Fungi on submerged wood in the Koito River,Japan

 Woody substrates were collected from the Koito River in Japan, and the biodiversity of fungi on these substrates was investigated. Twenty-eight species were identified, comprising 12 ascomycete and 16 anamorphic taxa. The common fungi included Chaetosphaeria sp., Ophioceras commune, Pseudohalonectria lignicola, and Savoryella lignicola. The occurrence of fungi on submerged wood is discussed, and three interesting taxa – Pseudohalonectria lignicola, Bactrodesmium arnaudii, and B. pallidum – are described and illustrated. Received: August 19, 2002 / Accepted: November 25, 2002 Acknowledgments We are grateful to the financial support of the Special Coordination Fund for Promoting Science and Technology from the Ministry of Education, Culture, Sports, Science and Technology, Japan. Profs. K. Nishimura and M. Miyaji are thanked for scientific invitation and assistance in Chiba, Japan. Dr. Norio Hayashi from the Natural History Museum and Institute, Chiba, is thanked for providing physicochemical data for the Koito River. Y.M. Leung and M.H. Ng are thanked for technical assistance. Correspondence to:C.K.M. Tsui     

Purification and properties of β-amylase from Bacillus circulans S31

A thermotolerant -amylase was purified from Bacillus circulans S31 isolated from soil in Hong Kong. The purified enzyme has an M _r of 64 kDa and was stable at 50°C and pH 7.0 for 30 min. Its K _m for starch was 0.9 mg/ml with a V _max of 0.3 mg/min. It was not activated by any metal ion although sulphydrys reagents were inhibitory.H.S. Kwan, K.H. So and K.Y. Chan are with the Department of Biology, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong S.C. Cheng is with the Department of Applied Biology and Chemical Technology, Hong Kong Polytechnic, Hung Hom, Hong Kong.     

Pleosporales in Japan (1): the genus <Emphasis Type="Italic">Lophiostoma</Emphasis>

 Seven species of the genus Lophiostoma were the subject of this study. Among these, Lophiostoma mucosum is described and illustrated as a new species. All other species, L. macrostomum, L. semiliberum, L. arundinis, L. caulium, L. caudatum, and L. winteri, are reported for the first time in Japan. A key to the species of Lophiostoma in Japan is given. Received: August 5, 2002 / Accepted: November 28, 2002 Acknowledgments We are grateful to Dr. Hideki Takahashi (curator of SAPA) for the loan of fungal material. Correspondence to:Y. Harada     

<Emphasis Type="Italic">Erysiphe patagoniaca</Emphasis>: a new species of <Emphasis Type="Italic">Erysiphe </Emphasis>sect. <Emphasis Type="Italic">Uncinula </Emphasis>from Patagonia,Argentina

 A new species of Erysiphe sect. Uncinula is described and illustrated from Patagonia, Argentina. Erysiphe patagoniaca sp. nov., found on leaves of Nothofagus × antarctica, is similar to E. nothofagi and E. kenjiana, but differs in its appendages being twisted throughout their length and the number of appendages, asci, and ascospores. The two endemic species of Erysiphe sect. Uncinula, E. magellanica and E. nothofagi, coexisted on the same leaves together with Erysiphe patagoniaca. Received: September 19, 2002 / Accepted: November 28, 2002 Acknowledgments The authors are grateful to Ms. Seiko Niinomi for providing the micrographs of ascomata of Erysiphe spp. on Nothofagus. Correspondence to:S. Takamatsu     

Competition between n-alkane-assimilating yeasts and bacteria during colonization of sandy soil microcosms

An n-alkane-assimilating strain of Candida tropicalis was selected in sandy soil inoculated with microorganisms from contaminated sites. Competition experiments with n-alkane utilizers from different strain collections confirmed that yeasts overgrow bacteria in sandy soil. Acidification of the soil is one of the colonization factors useful for the yeasts. It can be counteracted by addition of bentonite, a clay mineral with high ion exchange capacity, but not, however, by kaolin. Strains of different yeast species showed different levels of competitiveness. Strains of Arxula adeninivorans, Candida maltosa, and Yarrowia lipolytica overgrew strains of C. tropicalis, C. shehatae or Pichia stipitis. Two strains of C. maltosa and Y. lipolytica coexisted during several serial transfers under microcosm conditions. Received: 20 October 1999 / Received revision: 26 January 2000 / Accepted: 27 January 2000     

Pleosporales in Japan (3). The genus Massarina

 Twelve species of the genus Massarina collected from Japan are reported in this article. Among them, 4 new species, M. constricta, M. japonica, M. submediana, and M. uniserialis, are described, illustrated, and compared to similar species. Two species, M. scirpina and M. ryukyuensis, are described as new combination, and 4 species, M. arundinariae, M. fluviatilis, M. peerallyi, and M. rubi, are reported from Japan for the first time. One bambusicolous species, Metasphaeria tuberculosa, is considered to be a synonym of Massarina bambusina. Received: December 13, 2002 / Accepted: February 6, 2003 Acknowledgments We are grateful to Dr. Shuhei Tanaka, curator of YAM (Yamaguchi University), for the loan of specimens for this study.     

Isolation and characterization of restriction endonuclease EclHKI from an Enterobacter cloacae strain

An Enterobacter cloacae strain, producing restriction enzyme EclHKI, was isolated from a decaying potato. The enzyme is an isoschizomer of Eam1105I, which recognizes and cleaves GACNNN!NNGTC. EclHKI was produced at high activity (4×10⁴ U/g wet cells) and was purified from contaminants which interfere with restriction digestion by passing the cell lysate through DEAE-Sephacel and heparin columns. Activity was optimal at 37°C in a medium salt buffer.H.-Y. Chan, Y.-C. Chan and P.-C. Shaw are with the Department of Biochemistry, The Chinese University of Hong Kong, Hong Kong: K.-M. Kam is with the Institute of Pathology, Sai Ying Pun Jockey Club Clinic, Hong Kong.     

Pleosporales in Japan (2): the genus <Emphasis Type="Italic">Lophiotrema</Emphasis>

 Five species of the genus Lophiotrema are reported in this article. Of these, Lophiotrema vitigenum is described and illustrated as a new species. Three species, L. nucula, L. neohysterioides, and L. vagabundum, are newly added to the Japanease mycoflora. Lophiosphaera orientalis, Nodulosphaeria araucariae, and N. rosae, which had been recorded in Japan, are treated as synonyms of Lophiotrema fuckelii. A key to the species of Lophiotrema in Japan is given. Received: October 9, 2002 / Accepted: January 20, 2003 Acknowledgments We are grateful to Drs. Shuhei Tanaka (curator of YAM) and Ken Katumoto for the loan of fungal material. Correspondence to:K. Tanaka     

High constitutive peroxidase activity and constitutive thermotolerance in <Emphasis Type="Italic">Neurospora crassa</Emphasis>

 During the isolation of mutations in the heat-inducible hsp70-1 gene of Neurospora crassa by RIP (repeat-induced point mutations), several transformants were generated by electroporation of conidia with a plasmid harboring an incomplete copy of this gene. One isolate, designated E-45, containing ectopically integrated hsp70-1 DNA, exhibited a slow growth rate, low-temperature sensitivity, constitutive thermotolerance (without prior heat shock), and high constitutive peroxidase activity. The constitutive form of peroxidase (CP) was distinguishable from the heat-inducible form (HIP) by immunoinactivation employing polyclonal antiserum against the latter enzyme and by electrophoretic resolution in nondenaturing polyacrylamide gels. This enzyme was purified to near homogeneity and some of its properties examined. The relative molecular mass of native CP was in the range of 118–136 kDa, as estimated by gel filtration analysis on size exclusion matrices, whereas SDS-PAGE analysis yielded a size of ∼37 kDa for the polypeptide. Substrate saturation kinetics studies were conducted using ABTS [2,2′-azino-bis (3-ethylbenzthiazole-6-sulfonic acid)] and H₂O₂ as substrates: K _m, V _max, and K _cat values for H₂O₂ were ∼22 μM, ∼447 nmol mg⁻¹, and 0.33 s⁻¹, respectively, and those for ABTS were ∼55 μM, ∼453 nmol mg⁻¹, and 0.3 s⁻¹, respectively. Guaiacol was not used as a substrate by this enzyme. CP peroxidase was shown to be a heme-containing enzyme, stable at temperatures up to 58°C. Received: August 5, 2002 / Accepted: January 22, 2003 Acknowledgments This work was supported by an operating grant from the Natural Sciences and Engineering Research Council (NSERC) of Canada (to M.K.). The financial support provided to A. M. in the form of a graduate studentship award by the AHFMR (Alberta Heritage Foundation for Medical Research) and of a graduate teaching assistantship to A. S. by the Department of Biological Sciences, University of Calgary, is gratefully acknowledged. Correspondence to:M. Kapoor     

In Vitro Activities of New Triazole Antifungal Agents, Posaconazole and Voriconazole, Against Oral Candida Isolates from Patients Suffering from Denture Stomatitis

Denture stomatitis is often treated with antifungal agents but recurrences or new episodes are common, and certain episodes can be resistant. New triazoles, such as posaconazole and voriconazole, may represent useful alternatives for management. In vitro activities of amphotericin B, nystatin, miconazole, fluconazole, itraconazole, posaconazole and voriconazole against 150 oral Candida (101 C. albicans, 18 C. tropicalis, 12 C. glabrata, 11 C. guilliermondii, 4 C. parapsilosis, 2 Saccharomyces cerevisiae, 1 C. dubliniensis and 1 C. krusei) from 100 denture wearers were tested by the CLSI M27-A3 method. Resistant isolates were retested by Sensititre YeastOne and Etest. Most antifungal agents were very active. However, 4 C. glabrata (33.3%), 2 C. tropicalis (11.1%), 6 C. albicans (5.6%) and 1 C. krusei were resistant to itraconazole. Posaconazole was active against 143 yeast isolates (95.3%): 6 C. albicans (5.9%) and 1 C. tropicalis (5.6%) were resistant. Geometric mean MICs were 0.036 μg/ml for C. parapsilosis, 0.062 μg/ml for C. albicans, 0.085 μg/ml for C. tropicalis, 0.387 μg/ml for C. guilliermondii and 0.498 μg/ml for C. glabrata. Voriconazole was active against 148 isolates (98.7%) with geometric mean MICs ranging from 0.030 μg/ml for C. parapsilosis, 0.042 μg/ml for C. albicans, 0.048 μg/ml for C. tropicalis, 0.082 μg/ml for C. guilliermondii, to 0.137 μg/ml for C. glabrata. Only 2 C. albicans (2%) were resistant to voriconazole showing cross-resistance to other azoles. Posaconazole and voriconazole have excellent in vitro activities against all Candida isolates and could represent useful alternatives for recalcitrant or recurrent candidiasis.     

Antifungal Susceptibility and Virulence Attributes of Bloodstream Isolates of <Emphasis Type="Italic">Candida</Emphasis> from Hong Kong and Finland

Candida bloodstream infection has dramatically increased in the last decade due to the growing number of immunocompromised populations worldwide. In this study, we evaluated the antifungal susceptibility profiles and virulence attributes of Candida bloodstream isolates (CBIs) derived from Hong Kong and Finland, information which are vital for devising empirical clinical strategies. Susceptibility testing of a wide range of antifungals including fluconazole, itraconazole, voriconazole, ketoconazole, 5-fluorocytosine, amphotericin B and caspofungin was performed. Haemolytic activity and secretion of proteinase of CBIs were also examined. All CBIs derived from Hong Kong were susceptible to all the antifungals tested whilst some CBIs from Finland were resistant to azoles and caspofungin. C. albicans, C. glabrata and C. tropicalis showed higher haemolytic activity whereas C. parapsilosis and C. guilliermondii were non-haemolytic in general. Proteinase activity of the Finland C. albicans isolates was significantly higher than the Hong Kong isolates. Our data provide a glimpse of the possible evolutionary changes in pathogenic potential of Candida that may be occurring in different regions of the world. Therefore, continuous surveillance and availability of local data should be taken into consideration when treating candidemia patients.     

Radiation and reticulation: extensive introgressive hybridization in the carabid beetles Ohomopterus inferred from mitochondrial gene genealogy

 In the evolutionary process of an animal lineage, interactions in secondary contacts of differentiated populations and introgressive hybridization may play an important role. In the Japanese islands, the carabid subgenus Ohomopterus (genus Carabus) exhibits a marked differentiation in body size and genital morphology. Although geographical differentiation is apparent, two or three species usually coexist at many localities. Their reproductive isolation relies on body size differences, chemical cues for mate recognition, and a species-specific genital lock-and-key system. However, these isolation mechanisms are not always effective enough to prevent interspecific hybridization. An initial assessment of the species-level phylogeny with mitochondrial gene sequences revealed that the gene genealogy is highly inconsistent with the morphology-based taxonomy. A comparison of mitochondrial and nuclear gene genealogies showed that these are strongly incongruent with each other, while the nuclear gene genealogy is more consistent with traditional taxonomy, indicating the repeated occurrence of introgression of mitochondria across species. Here, two different cases of mitochondrial introgression among Ohomopterus species are described in detail, one for parapatric species and the other for sympatric species. First, mitochondrial haplotypes and sequences were studied in Carabus insulicola and three taxa parapatric with C. insulicola, at least two of which hybridize with C. insulicola naturally. Among the four species studied, directional introgressions of mitochondria across boundary zones were detected. Second, in the Mt. Kongo area in central Honshu, which harbors five species, introgression of mitochondria among four out of the five species was detected, despite the apparent absence of on-going natural hybridization. These inferred cases of mitochondrial introgression indicate that species interactions through hybridization could have played an important role at various stages in the evolution of Ohomopterus. Received: April 12, 2002 / Accepted: October 17, 2002 Acknowledgments I am grateful to Alfried P. Vogler for a long-lasting collaboration in the molecular phylogenetic study of Ohomopterus. R. Ishikawa, K. Kubota, M. Ujiie, Y. Takami, and F. Kusumoto have also collaborated at various stages of this study. Thanks are also due to K. Miyashita, T. Funakoshi, H. Fujimoto, T. Dejima, Y. Nagahata, T. Miyagawa, K. Yodoe, H. Kadowaki, S. Nakamine, Y. Oka, H. Tanaka, T. Tanabe, K. Kusakari, and T. Okumura for their care of specimens. Supported by grants-in-aid from the Japan Society for the Promotion of Science (Nos. 09640748, 11304056).     

Identification and production of 3-hydroxy-Δ9-cis-1,18-octadecenedioic acid by mutants of Candida tropicalis

 The transformation of oleic acid by mutants of Candida tropicalis was studied in fed-batch cultures. Besides Δ⁹-cis–1,18-octadecenedioic acid, 3-hydroxy-Δ⁹-cis–1,18-octadecenedioic acid was detected as the main fermentation product. Here we describe the production, isolation and the complete chemical characterization of the purified 3-hydroxy-Δ⁹-cis–1,18-octadecenedioic acid. The geometric configuration of the double bond was not changed during bioconversion. The enantiomeric excess of the compound was 76%. Mutagenesis of C. tropicalis DSM 3152 with N-methyl-N-nitro-N′-nitrosoguanidine and selection with oleic acid as the sole carbon source led to mutant M 25, which produced the 3-hydroxy-Δ⁹-cis–1,18-octadecenedioic acid at a 1.8-fold higher concentration in the medium as compared to the parent strain. The maximum concentration of the hydroxy dioic acid was 19.4 g/l after 223 h fermentation. Received: 24 August 1995/Received revision: 21 September 1995/Accepted: 4 October 1995     

Primary structure and properties of the formyltransferase from the mesophilic Methanosarcina barkeri: comparison with the enzymes from thermophilic and hyperthermophilic methanogens

The ftr gene encoding formylmethanofuran: tetrahydromethanopterin formyltransferase (Ftr) from Methanosarcina barkeri was cloned, sequenced, and functionally expressed in Escherichia coli. The overproduced enzyme was purified eightfold to apparent homogeneity, and its catalytic properties were determined. The primary structure and the hydropathic character of the formyltransferase from Methanosarcina barkeri were compared with those of the enzymes from Methanobacterium thermoautotrophicum, Methanothermus fervidus, and Methanopyrus kandleri. The amino acid sequence of the enzyme from Methanosarcina barkeri was 64%, 61%, and 59% identical to that of the enzyme from Methanobacterium thermoautotrophicum, Methanothermus fervidus, and Methanopyrus kandleri, respectively. A negative correlation between the hydrophobicity of the enzymes and both the growth temperature optimum and the intracellular salt concentration of the four organisms was observed. The hydrophobicity of amino acid composition was +21.6 for the enzyme from Methanosarcina barkeri (growth temperature optimum 37° C, intracellular salt concentration ≈ 0.3 M), +9.9 for the enzyme from Methanobacterium thermoautotrophicum (65°C, ≈ 0.7 M), –20.8 for the enzyme from Methanothermus fervidus (83° C, ≈ 1.0 M) and –31.4 for the enzyme from Methanopyrus kandleri (98° C, > 1.1 M). Generally, a positive correlation between hydrophobicity and thermophilicity of enzymes and a negative correlation between hydrophobicity and halophilicity of enzymes are observed. The findings therefore indicate that the hydropathic character of the formyltransferases compared is mainly determined by the intracellular salt concentration rather than by temperature. Sequence similarities between the formyltransferases from methanogens and an open reading frame from Methylobacterium extorquens AM1 are discussed. Received: 7 September 1995 / Accepted: 7 November 1995     

A new species ofClohiesia from Hong Kong

Clohiesia lignicola sp. nov. (freshwater ascomycetes) is introduced based on a specimen collected on submerged wood in the Tung Chung River, Hong Kong. Ascomata are clypeate, asci are cylindric-clavate with a relatively massive apical apparatus and ascospores are fusoid-ellipsoidal.Clohiesia lignicola differs fromC. corticola in having wider asci and wider fusoid-ellipsoidal ascospores, and larger ascomata.Clohiesia lignicola is described and illustrated with light micrographs and is also compared with species in the genus,Annulatascus.