Infections caused by fungi have increased in recent years. Accurate and rapid identification of fungal pathogens is important for appropriate treatment with antifungal agents. On the basis of the internal transcribed spacer 1 (ITS 1) and ITS 2 sequences of the rRNA genes, an oligonucleotide array was developed to identify 64 species (32 genera) of clinically important filamentous (or dimorphic) fungi. These 64 species included fungi causing superficial, cutaneous, subcutaneous, and invasive infections. The method consisted of PCR amplification of the ITS regions using a pair of universal primers, followed by hybridization of the digoxigenin-labeled PCR products to a panel of species-or group-specific oligonucleotides immobilized on a nylon membrane. Of 397 fungal strains (290 target and 107 nontarget strains) tested, the sensitivity and specificity of the array was 98.3% (285/290) and 98.1% (105/107), respectively. Misidentified strains were usually those belonging to the same genus of the target species or having partial homology with oligonucleotide probes on the membrane. The whole procedure can be finished within 24 h starting from isolated colonies; reproductive structures, which are essential for the conventional identification methods, are not needed. In conclusion, the present array is a powerful tool for identification of clinically important filamentous fungi and may have the potential to be continually extended by adding further oligonucleotides to the array without significantly increasing the cost or complexity.

Three apiculate yeast strains, EJ7M09T , GJ5M15 and GJ15M04, isolated from mushrooms in Taiwan were found to represent a novel species of the genus Kloeckera. The phylogenetically closest relative of this novel species is Hanseniaspora occidentalis, but the type strain of H. occidentalis differed by 4.6 % divergence (25 substitutions; 5 gaps) in the sequence of the D1/D2 domain of the large subunit rRNA gene. This difference clearly suggests that the three strains represent a distinct species. As none of the strains that were examined in this study produced ascospores or exhibited conjugation on common sporulation medium either alone or in a pairwise mixture, this species could be considered as an anamorphic member of the genus Hanseniaspora, and a novel species, Kloeckera taiwanica sp. nov., is proposed, with EJ7M09 The ascosporogenous genus Hanseniaspora and its anamorphic genus Kloeckera are characterized by bipolar budding as apiculate yeasts (Smith, 1998a, b). Several novel apiculate yeasts have been described since Reess (1870) proposed a novel species Saccharomyces apiculatus (5Kloeckera apiculata, an anamorph of Hanseniaspora uvarum) to accommodate yeast strains that reproduce by bipolar budding. Meyer et al. (1978) carried out detailed taxonomic studies based on DNA-DNA reassociation and classified these apiculate yeasts into six species in the genus Hanseniaspora, proposed by Zikes (1911), and one species in the anamorphic genus Kloeckera, proposed by Janke (1923 Therefore, 13 Hanseniaspora species and two Kloeckera species are currently recognized.Members of the genera Hanseniaspora and Kloeckera have been reported in various habitats such as fruit, flowers, soil and fermenting juice, as well as insect-associated samples (Jindamorakot et al., 2009;Cadez et al., 2003Cadez et al., , 2006. During an investigation of yeast diversity in mushrooms in Taiwan, many Hanseniaspora and Kloeckera strains were also isolated from the fruiting bodies of mushrooms. Of these isolates, three strains (EJ7M09 T , GJ5M15 and GJ15M04) were distinctly different from members of currently recognized species. These strains were highly similar to each other in their morphological and molecular characteristics, indicating that they are conspecific. Strains EJ7M09 T , GJ5M15 and GJ15M04 were isolated from the fruiting bodies of the mushrooms Russula sp., Mycena sp., and Bisporella sp., respectively; EJ7M09 T was found in Hsinchu county, whereas GJ5M15 and GJ15M04 were from Pingtung county, Taiwan. Based on our data, we propose that these strains represent a novel species.The yeast strains were isolated from mushroom fruiting bodies as described by Hsieh et al. (2010). Approximately 1.0 g fruiting body from each sample was placed into a tube containing 9 ml yeast extract-malt extract (YM) broth (1 % glucose, 0.5 % peptone, 0.3 % yeast extract, 0.3 % malt extract, pH 5.4) supplemented with 50 mg chloramphenicol ml 21 and then vortex-mixed. One-tenth of a millilitre of successive decimal dilutions was spread on acidified YM agar (pH...

Two novel ascomycetous yeast species, Saturnispora serradocipensis and Saturnispora gosingensis, were isolated from leaf detritus in a tropical stream of Southeastern Brazil and a mushroom collected in Taiwan, respectively. Analysis of the D1/D2 domains of the large-subunit of the rRNA gene of these strains showed that these species are related to Saturnispora hagleri, their closest relative. Saturnispora serradocipensis and S. gosingensis differed from S. hagleri, respectively, by seven nucleotide substitutions and two indels and three nucleotide substitutions and three indels in D1/D2 rRNA sequences. The two new species differ from each another by four nucleotide substitutions and one indel in D1/D2 rRNA sequences. However, the ITS sequences of S. serradocipensis, S. gosingensis and S. hagleri were quite divergent, showing that they are genetically separate species. The type strain of S. serradocipensis is UFMG-DC-198(T) (=CBS 11756(T) = NRRL Y-48717(T)), and of S. gosingensis GA4M05(T) is (CBS 11755(T) = NRRL Y-48718(T)).

Polyurethanes (PU) with suitable soft segments have been found to be good blood-compatible polymers and have attracted much attention recently. In this study, various molar amounts of 4,4'-methylene bisphenyl isocyanate reacted with poly(tetramethy1ene oxide) were synthesized to explore the optimal ratio of hard/soft segments for cell attachment and proliferation in in uitro systems. Differential scanning calorimetry and dynamic mechanical analysis were used to determine the physical properties, hydrogen bonding index (HBI) and transmission electron microscopy to observe the phase-separation phenomena in the materials, and 3T3 fibroblast to evaluate the dependence of the cell proliferation at 37°C on the material properties. Our results show that cell attachment and proliferation are closely related to the cell growth surface, which in turn is controlled by (1) the ratio of hard to total segment concentration and (2) the recrystallization temperature (T,) of PU. To obtain a good cell growth surface, the ratio of hard to total segment concentration is found to be between 0.4 and 0.6, and HBI is between 1.5 and 2.1. Furthermore, when the T, of PU is near the physiology temperature, a stable surface for cell growth can be provided. The shorter molecules in the soft segment region can rearrange the molecular chain at 37°C. K e y words: polyurethane, cell growth, phase separation, hydrogen bonding index, cytotoxicity.

Moth orchids (Phalaenopsis) are among the top-traded blooming potted plants in the world. To explore mitochondrial DNA (mtDNA) markers for species identification, we located simple sequence repeats in the mtDNA of Phalaenopsis aphrodite subsp. formosana and then pre-screened them for polymorphic markers by their comparison with corresponding mtDNA regions of P. equestris. The combination of 13 selected markers located in intergenic spacers could unambiguously distinguish 15 endemic moth orchids. Five most variable markers with polymorphic information content (PIC) ≥ 0.7 could be combined to classify 18 of 19 endemic moth orchids including parental strains most commonly used in breeding programs. The sequences of four selected mtDNA regions were highly variable, and one region (MT2) could be used to completely distinguish 19 endemic moth orchids. Though mitochondrial introns were highly conserved among moth orchids, evolutionary hotspots, such as variable simple sequence repeats and minisatellite repeats, were identified as useful markers. Furthermore, a marker technology was applied to reveal the maternal inheritance mode of mtDNA in the moth orchids. Moreover, phylogenetic analysis indicates that the mtDNA was nonmonophyletic below the Phalaenopsis genus. In summary, we have revealed a set of mtDNA markers that could be used for identification and phylogenetic study of Phalaenopsis orchids.