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Monuments and artistic stone surfaces are often consolidated and protected with synthetic polymers, in particular, acrylics. Although it is generally thought that acrylic polymers are resistant to biodeterioration, we report for the first time the systematic occurrence of dematiaceous meristematic fungi on many marble samples of the cathedral in Milan (Italy) previously treated with this material. Fourier transform infrared spectroscopy applied to the Milan cathedral stone samples revealed characteristic features of biodeteriorated synthetic resins that differentiated them from the aged but nonbiodeteriorated samples. Samples showing biological colonization were analyzed for the presence of fungi. Cultivation and morphological characterization and methods independent from cultivation, such as denaturing gradient gel electrophoresis coupled with partial 18S rRNA gene sequencing and immunofluorescence staining with melanin-binding antibodies, showed that melanin-producing species are heavily present on stone surfaces protected with acrylic resins. This observation raises the question of the effectiveness of acrylics in protecting stone artworks.

Three expedition huts in the Ross Sea region of Antarctica, built between 1901 and 1911 by Robert F. Scott and Ernest Shackleton, sheltered and stored the supplies for up to 48 men for 3 years during their explorations and scientific investigation in the South Pole region. The huts, built with wood taken to Antarctica by the early explorers, have deteriorated over the past decades. Although Antarctica has one of the coldest and driest environments on earth, microbes have colonized the wood and limited decay has occurred. Some wood in contact with the ground contained distinct microscopic cavities within secondary cell walls caused by soft rot fungi. Cadophora spp. could be cultured from decayed wood and other woods sampled from the huts and artifacts and were commonly associated with the soft rot attack. By using internal transcribed spacer sequences of ribosomal DNA and morphological characteristics, several species of Cadophora were identified, including C. malorum, C. luteo-olivacea, and C. fastigiata. Several previously undescribed Cadophora spp. also were found. At the Cape Evans and Cape Royds huts, Cadophora spp. commonly were isolated from wood in contact with the ground but were not always associated with soft rot decay. Pure cultures of Cadophora used in laboratory decay studies caused dark staining of all woods tested and extensive soft rot in Betula and Populus wood. The presence of Cadophora species, but only limited decay, suggests there is no immediate threat to the structural integrity of the huts. These fungi, however, are widely found in wood from the historic huts and have the capacity to cause extensive soft rot if conditions that are more conducive to decay become common.

In this study, we sequenced 18S rRNA genes (rDNA) from 49 fungal strains representing 31 species from 15 genera. Most of these species are common airborne fungi and pathogens that may cause various public health concerns. Sequence analysis revealed distinct divergence between Zygomycota and Ascomycota. Within Ascomycota, several strongly supported clades were identified that facilitate the taxonomic placement of several little-studied fungi. Wallemia appeared as the group most diverged from all the other Ascomycota species. Based on the 18S rDNA sequence variation, 108 oligonucleotide probes were designed for each genus and species included in this study. After homology searches and DNA hybridization evaluations, 33 probes were verified as genus or species specific. The optimal hybridization temperatures to achieve the best specificity for these 33 probes were determined. These new probes can contribute to the molecular diagnostic research for environmental monitoring.

Two PCR primer pairs were designed to amplify rRNA genes (rDNA) from all four major phyla of fungi: Ascomycota, Basidiomycota, Chytridomycota, and Zygomycota. PCRs performed with these primers showed that both pairs amplify DNA from organisms representing the major taxonomic groups of fungi but not from nonfungal sources. To test the ability of the primers to amplify fungal rDNA from environment samples, clone libraries from two avocado grove soils were constructed and analyzed. These soils possess different abilities to inhibit avocado root rot caused by Phythophthora cinnamomi. Analysis of the two rDNA clone libraries revealed differences in the two fungal communities. It also revealed a markedly different depiction of the soil fungal community than that generated by a culture-based analysis, confirming the value of rDNA-based approaches for identifying organisms that may not readily grow on agar media. Additional evidence of the usefulness of the primers was obtained by identifying fungi associated with avocado leaves. In both the soil and leaf analyses, no nonfungal rDNA sequences were identified, illustrating the selectivity of these PCR primers. This work demonstrates the ability of two newly developed PCR primer sets to amplify fungal rDNA from soil and plant tissue, thereby providing unique tools to examine this vast and mostly undescribed community of organisms.

Significant substratum damage can occur when plasticized PVC (pPVC) is colonized by microorganisms. We investigated microbial colonization of pPVC in an in situ, longitudinal study. Pieces of pPVC containing the plasticizers dioctyl phthalate and dioctyl adipate (DOA) were exposed to the atmosphere for up to 2 years. Fungal and bacterial populations were quantified, and colonizing fungi were identified by rRNA gene sequencing and morphological characteristics. Aureobasidium pullulans was the principal colonizing fungus, establishing itself on the pPVC between 25 and 40 weeks of exposure. A group of yeasts and yeast-like fungi, including Rhodotorula aurantiaca and Kluyveromyces spp., established themselves on the pPVC much later (after 80 weeks of exposure). Numerically, these organisms dominated A. pullulans after 95 weeks, with a mean viable count +/- standard error of 1,000 +/- 200 yeast CFU cm(-2), compared to 390 +/- 50 A. pullulans CFU cm(-2). No bacterial colonization was observed. We also used in vitro tests to characterize the deteriogenic properties of fungi isolated from the pPVC. All strains of A. pullulans tested could grow with the intact pPVC formulation as the sole source of carbon, degrade the plasticizer DOA, produce extracellular esterase, and cause weight loss of the substratum during growth in vitro. In contrast, several yeast isolates could not grow on pPVC or degrade DOA. These results suggest that microbial succession may occur during the colonization of pPVC and that A. pullulans is critical to the establishment of a microbial community on pPVC.

Fungal infections, especially those caused by opportunistic species, have become substantially more common in recent decades. Numerous species cause human infections, and several new human pathogens are discovered yearly. This situation has created an increasing interest in fungal taxonomy and has led to the development of new methods and approaches to fungal biosystematics which have promoted important practical advances in identification procedures. However, the significance of some data provided by the new approaches is still unclear, and results drawn from such studies may even increase nomenclatural confusion. Analyses of rRNA and rDNA sequences constitute an important complement of the morphological criteria needed to allow clinical fungi to be more easily identified and placed on a single phylogenetic tree. Most of the pathogenic fungi so far described belong to the kingdom Fungi; two belong to the kingdom Chromista. Within the Fungi, they are distributed in three phyla and in 15 orders (Pneumocystidales, Saccharomycetales, Dothideales, Sordariales, Onygenales, Eurotiales, Hypocreales, Ophiostomatales, Microascales, Tremellales, Poriales, Stereales, Agaricales, Schizophyllales, and Ustilaginales).

DNA sequences of single-copy genes coding for chitin synthases (UDP-N-acetyl-D-glucosamine:chitin 4-beta-N-acetylglucosaminyltransferase; EC 2.4.1.16) were used to characterize ectomycorrhizal fungi. Degenerate primers deduced from short, completely conserved amino acid stretches flanking a region of about 200 amino acids of zymogenic chitin synthases allowed the amplification of DNA fragments of several members of this gene family. Different DNA band patterns were obtained from basidiomycetes because of variation in the number and length of amplified fragments. Cloning and sequencing of the most prominent DNA fragments revealed that these differences were due to various introns at conserved positions. The presence of introns in basidiomycetous fungi therefore has a potential use in identification of genera by analyzing PCR-generated DNA fragment patterns. Analyses of the nucleotide sequences of cloned fragments revealed variations in nucleotide sequences from 4 to 45%. By comparison of the deduced amino acid sequences, the majority of the DNA fragments were identified as members of genes for chitin synthase class II. The deduced amino acid sequences from species of the same genus differed only in one amino acid residue, whereas identity between the amino acid sequences of ascomycetous and basidiomycetous fungi within the same taxonomic class was found to be approximately 43 to 66%. Phylogenetic analysis of the amino acid sequence of class II chitin synthase-encoding gene fragments by using parsimony confirmed the current taxonomic groupings. In addition, our data revealed a fourth class of putative zymogenic chitin synthesis.

The fungicide benomyl has long been known to differentially affect major taxonomic groups of fungi. In the present study 163 species or aggregates of closely similar species of medically important fungi and actinomycetes, as well as species commonly isolated as clinical contaminants, were tested to determine their reactions to three concentrations of benomyl. Fungi of basidiomycetous, endomycetous, and microascaceous affinities were highly resistant, including all common yeasts and Geotrichum, Pseudallescheria, Scedosporium, and Scopulariopsis species. Also resistant were fungi of pleosporalean affinities with poroconidial anamorphs, such as Alternaria, Bipolaris, Curvularia, and Exserohilum species. Most other fungi of ascomycetous affinity were moderately to strongly susceptible. Such fungi included dermatophytes; Coccidioides, Blastomyces, and Histoplasma species; Sporothrix schenckii; medically important aspergilli; and "black yeasts." Benomyl testing aided in the provisional identification of nonsporulating mycelia, including common basidiomycetous isolates obtained as contaminants as well as nonsporulating Aspergillus fumigatus from pulmonary sources.