-producing fungi in indoor environments, e.g., damp homes and buildings. The principal mycotoxins that contaminate food and feed (alfatoxins, fumonisins, ochratoxin A, deoxynivalenol, zearalenone) are rarely if ever found in indoor environments, but their toxicological properties provide an insight into the difficulties of assessing the health effects of related mycotoxins produced by indoor molds. Although the Penicillium and Aspergillus genera of fungi are major contaminants of both food and feed products and damp buildings, the particular species and hence the array of mycotoxins are quite different in these environments. The mycotoxins of these indoor species and less common mycotoxins from Stachybotrys and Chaetomium fungi are discussed in terms of their health effects and the need for relevant biomarkers and long-term chronic exposure studies.     

Air- and Dustborne Mycoflora in Houses Free of Water Damage and Fungal Growth

Typically, studies on indoor fungal growth in buildings focus on structures with known or suspected water damage, moisture, and/or indoor fungal growth problems. Reference information on types of culturable fungi and total fungal levels are generally not available for buildings without these problems. This study assessed 50 detached single-family homes in metropolitan Atlanta, Ga., to establish a baseline of “normal and typical” types and concentrations of airborne and dustborne fungi in urban homes which were predetermined not to have noteworthy moisture problems or indoor fungal growth. Each home was visually examined, and samples of indoor and outdoor air and of indoor settled dust were taken in winter and summer. The results showed that rankings by prevalence and abundance of the types of airborne and dustborne fungi did not differ from winter to summer, nor did these rankings differ when air samples taken indoors were compared with those taken outdoors. Water indicator fungi were essentially absent from both air and dust samples. The air and dust data sets were also examined specifically for the proportions of colonies from ecological groupings such as leaf surface fungi and soil fungi. In the analysis of dust for culturable fungal colonies, leaf surface fungi constituted a considerable portion (>20%) of the total colonies in at least 85% of the samples. Thus, replicate dust samples with less than 20% of colonies from leaf surface fungi are unlikely to be from buildings free of moisture or mold growth problems.     

Air- and dustborne mycoflora in houses free of water damage and fungal growth

Typically, studies on indoor fungal growth in buildings focus on structures with known or suspected water damage, moisture, and/or indoor fungal growth problems. Reference information on types of culturable fungi and total fungal levels are generally not available for buildings without these problems. This study assessed 50 detached single-family homes in metropolitan Atlanta, Ga., to establish a baseline of "normal and typical" types and concentrations of airborne and dustborne fungi in urban homes which were predetermined not to have noteworthy moisture problems or indoor fungal growth. Each home was visually examined, and samples of indoor and outdoor air and of indoor settled dust were taken in winter and summer. The results showed that rankings by prevalence and abundance of the types of airborne and dustborne fungi did not differ from winter to summer, nor did these rankings differ when air samples taken indoors were compared with those taken outdoors. Water indicator fungi were essentially absent from both air and dust samples. The air and dust data sets were also examined specifically for the proportions of colonies from ecological groupings such as leaf surface fungi and soil fungi. In the analysis of dust for culturable fungal colonies, leaf surface fungi constituted a considerable portion (>20%) of the total colonies in at least 85% of the samples. Thus, replicate dust samples with less than 20% of colonies from leaf surface fungi are unlikely to be from buildings free of moisture or mold growth problems.     

An aerobiological study in the rural areas of Aragon (Spain) with a high population of pigs

The development of intensive livestock farming in the swine industry has created problems associated not only with disease control but also with the disposal of excreta and other waste products. Both waste products and animals can contaminate the air of confinement swine buildings and their surroundings. The objective of this work was to determine the concentration and species composition of the bacterial and fungal microflora in the air of different villages with a high population of pigs in their surroundings. Outdoor air sampling was conducted in 24 rural urban nucleus of Aragon (Spain). In each site, three sampling points were done: one in the center of the town, another in the outskirts of the same town, and the last one in the surrounding farms. Microorganisms were isolated with a sampler Surface Air System (S.A.S.). In the quantitative analysis, the potential risk of exposure to airborne microorganisms decreased outside and near of the farms and, of course, in other sampling points, because the airborne levels of total bacteria and fungi found in the three points are similar, slightly higher in the surrounding farms and lower inside of the buildings. The genera Staphylococcus and Bacillus among isolated Grampositive bacteria were the most common, while among the Gram-negative genera often were Alcaligenes and Acinetobacter in most cases observed. In fungi, the most isolated genera were Cladosporium, Aspergillus, Penicillium and Alternaria, which are very important aeroallergens. In this work, were found no differences between the three sampling, which are indicative of few microbial contamination of the air from swine confinement buildings at short distances.     

Preliminary survey of indoor and outdoor airborne microfungi at coastal buildings in Egypt

Forty six species and two sterile fungi and yeast species were isolated from samples collected both indoors and outdoors of coastal buildings located in an Egyptian coastal city. Twenty flats from ten buildings were investigated; children living in these buildings have been reported to suffer from respiratory illnesses. Samples were taken using a New Brunswick sampler (model STA-101) operating for 3.0 min at a flow rate of 6.0 l/min. Most of the species isolated have been associated with symptoms of respiratory allergies. Indoors the total culturable fungal count was 1548 CFU/m³; outdoors, it was 1452 CFU/m³. Indoor values of culturable fungal count, total spores count and ergosterol content ranged from 52 to 124 CFU/m³, 100 to 400 spore/m³ and 5 to 27.7 mg/m³, respectively, whereas outdoor levels typically varied between 25 and 222 CFU/m³, 110 and 900 spore/m³ and 3.3 and 67.2 mg/m³, respectively. The maxima for these parameters were detected indoors in house no. 6 and outdoors, outside of house no. 7. The most abundant species were primarily mitosporic (2832 CFU/m³). The most frequent species in both the indoor and outdoor samples were Cladosporium cladosporioides followed by Alternaria alternata and Penicillium chrysogenum,with inside:outside ratios of 1.4, 1.8 and 1.9, respectively. The patterns of fungal abundance were influenced to some extent by changes in the relative humidity and temperature. Other factors, such as type of culture media, rate of sedimentation, size, survival rates of spore and species competition,also affected fungal counts and should be taken into consideration during any analysis of bioaerosol data.     

Contamination of air systems: 1987–88 assessment and prospects of the Grenoble intervention group

Summary In order to evaluate bacterial and fungal contamination, the authors used a qualitative and quantitative procedure and investigated 47 air conditioning and humidifying units. Air samples were studied using Biotest RCS biocollector (160×6 liters on 6 different selective media), whilst dust samples were analysed after spreading 10 mg onto fungal media. Selective research ofLegionella and fungi were performed using water filtrates and specific media. Even though, the most frequently identified species were quite common environmental fungi, mainlyPenicillium, Alternaria andAspergillus fumigatus (36 out of 47 collections), the health effects of intense exposure to these common moulds are not really known.Staphylococcus aureus was identified 6 times and thermophilic actinomyces species (A. candidus) once. From the quantitative analysis a significant relationship was found between some risk factors and airborne contamination. Indeed bacteria and fungi number depends on the humidifying system (steam or washer), efficiency of secondary filters, relative humidity percentage (< or >60%), installation maintenance, industrial activity and complaints of people at risk. The presence of air recycling and biocide use seems only to have an influence on fungal flora. Having determined a global risk score for each air conditioning unit, the authors underline the fact that bacterial and fungal airborne contamination increases with score. Moreover, for a global, metrological, medical and technical evaluation, a multidisciplinary approach has proved itself to be indispensable.     

Biodiversity and concentration of airborne fungi in a hospital environment

The biodiversity and concentration of airborne fungi were monitored over a period of 6 months in a special-care unit of a hospital. Air sampling was performed in a corridor that was also accessible to visitors and in an adjacent bone-marrow transplantation (BMT) unit using an air sampler and two isolation media. Altogether, 98 fungal species could be identified, among them Aspergillus fumigatus and A. terreus as well as 48 other species reported as potential pathogens. The average contamination values of the corridor air ranged from 124 to 485 cfu m⁻³. Neither the degree of fungal air contamination nor the species composition inside the special care unit differed from those found in the corridor. By means of data obtained with a light-activated sensor, a possible influence of human activities on diurnal changes of fungal propagule concentration was shown. This revised version was published online in July 2006 with corrections to the Cover Date.     

Identification and seasonal distribution of airborne fungi in three horse stables in Italy

Fungal agents are responsible for a variety of respiratory diseases both in humans and animals. The nature and seasonal variations of fungi have been investigated in many environments with wide ranging results. The aims of the present report were (i) to evaluate the quality and magnitude of exposure to airborne fungi in three differently structured equine stalls (open air, partially and completely enclosed buildings) during a one-year period, using an air sampling technique and (ii) to compare the distribution and frequency of fungal species, with regards to these different environments. Air samples were collected monthly from December 2001 to November 2002 by means of a surface air sampler (SAS) Super-90, (PBI International, Milan, Italy). Penicillium and Aspergillus spp. were cultured from all the stables in all seasons. Mucoraceae were also recovered in all seasons in stalls 1 and 2, while they were not isolated in spring and fall in stall 3. These fungi were detected in 28.4%, 72.9% and 60.5% of the total number of samples, respectively. Other fungal genera such as Alternaria, Cladosporium, Fusarium, Beauveria and Drechslera were also occasionally recovered.Viable fungal concentrations varied greatly, ranging from below the limit of detection to more than 3000 CFU/m³ for stables 1 and 2, and 1750 CFU/m³ for stable 3. The median fungal concentration was approximately 178 CFU/m³. Total fungal concentration appeared to be highest in summer, winter and spring, and lowest in the fall.     

The importance of combining air sampling and surface analysis when studying problematic houses for mold biodiversity determination

Houses that underwent water damages are often responsible for heath problems of the occupants. Since there is no universally used protocol for the analysis, we wanted to verify the usefulness of surface sampling versus air sampling for the evaluation of mold diversity in problematic houses and the value of the number of visible mold growth zones to predict air quality. Seventeen houses were sampled for culturable molds in the air and on the surfaces showing contamination. We compared the mold taxa found in the air and on the surfaces and verified the correlation between the number of moldy surfaces and airbone mold concentration. This study demonstrated that, surprisingly, some of the so called wet spore molds (e.g. Stachybotrys) were found more often from air than surface samples whereas, some dry spore molds (e.g. Asp. fumigatus) was more easily isolated from surface samples. There was a good correlation between the number of visible mold growth zones and the concentration of airborne molds. We conclude that air and surface sampling are necessary to evaluate mold diversity in problematic houses and that the number of mold growth zones is a good predictor of airborne mold concentration.     

Temporal and spatial variation of indoor and outdoor airborne fungal spores,pollen, and (1→3)-<Emphasis Type="Italic">β</Emphasis>-<Emphasis Type="SmallCaps">d</Emphasis>-glucan

Characterizing the variation in bioaerosol concentrations is important for the estimation of health effects associated with bioaerosols and planning exposure assessment strategies. This investigation was conducted in order to develop a better understanding of exposure to fungal spores, pollen, and (1→3)-β-d-glucan, by determining the variations of their concentrations between and within homes. In the study, 24-h air sampling was performed on five consecutive days in four Cincinnati area homes. The samples (a total of 160) were taken simultaneously in four different rooms inside each home and at four different outside locations near the home using Button Personal Inhalable Aerosol Samplers. The relative sizes of the between- and within-home variability to the total variability were calculated for each outcome. The relative sizes of the between- and within-home variability in indoor air ranged from 0.10 to 0.52 and 0.09 to 0.10, respectively. For outdoor air, the between- and within-home variability ranged from 0.27 to 0.50 and 0.09 to 0.10, respectively. Thus, the ranges of within-home variability, both indoors and outdoors, were much less than the variability between different homes. The results suggest that, if long-term sampling methods are employed to characterize the bioaerosol exposure for a population, the sampling should be repeated in a larger number of homes as an alternative to replicate sampling in a fewer number of homes. When characterizing exposure within one home, the sampling should be repeated in different rooms, rather than repeating it on different days.     

Prevalence and distribution of airborne and waterborne fungi and actinomycetes in the Nile river

The objective of this research was to investigate the prevalence and distribution of airborne and waterborne fungi and actinomycetes along the main stream of the Nile river during April to July, 2005. Air and water samples were collected at eight sites within a ~50 km stretch of the river. The distribution and prevalence of air and water microorganisms varied with location. The highest counts of airborne fungi (516 CFU/p/h) and actinomycetes (222 CFU/p/h) were detected at suburban sites near cultivated areas. However, the highest counts of waterborne fungi (56.4 CFU/ml) and actinomycetes (15.4 CFU/ml) were detected at Al-Galaa (city centre) and Kafr-El-elwe (south Cairo), respectively. A total of 1,816 fungal colonies (943 isolates from air and 873 from water samples) belonging to 27 genera were identified. Aspergillus, Alternaria, Cladosporium, and yeasts were the predominant fungal types in both air and water environments. Dreschlera, Emericella, Nigrospora, Spicaria, Stachybotrys, and Verticillium were only detected in the air, and Epicoccum, Philaphora, Phoma and Ulocladium were only detected in the water. Mycotoxin-producing fungi represented by Aspergillus flavus, Aspergillus parasiticus, Penicillium, Fusarium, and Trichoderma were found in the air and water environments. Significant differences (P ≤ 0.05) were found between fungal populations in air and water at different sampling sites. No significant differences (P ≥ 0.05) were found between waterborne actinomycetes. Sampling location, human activity, and pollution load are the main factors affecting the variability and biodiversity of microorganisms in different microenvironments.     

Detection of culturable and nonculturable Legionella species from hot water systems of public buildings in Japan

Aims: To investigate the prevalence of culturable and nonculturable Legionella species in hot water systems of public buildings in Japan and assess the risk factors associated with Legionella contamination in hot water systems. Methods and Results: Legionella species were detected by conventional culture and molecular methods in 130 water samples collected from 40 buildings. A total of 26 (20·0%) water samples from 17 (42·5%) buildings were positive by culture, qualitative PCR or both methods: Legionella pneumophila and Leg. anisa were detected in four samples by a culture method, whereas 23 samples were positive by qualitative PCR, with the presence of various Legionella species confirmed by sequencing. Of these 23 samples, bacterial counts were quantifiable in 21 by real‐time PCR (from 1·7 × 10⁵ to 2·6 × 10¹¹ cells per litre). Phylogenetic analysis of amplified partial 16S rRNA gene showed close relations to various species of Legionella, including Leg. anisa and Leg. micdadei, all of which have been associated with respiratory diseases or increased antibody titres in human sera. Assessment of risk factors showed that turbidity, free chlorine concentration, iron concentration and heterotrophic plate count (HPC) were significantly associated with Legionella contamination (P < 0·05). Conclusions: Contamination of hot water systems of public buildings with culturable and nonculturable Legionella species may be a potential risk factor for Legionella infection in Japan. Adequate levels of chlorine, low levels of iron and HPC are important maintenance measures in the reduction of Legionella contamination in hot water systems. Significance and Impact of the Study: More than 40% of hot water systems in the Japanese public buildings examined were contaminated by not only culturable Leg. pneumophila and Leg. anisa but also by nonculturable pathogenic species. To our knowledge, this is the first report of both culturable and nonculturable Legionella contamination in hot water systems of public buildings in Japan.     

Prevalence of fungi in carpeted floor environment: Analysis of dust samples from living-rooms,bedrooms, offices and school classrooms

The results of 100 carpet dust analyses from atopic individuals' environment were compared according to the sampling period or the location. Dust samples were collected with a standard domestic vacuum cleaner, in locations with carpeted floor: in residences (living-room and/or bedroom), in school classrooms and in offices. The quantities of fungi vary from 5000 CFU/g to 66 000 000 CFU/g of dust. More than 100 species were isolated by dilution plating. The main species found in carpet dust wereEurotium repens, Penicillium chrysogenum, Alternaria alternata, Aureobasidium pullulans andPhoma herbarum. Strict xerophilic species were rather rare and detected in small quantities. Differences in the distribution of the CFU concentrations were examined for the four different sampling locations and were statistically significant (P=0.0174). In this study, schools were open spaces, and offices, mostly with air conditioning systems, were locations in which air is not confined. This, added to frequent professional carpet cleaning, probably explains the lowest levels of fungal concentration found in these locations. The majority of the homes had the largest fungal concentration in the living-room (median: 2×10⁵ CFU/g) while some bedrooms (median: 7×10⁴ CFU/g) had the highest concentrations. It is suggested that, when fungi are suspected to be the origin of respiratory allergy or irritating symptoms, the mycoflora of the bedroom, principally, should be investigated first.     

Profiles of Airborne Fungi in Buildings and Outdoor Environments in the United States

We examined 12,026 fungal air samples (9,619 indoor samples and 2,407 outdoor samples) from 1,717 buildings located across the United States; these samples were collected during indoor air quality investigations performed from 1996 to 1998. For all buildings, both indoor and outdoor air samples were collected with an Andersen N6 sampler. The culturable airborne fungal concentrations in indoor air were lower than those in outdoor air. The fungal levels were highest in the fall and summer and lowest in the winter and spring. Geographically, the highest fungal levels were found in the Southwest, Far West, and Southeast. The most common culturable airborne fungi, both indoors and outdoors and in all seasons and regions, were Cladosporium, Penicillium, nonsporulating fungi, and Aspergillus. Stachybotrys chartarum was identified in the indoor air in 6% of the buildings studied and in the outdoor air of 1% of the buildings studied. This study provides industrial hygienists, allergists, and other public health practitioners with comparative information on common culturable airborne fungi in the United States. This is the largest study of airborne indoor and outdoor fungal species and concentrations conducted with a standardized protocol to date.     

辽宁碱蓬根际土壤真菌多样性的季节变化及其耐盐性

采用稀释平板法和形态学鉴定法进行了辽宁碱蓬(Suaeda liaotungensis)根际土壤真菌的分离鉴定及季节、盐度对真菌多样性的影响分析。结果表明:春、夏、秋、冬四季辽宁碱蓬根际土壤中的真菌菌落数分别为6410、4180、5730和3340,种类分别为6属13种、9属16种、11属31种、6属12种,共分离鉴定出13属42种真菌;其根际土壤真菌的多样性指数、丰富度指数和均匀度指数,从大到小的次序均为秋夏春冬,3种指数均在秋季达到峰值;多样性指数和丰富度指数差异较大,均匀度指数波动较小,春季和冬季的均匀度指数相近。辽宁碱蓬根际土壤真菌种类明显多于无植被土壤,是无植被土壤的7倍,多样性指数和均匀度指数分别是无植被土壤的4—8倍和1—1.2倍;其根际土壤真菌优势种群包括青霉属(Penicillium)、葡萄穗霉属(Stachybotrys)、枝孢属(Cladosporium)、木霉属(Trichoderma)、曲霉属(Aspergillus)和镰孢属(Fusarium);4个季节的优势菌不尽相同,但均有一个共同的优势菌属青霉属。以Na Cl浓度梯度法制造盐胁迫生境,检测辽宁碱蓬根际土壤真菌对盐胁迫的响应,结果表明:60%左右的菌种能耐受5%以下盐度、15%左右的菌种能耐10%—20%盐度;筛选出了6株高度耐盐菌:细交链孢霉、草酸青霉、产黄青霉、烟曲霉、细极链格孢和赭曲霉,其中最高耐盐菌种赭曲霉可耐20%盐度。创新之处在于以盐度和季节为变量检测辽宁碱蓬根际土壤真菌的种群构成,为其深入研究奠定了理论基础。     

Investigating the inoculum dynamics of Cladosporium on the surface of raspberry fruits and in the air

Raspberry production is under threat from the emerging fungal pathogenic genus Cladosporium. We used amplicon-sequencing, coupled with qPCR, to investigate how fruit age, fruit location within a polytunnel, polytunnel location and sampling date affected the fruit epiphytic microbiome. Fruit age was the most important factor impacting the fungal microbiome, followed by sampling date and polytunnel location. In contrast, polytunnel location and fruit age were important factors impacting the bacterial microbiome composition, followed by the sampling date. The within-tunnel location had a small significant effect on the fungal microbiome and no effect on the bacterial microbiome. As fruit ripened, fungal diversity increased and the bacterial diversity decreased. Cladosporium was the most abundant fungus of the fruit epiphytic microbiome, accounting for nearly 44% of all fungal sequences. Rotorod air samplers were used to study how the concentration of airborne Cladosporium inoculum (quantified by qPCR) varied between location (inside and outside the polytunnel) and time (daytime vs. nighttime). Quantified Cladosporium DNA was significantly higher during the day than the night and inside the polytunnel than the outside. This study demonstrated the dynamic nature of epiphytic raspberry fruit microbiomes and airborne Cladosporium inoculum within polytunnels, which will impact disease risks on raspberry fruit.     

Fungi and fungal products in some Canadian houses

Building related illness prompted a study in the winter of 1986 to identify and quantify and fungal products present in c. 50 Canadian homes. Of these, 70% had been reputedly associated with health problems. Building parameters, i.e. air change rate and the internal moisture levels, were measured, and the fungi present were characterized and quantified along with their metabolites. Air and dust samples were analyzed and the fungal biomass in the dust was measured by a procedure which involved determination of ergosterol by a gas chromatograph/mass spectrometer system. Some 42 fungal species were identified in air, samples of which were further analyzed for fungal volatiles. Penicillium was the most common genus in both air and dust, together with Cladosporium and Alternaria. The potentially hazardous fungus Aspergillus fumigatus was found in only two houses, and Strachybotry atra in only one. New criteria are suggested to define the acceptable standards for indoor fungal levels in air during winter.