A novel PCR-based approach for the detection and classification of potential cellulolytic fungal strains isolated from museum items and surrounding indoor environment.

AIMS: To develop a novel PCR-based method able to detect potential cellulolytic filamentous fungi and to classify them exploiting the amplification of the cellobiohydrolase gene (cbh-I) and its polymorphism. METHODS AND RESULTS: A mixed approach including the combination of (i) fungal cultivation and isolation, (ii) classification of fungal isolates through the amplification of the cbh gene using a fluorescently labelled primer (f-CBH-PCR) and (iii) final fungal identification based on amplification and sequencing of the ITS1-5.8S rDNA-ITS2 region of the selected fungal strains was developed. By this approach, it was possible to screen 77 fungal strains belonging to 14 genera and 26 species. CONCLUSIONS: The f-CBH-PCR permitted the discrimination of fungal species, producing typical f-CBH profiles. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study, the cbh gene was used as a preliminary classification tool able to differentiate among themselves the fungal members isolated from indoor museum items and surrounding environment. Such mixed approach consented the fast identification of all isolated fungal strains. The f-CBH-PCR method demonstrated its discrimination power, and it can be considered as a new molecular system suitable for the classification of fungal strains isolated from different environments.

Selection and identification of autochthonous yeasts in Slovakian wine samples using a rapid and reliable three-step approach.

AIMS: The investigation of yeast microflora during the must fermentation of two wine varieties (Frankovka modra - Blaufränkisch and Veltlinske zelene - Grüner Veltliner) from two consecutive vintages was performed using a three-step approach. METHODS AND RESULTS: The investigation strategy consisted of the combination of yeast cultivation, selection of the isolated yeasts based on the amplification of internal transcribed spacer 2 using a fluorescence-labelled primer (f-ITS-PCR) and a final identification step based on amplification and sequencing of the ITS1-5.8S rDNA-ITS2 region of the selected yeasts. By this three-step approach, it was possible to screen 433 yeasts isolates that belonged to 13 different species. CONCLUSIONS: The f-ITS-PCR allowed the unambiguous differentiation of all isolated yeast species that produced their typical f-ITS-PCR profile. SIGNIFICANCE AND IMPACT OF THE STUDY: This is one of few reports that treat the yeast diversity in Slovakian wines and in two varieties largely cultivated in Central Europe. The three-step approach permitted the rapid and reliable identification of isolated yeasts. The f-ITS-PCR with its good discrimination power can represent a suitable molecular tool for the selection of yeast members recovered from food or other environments.

Wooden art objects and the museum environment: identification and biodegradative characteristics of isolated microflora.

AIMS: The identification of culturable microbial communities on wooden art objects and from indoor air, and the analysis of their biodegradative properties. METHODS AND RESULTS: Common and newly-developed agar media were used for the isolation of fungal and bacterial microflora. The identification was carried out by traditional methods and by the sequencing of 16S or 18S rDNA PCR products. Different plate assays were employed to screen the lignolytic and cellulolytic activities of the isolated microflora. Interesting bacteria were isolated from art objects even though the fungi were the principal contaminants of art works. Various fungal and bacterial species exhibited their lignolytic and cellulolytic activity by the decolorization of Remazol Brilliant Blue R, Phenol Red, Azure B and Ostazin Brilliant Red H-3B. CONCLUSIONS: The microbial communities on wooden art objects exposed in an indoor environment were identified. The study showed the biodegradative power of many microorganisms, and new data were added to this field barely investigated. SIGNIFICANCE AND IMPACT OF THE STUDY: By the development of new culture media and the evaluation of different biodegradative plate assays, a strategy for the analysis of microflora in wooden art objects was established. Several aspects of the study could be also exploited for biotechnology applications.