Evaluation of quantitative PCR and culture methods for detection of house dust fungi and streptomycetes in relation to moisture damage of the house.

AIMS: Microbial concentrations in vacuumed house dust samples (n = 71) were analysed by culture and quantitative polymerase chain reaction (qPCR) methods and their association with extent of moisture damage in the house was studied. METHODS AND RESULTS: Microbial concentrations measured by qPCR correlated with concentrations obtained by culture method, but were orders of magnitude higher. qPCR also had better sensitivity. Concentrations of several microbes in house dust, determined with qPCR, were associated with the extent of moisture damage in the house. This association was strongest for Penicillium brevicompactum, one of the fungi detected in highest concentrations by qPCR. Furthermore, house dust concentrations of Wallemia sebi, Trichoderma viride, Cladosporium sphaerospermum, Eurotium amstelodami and the combined assay group for Penicillium spp., Aspergillus spp. and Paecilomyces variotii were significantly associated with the extent of the moisture damage. CONCLUSION: These species or assay groups could probably be used as indicators of moisture damage in the house. SIGNIFICANCE AND IMPACT OF THE STUDY: This finding indicates the benefits of the qPCR method, which is sensitive enough to reveal the differences in microbial concentrations of house dust between moisture-damaged and undamaged houses.

pH effects on 10 Streptomyces spp. growth and sporulation depend on nutrients.

AIMS: Streptomycetes are regarded to prefer neutral to alkaline environmental pH, although they commonly occur at remarkably variable pH and nutritional conditions. Therefore, the dependence of 10 Streptomyces spp. pH tolerance on nutrients was determined. METHODS AND RESULTS: Ten environmental Streptomyces spp. were grown and sporulated between pH 4.0 and 11.5, at the interval of 1.5, on starch-casein-KNO(3), tryptone-yeast extract-glucose, glycerol-arginine and tryptone-soy agars, and three their modifications. On media with starch and casein; glucose, tryptone and yeast extract; tryptone and soy peptone; and glycerol-arginine and yeast extract strains grew over a broad pH range between 4.0-5.5 and 10.0-11.5. On glycerol-arginine and on medium with Na-propionate, NH(4)NO(3) and yeast extract, streptomycetes grew optimally at pH 7.0 and above. The high organic load enabled the growth over a wide pH range. The sporulation pH ranges followed those for growth. CONCLUSIONS: The high organic load enabled the growth over a wide pH range. The strain-specific differences in sporulation were greater than those caused by pH. The best medium for sporulation contained glucose and tryptone with minerals of glycerol-arginine agar at pH 5.5. SIGNIFICANCE AND IMPACT OF THE STUDY: The growth pH ranges, pH ranges for the optimal growth, and sporulation were strongly dependent on nutrients.

Effect of growth medium on potential of Streptomyces Anulatus spores to induce inflammatory responses and cytotoxicity in RAW264.7 macrophages.

Epidemiological studies have shown an association between microbial growth in buildings and increased risk of respiratory symptoms and disease related to inflammatory reactions in the inhabitants96. The current study examined the affects of growth conditions of Streptomyces anulatus, isolated from indoor air of a moldy building, on the inflammatory potential of spores of this microbe. Spores were harvested from 15 growth media formulations, applied to RAW264.7 macrophages (10(5), 10(6), or 10(7) spores/million cells), and evaluated for the ability to stimulate production of inflammatory mediators and cytotoxicity in these cells 24 h after exposure. Streptomyces anulatus spores induced dose-dependent production of nitric oxide (NO) in macrophages, reaching a level from 4.2 microM to 39.2 microM depending on the composition of the growth medium of the microbe. Expression of inducible NO synthase (iNOS) was detected in macrophages after exposure to spores collected from all growth media. Production of reactive oxygen species (ROS) was significantly increased only by the highest dose of S. anulatus spores grown on glycerol-arginine agar. Furthermore production of cytokines was affected by growth medium; the highest dose-dependent levels of interleukin 6 (IL-6) ranged from 900 to 7800 pg/ml, and the levels of tumor necrosis factor alpha (TNFalpha) varied from 490 to 3200 pg/ml. The amount of dead macrophages after the exposure varied from 11% to 96%, depending also on the growth media of the microbe. Altogether, our results suggest that the growth medium of S. anulatus has a fundamental role in the ability of the spores to induce inflammatory responses and cytotoxicity in mammalian cells.