The Fungal to Allergen Index is a new concept aiming at giving better information of the Indoor Air Quality. Parallel air samples are analysed for total fungi and total allergens. This gives information of the potential exposure to fungi and allergens but also an indication of the risk of a mould source. Investigations of “non-problem buildings” having no apparent moisture or mould problems, and a few buildings with mould/moisture problems showed that the median value was 8.5 % in non-problem buildings and all values was above 56% in buildings with mould/moisture problems.

This paper reports results obtained from a surface (both visually clean and dirty/dusty surfaces) and active (aggressive or activated) air testing scheme on 140 residential rooms in England, without visible water damage or mould growth, along with a few rooms with visible mould growth/water damage tested for comparison purposes. The aim was to establish normal background levels of mould in non-water-damaged interiors to benchmark a ‘normal’ indoor environment, and in turn when there is a need for further investigation, and, possibly, remediation. Air and surface mould was quantified based on the activity of β-N-acetylhexosaminidase (EC 3.2.1.52; NAHA). The obtained readings showed a log-normal distribution. Ninety-eight percent of the samples obtained from visually clean surfaces were equal to or less than 25 relative fluorescence units (RFU), which is suggested to be the higher bound for the range which can be used as a success criterion for surface cleaning/remediation. Of samples obtained from visually dirty/dusty surfaces, around 98% were below 450 RFU, which is suggested to define the lower-bound for abnormally high levels of mould, rare even on dirty/dusty surfaces. Similarly, around 98% of the air samples were found to have 1700 RFU or below. Values above 1700 RFU are therefore deemed unlikely in a non-problem indoor environment and can be indicative of a possible problem inducing mould growth. The samples with values below 1700 were further divided into three proposed sub-categories. Finally, the obtained RFU values and the suggested benchmarks were compared to those obtained from 17 non-residential indoor environments tested previously in Copenhagen, and the benchmarks that are currently used in Danish national standards, and they were both found to be highly congruent, suggesting that local climate regimes and room functions might not be as influential on indoor mould levels as commonly thought, or that the nuances between England and Denmark in terms of these factors are not strong enough to lead to sizable changes in the typical indoor mould levels in these countries’ building stocks.

This paper reports results obtained from a surface (both visually clean and dirty/dusty surfaces) and active (aggressive) air testing scheme on 140 residential rooms in England, without visible water damage or mould growth, along with a few rooms with visible mould growth/water damage tested for comparison purposes, with the aim of providing background levels of mould in non-water-damaged interiors to benchmark a normal indoor environment, and in turn when there is a need for further investigation, and, possibly, remediation. Air and surface mould was quantified based on the activity of β-N-acetylhexosaminidase (EC 3.2.1.52; NAHA). The obtained readings showed a log-normal distribution. 98% of the samples obtained from visually clean surfaces were equal to or less than 25 relative fluorescence units (RFU), which is suggested to be the higher bound for the range which can be used as a success criterion for surface cleaning/remediation in non-problem buildings. Of samples obtained from visually dirty/dusty surfaces, around 98% were below 450 RFU, which is suggested to define the lower-bound for abnormally high levels of mould, rare even on dirty/dusty surfaces. Similarly, around 98% of the air samples were found to have 1700 RFU or below. Values above 1700 RFU are therefore unlikely in a non-problem indoor environment and can be indicative of a possible problem inducing mould growth. The samples with values below 1700 were further divided into three proposed sub-categories. Finally, these values were compared to those obtained in Denmark in a similar study and are currently used in national standards, and they were found highly congruent, suggesting that local climate regimes and room functions might not be as influential on indoor mould levels, or that the nuances between UK and Denmark in terms of these factors are not strong enough to lead to sizable changes in the typical indoor mould levels in these countries.

Despite indoor mould being one of the most common problems in residential properties in the UK, there are not any widely accepted methodologies for its measurement. This paper focusses on this problem of measurement and reports on the findings from a rigorous testing scheme carried out to quantify air and surface mould concentrations and particle counts within 71 rooms from 64 properties in North London, some with and some without visible mould. The aim was to investigate the potential of passive and active air sampling strategies (sampling from still and actively mixed air, respectively) to explain visible mould, and understand how home/room characteristics correlate with the obtained readings. Airborne mould levels were quantified using an Andersen sampler (passively and actively), as well as by a chemical method based on the quantification of the N-acetylhexosaminidase (NAHA) activity (actively), which was also used to quantify surface mould. The mould levels were then correlated against physical characteristics of the tested homes/rooms, collected by means of survey sheets developed as part of this study. The findings did not reveal any independent variable governing all or most of the response variables, but a complex analysis suggested that whether it is a house or a flat could depict mould levels in the air and on the surfaces. It was also shown that a robust testing protocol should combine air and surface based methods, and an active air sampling strategy leads to a more accurate appraisal of airborne mould levels. Finally, the results showed that while there is some correlation between visible mould (and other moisture induced problems such as condensation) and measured air mould concentrations, lack of visible mould within a room does not necessarily mean low air mould concentrations, and thus one should not rely solely on visual inspection.

Mould in buildings constitutes a threat to health. Present methods to determine the moulds comprise counting of spores or determination of viable moulds which give imprecise measures of total mould cell biomass. Analysis of ergosterol and β-glucan as markers of mould cell biomass is expensive and cumbersome. To evaluate if airborne enzyme activity was related to mould in buildings air samples were taken using an impinger technique or cellulose filters in 386 rooms in 141 buildings. The samples were analysed for the activity of N-acetylhexosaminidase (NAHA) and expressed as enzyme units per m(3) (EU per m(3)). The highest value found in a building was used for the classification of the building and was related to the results from the subsequent technical inspection. In buildings without mould damage, the NAHA activity was generally below 20 EU per m(3). In buildings with mould damage, almost all the buildings had activities above 20 EU per m(3) (specificity 85%). At 30 EU per m(3) the specificity was 100%. Measurements of airborne enzyme activity have a high sensitivity and specificity to identify buildings with mould problems. The method can be used in the investigations of building related symptoms or for home exposure characteristics when investigating diseases such as asthma that can be related to mould exposure.

The purpose of the study was to measure the extent of fungal contamination in laboratory animal cage beddings over time. The material was analysed for the content of fungal enzyme N-acetylhexosaminidase and the fungal cell wall agent 1,3-beta-glucan at 0-7 days after use. In some cages the values were increased above baseline already at 3 days and at 7 days practically all beddings showed a fungal contamination. It is suggested that the fungal enzyme test can be used for bedding quality control purposes and to monitor fungal contamination in animal cages to prevent pulmonary and other pathologies.

OBJECTIVES:The aim of this study was to investigate how the microbial conditions of kitchen facilities differ from those in other school facilities. The health status of the personnel was also studied.MATERIALS AND METHODS:The microbial investigations were conducted in six moisture-damaged schools and two reference schools. The symptoms of the kitchen personnel were surveyed with questionnaires and inflammatory responses in nasal lavage (NAL) fluid were measured.RESULTS:The total concentrations of airborne microbes were lower in kitchens than in other facilities of the schools. However, the occurrence of moisture damage increased the airborne microbial concentrations both in kitchens, and in other facilities. Bacterial concentrations were high on surfaces in the damaged kitchens. Gram-negative bacteria predominated, but also thermophilic bacteria and mycobacteria were detected. Respiratory and general symptoms were prevalent both among kitchen workers and clerical personnel in the moisture-damaged environments. Reported allergies and repeated respiratory infections were connected with high IL-4 concentrations in NAL fluid. Median concentrations of studied inflammatory mediators (NO, IL-4, IL-6 and TNF-alpha) were slightly higher in NAL samples of kitchen workers than among the clerical personnel.CONCLUSIONS:Kitchen facilites differ from other facilities of the school building for their moisture conditions and microbial contamination. Thus, they represent a specific type of environment that may affect the health status of the personnel.

ABSTRACT Two mold species, Stachybotrys chartarum and Aspergillus versicolor , were inoculated onto agar overlaid with cellophane, allowing determination of a direct measurement of biomass density by weighing. Biomass density, ergosterol content, and beta- N -acetylhexosaminidase (3.2.1.52) activity were monitored from inoculation to stationary phase. Regression analysis showed a good linear correlation to biomass density for both ergosterol content and beta- N -acetylhexosaminidase activity. The same two mold species were inoculated onto wallpapered gypsum board, from which a direct biomass measurement was not possible. Growth was measured as an increase in ergosterol content and beta- N -acetylhexosaminidase activity. A good linear correlation was seen between ergosterol content and beta- N -acetylhexosaminidase activity. From the experiments performed on agar medium, conversion factors (CFs) for estimating biomass density from ergosterol content and beta- N -acetylhexosaminidase activity were determined. The CFs were used to estimate the biomass density of the molds grown on gypsum board. The biomass densities estimated from ergosterol content and beta- N -acetylhexosaminidase activity data gave similar results, showing significantly slower growth and lower stationary-phase biomass density on gypsum board than on agar.

The production of extracellular enzymes by the thermophilic fungus Thermomyces lanuginosus was studied in chemostat cultures at a dilution rate of 0.08 h−1 in relation to variation in the ammonium concentration in the feed medium. Under steady state conditions, three growth regimes were recognised and the production of several extracellular enzymes from T. lanuginosus was recorded under different nutrient limitations ranging from nitrogen limitation to carbon/energy limitation. The range and the production of carbohydrate hydrolysing enzymes and lipase increased from Regime I (NH4Cl ≤ 600 mg l−1) to Regime III (NH4CI ≥ 1200 mg l−1), whereas production of protease was highest in Regime II (600 mg l−1 < NH4Cl <1200 mg l−1).

In order to identify factors responsible for production of multiple forms of glucoamylase (GA) by Aspergillus niger Bo-1, the fungus was cultured in both complex and defined media in pH-controlled batch fermenters and chemostats. At all culture conditions three forms of GA were produced with molecular weights of approx. 91 (GAI), 73 (GAII), and 59 kDa (GAIII). Data from batch fermentations with constant pH 3.0 and 5.0 showed a uniform distribution of extracellular GA forms throughout the fermentations and independent of culture growth phases. Furthermore, steady-state data from chemostat cultivations at constant pH 3.0 and 5.0 showed a similar distribution of extracellular GA forms and established that the nitrogen concentration of the medium (C/N ratio) did not affect the distribution of multiple forms of GA. The extracellular acid protease activity was only moderate when the fungus was cultivated in batch and continuous fermentations with a constant pH of 3.0 or 5.0, whereas acidification of both complex and defined batch culture media after 20 h of growth induced a significant secretion of acid protease(s). The protease(s) present in the complex medium catalysed modification of the extracellular profile of the multiple forms of GA by degradation of GAI to GAII, whereas no proteolytic processing of the GA profile was observed in the defined medium. In vitro experiments confirmed that the pH-induced modifications of the GA multiple-form profile were caused by proteolytic and not spontaneous degradation of the GA forms at low pH. It was concluded that the observed modifications of the extracellular profile of GA isoforms in A. niger Bo-1 are due to changes in pH and medium composition.

ABSTRACT Our objective was to determine if 4-methylumbelliferyl-labelled enzyme substrates could be used to detect and quantify specific components of chitinase and cellulase activities as specific indicators of the presence and activity of fungal biomass. The fluorogenic substrates 4-methylumbelliferyl (MUF)N-acetyl-β-d-glucosaminide and MUF β-d-lactoside were used for the detection and quantification of β-N-acetylglucosaminidase (EC 3.2.1.30 ) (NAGase) and endo 1,4-β-glucanase (EC 3.2.1.4 )/cellobiohydrolase (EC3.2.1.91 ) (CELase), respectively. Culture screenings on solid media showed a widespread ability to produce NAGase among a taxonomically diverse selection of fungi on media with and without added chitin. NAGase activity was expressed only in a limited number of bacteria and on media supplemented with chitin. The CELase activity was observed only in a limited number of fungi and bacteria. Bacterial CELase activity was expressed on agar media containing a cellulose-derived substrate. In soil samples, NAGase activity was significantly correlated with estimates of fungal biomass, based on the content of two fungus-specific indicator molecules, 18:2ω6 phospholipid fatty acid (PLFA) and ergosterol. CELase activity was significantly correlated with the PLFA-based estimate of fungal biomass in the soil, but no correlation was found with ergosterol-based estimates of fungal biomass.

The yeast-mycelial dimorphic fungus Aureobasidium pullulans was grown in chemostat culture in mineral medium under zinc-limiting and zinc-sufficient conditions. The Zn2+ uptake of both the yeast and mycelial growth form conformed to saturation kinetics and the Zn2+ uptake appeared to be energy dependent and no efflux was demonstrated. No significant difference between the K-m and V-max values of the yeast and mycelial growth forms were found when grown under glucose-limiting (adequate zinc) conditions, with 7.6 mu M Zn2+ in the feed medium. Growth conditions (zinc limitation or adequate zinc) were shown to influence the Zn2+ uptake capabilities as yeast cells harvested from zinc-limited chemostat culture had a higher affinity (K-m = 0.13 mu M) and a higher V-max (1.67 nmol Zn2+ min(-1) (mg dry wt.)(-1)) than yeast cells harvested from zinc adequate chemostat culture (K-m = 0.36 mu M; V-max = 0.64 nmol Zn2+ min(-1) (mg dry wt.)(-1)). (C) 1998 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.

The yeast-mycelial dimorphic fungus Aureobasidium pullulans was grown in chemostat culture in mineral medium under zinc-limiting and zinc-sufficient conditions. The Zn2+ uptake of both the yeast and mycelial growth form conformed to saturation kinetics and the Zn2+ uptake appeared to be energy dependent and no efflux was demonstrated. No significant difference between the Km and Vmax values of the yeast and mycelial growth forms were found when grown under glucose-limiting (adequate zinc) conditions, with 7.6 μM Zn2+ in the feed medium. Growth conditions (zinc limitation or adequate zinc) were shown to influence the Zn2+ uptake capabilities as yeast cells harvested from zinc-limited chemostat culture had a higher affinity (Km= 0.13 μM) and a higher Vmax (1.67 nmol Zn2+ min−1 (mg dry wt.)−1) than yeast cells harvested from zinc adequate chemostat culture (Km= 0.36 μM; Vmax= 0.64 nmol Zn2+ min−1 (mg dry wt.)−1).

The yeast-mycelial dimorphic Aureobasidium pullulans var. pullulans was grown in chemostat culture under zinc-limitation at pH values in the range 2-7. Steady state was obtained and the growth yield with respect to zinc was independent of pH. The culture grew entirely as yeast, independently of the pH in the pH interval 3-7. The exopolysaccharide (EPS) production from the yeast cells was highest in the pH range 3-6 with a maximum at pH 4.0. Only little EPS was produced a pH 7.0 and no EPS was produced at pH 2.1. At pH 2.1 about 20% of the total biomass was in the filamentous growth form and some chlamydospores were present. A change in the carbon source from glucose to sucrose resulted in a more than doubling of the steady-state EPS concentration from 3.1 to 6.9 g l(-1). A simple mass balance on carbon was constructed which could account fur more than 95% of the carbon.

The influence of the ammonium-glucose ratio on exopolysaccharide (EPS) production and morphology of Aureobasidium pullulans was studied in continuous cultivations. A simple salt medium was used and the feed concentration of ammonium sulphate (0.3–4.2 g l−1), the only nitrogen source, was varied. Four distinct growth regimes were recognized based on steady-state concentrations of biomass, EPS, glucose, ammonium, nitrogen in the cells (% of dry wt.), and the morphology. At low feed concentrations of ammonium sulphate (0.3 to around 1.1 g 1−1) the cultures were N-limited while at high concentrations (> 2.8 g l−1) the cultures were glucose-limited. At intermediate concentrations two transition regimes were recognized in which both ammonium and glucose were exhausted and physiological responses to insufficiency in both N and C were recognized. Under N limitation the culture consisted of almost equal amounts of yeast (40%) and mycelium (60%) and the specific EPS production was constant at about 36 mg g −1 h −1. When cultures were glucose-limited the mycelial fraction of the biomass was 85–95% and no EPS was produced.

Cultivation of Aureobasidium pullulans in medium with a low concentration of yeast extract (0.4 g/l) led to a decrease in the growth rate early in the fermentation as compared to cultivations in medium with high concentration of yeast extract. When this medium was supplemented with zinc and iron the cultivation closely resembled that obtained in medium with high concentration of yeast extract (4.0 g/l). The culture retained a high growth rate throughout the fermentation and the initiation of the mycelial to yeast (M-Y) transition and the exopolysaccharide production was delayed. In a defined medium or in defined medium without iron only a little exopolysaccharide was produced and the yeast fraction of the total biomass at the onset of the stationary phase was 22%–25%. However, cultivation in the defined medium without zinc resulted in a high production of exopolysaccharide and an increased intensity of the M-Y transition, which led to a yeast fraction of 41%.

Broth from batch fermentations with Aureobasidium pullulans was investigated with a vibrating reed viscometer which showed to be sensitive to changes in the concentration of exopolysaccharide as well as in the concentration of mycelial biomass. Samples were withdrawn at 11.5, 13.5, and 20 hours after inoculation and measurements on dilution series of mycelium showed that the change in the age and therefore in the morphology of the mycelium markedly affected the sensor signal.

Penicillium commune, Aureobasidium pullulans, and Paecilomyces farinosus were grown on two different media solidified with agar, Pluronic F-127, Carrageenan X-4910, or Carrageenan X-4910 overlaid with cellophane. Growth on Carrageenan X-4910 was generally the same as that on agar, as was the visual appearance of the colonies, e.g., the pigmentation. The Carrageenan X-4910 gels had a melting point, depending on the medium, of 41 to 46(deg)C, and the dry weights of the colonies were readily determined at 60(deg)C. To determine the dry weights of the colonies grown on agar plates, the gels were boiled for 10 min to melt the agar. Comparison of these two procedures showed that the boiling procedure resulted in a 22% reduction of the biomass dry weight. Cellophane membranes did not affect the radial growth rate profoundly. The biomass density was almost halved for P. commune and P. farinosus grown with membranes, whereas the presence of the membrane did not affect the biomass density of A. pullulans. The biomass densities of the colonies grown on Pluronic F-127 were significantly reduced, while in most cases, the radial growth rates of colonies grown on Pluronic F-127 were significantly higher than those obtained on agar or Carrageenan X-4910. Furthermore, the morphology of the leading hyphae was altered, and the hyphal growth unit length was more than twice that obtained on agar and Carrageenan X-4910. Carrageenan X-4910 is a valuable gelling compound for the study of the growth of fungi, as the biomass dry weight is readily determined and growth is similar to that obtained on agar gels.

The yeast-mycelium dimorphism of Aureobasidium pullulans was studied in continuous culture in a defined medium. At a constant dilution rate (0.08 h-1) the morphological status of the culture could be controlled by the input concentration of Zn2+. As the input concentration of Zn2+ was increased (in intervals from 0 to 7.6 microM) the culture shifted from a zinc-limited to a carbon-limited state. In this interval the culture gradually passed through three growth regimes based on morphology and concentration of exopolysaccharide and biomass. The first growth regime was found when the input concentration of Zn2+ was kept below 0.45 microM. Growth in this regime was zinc-limited and more than 90% of the biomass was in the yeast growth form. An increase in the input concentration of Zn2+ in this growth regime led to a proportional increase in both the biomass and the concentration of exopolysaccharide. When the input concentration of Zn2+ was varied between 0.45 microM and 0.80 microM a second growth regime could be detected where simultaneous limitations in two nutrients were recognized. Although the carbon source (glucose) was exhausted an increase in the input concentration of Zn2+ led to a proportional increase in the steady-state biomass concentration. The increase in biomass concentration was at the expense of exopolysaccharide production, which gradually decreased. The culture, still being primarily limited by Zn2+, remained in the yeast growth form. In a third growth regime (input concentration of Zn2+ above 0.80 microM) no increase in the steady-state biomass was seen when the input concentration of Zn2+ was increased.(ABSTRACT TRUNCATED AT 250 WORDS)