Cryoprotective properties and preliminary characterization of exopolysaccharide (P-Arcpo 15) produced by the Arctic bacterium Pseudoalteromonas elyakovii Arcpo 15

Twenty-two bacterial strains that secrete exopolysaccharides (EPS) were isolated from marine samples obtained from the Chukchi Sea in the Arctic Ocean; of these, seven strains were found to be capable of producing cryoprotective EPS. The ArcPo 15 strain was isolated based on its ability to secrete large amounts of EPS, and was identified as Pseudoalteromonas elyakovii based on 16S rDNA analysis. The EPS, P-ArcPo 15, was purified by protease treatment and gel filtration chromatography. The purified EPS (P-ArcPo 15) had a molecular mass of 1.7 × 10⁷ Da, and its infrared spectrum showed absorption bands of hydroxyl and carboxyl groups. The principal sugar components of P-ArcPo 15 were determined to be mannose and galacturonic acid, in the ratio of 3.3:1.0. The cryoprotective properties of P-ArcPo 15 were characterized by an Escherichia coli viability test. In the presence of 0.5% (w/v) EPS, the survival percentage of E. coli cells was as high as 94.19 ± 7.81% over five repeated freeze–thaw cycles. These biochemical characteristics suggest that the EPS P-ArcPo 15 may be useful in the development of cryoprotectants for biotechnological purposes, and we therefore assessed the utility of this novel cryoprotective EPS.     

Common genotypes (RFLP) within a diverse collection of yellow-green aspergilli used to produce traditional Oriental fermented foods

 DNA fingerprinting was performed on 72 strains of Aspergillus oryzae and 9 strains of Aspergillus sojae isolated from chu (China) or koji (Japan) mold inoculum used in the production of traditional Oriental fermented beverages or foods including soy sauce, miso, and sake. The cultures were deposited with the ARS Culture Collection (NRRL) between 1909 and 2001. PstI digests of total genomic DNA from each isolate were probed using the pAF28 repetitive sequence. All strains of A. sojae that we examined produced an identical DNA fingerprint and belong to the same DNA fingerprint group (GTAo-9). Strains of A. oryzae were distributed among 41 DNA fingerprint groups, including GTAo-12 represented by 11 strains, GTAo-19 represented by 5 strains, GTAo-5 and GTAo-15 each represented by 4 strains, and GTAo-8, GTAo-17, and GTAo-24 each represented by 3 strains. Thirty-three single strain isolates of A. oryzae produced unique fingerprints. Our data offer evidence suggesting that (1) the successful domestication of A. parasiticus genotypes yielding A. sojae occurred far less frequently than among genotypes of A. flavus var. oryzae; and (2) some Aspergillus genotypes employed in different fermentations and regions were derived from a common ancestral clonal population. Received: February 18, 2002 / Accepted: April 22, 2002     

Genotypic Diversity of Oscillatoriacean Strains Belonging to the Genera <Emphasis Type="Italic">Geitlerinema</Emphasis> and <Emphasis Type="Italic">Spirulina</Emphasis> Determined by 16S rDNA Restriction Analysis

Genotypic diversity of several cyanobacterial strains mostly isolated from marine or brackish waters, belonging to the genera Geitlerinema and Spirulina, was investigated by amplified 16S ribosomal DNA restriction analysis and compared with morphological features and response to salinity. Cluster analysis was performed on amplified 16S rDNA restriction profiles of these strains along with profiles obtained from sequence data of five Spirulina-like strains, including three representatives of the new genus Halospirulina. Our strains with tightly coiled trichomes from hypersaline waters could be assigned to the Halospirulina genus. Among the uncoiled strains, the two strains of hypersaline origin clustered together and were found to be distant from their counterparts of marine and freshwater habitat. Moreover, another cluster, formed by alkali-tolerant strains with tightly coiled trichomes, was well delineated. Received: 14 May 2002 / Accepted: 22 July 2002     

Isolation of freeze-tolerant laboratory strains of Saccharomyces cerevisiae from proline-analogue-resistant mutants

Since some amino acids, polyols and sugars in cells are thought to be osmoprotectants, we expected that several amino acids might also contribute to enhancing freeze tolerance in yeast cells. In fact, proline and charged amino acids such as glutamate, arginine and lysine showed a marked cryoprotective activity nearly equivalent to that of glycerol or trehalose, both known as major cryoprotectants for Saccharomyces cerevisiae. To investigate the cryoprotective effect of proline on the freezing stress of yeast, we isolated proline-analogue-resistant mutants derived from a proline-non-utilizing strain of S. cerevisiae. When cultured in liquid minimal medium, many mutants showed a prominent increase, two- to approximately tenfold, in cell viability compared to the parent after freezing in the medium at −20 °C for 1 week. Some of the freeze-tolerant mutants were found to accumulate a higher amount of proline, as well as of glutamate and arginine which are involved in proline metabolism. It was also observed that proline-non-utilizer and the freeze-tolerant mutants were able to grow against osmotic stress. These results suggest that the increased flux in the meta-bolic pathway of specific amino acids such as proline is effective for breeding novel freeze-tolerant yeasts. Received: 6 November 1996 / Accepted: 7 December 1996     

Influence of cryopreservation on the pulpal tissue of immature third molars in vitro

The purpose of this study was to evaluate in vitro the viability of isolated and non-isolated pulpal tissue of immature third molars after cryopreservation. This study was divided in three different experiments. Experiment 1: Pulpal tissue isolated from 19 third molars was divided in horizontal segments. Each segment was cultured separately in order to evaluate whether differences in growth capacity within the tissue could be found. Experiment 2: Pulpal tissue isolated from 27 third molars was divided in a mesial and a distal part. One part was cryopreserved before culturing, the other part was cultured immediately. Growth capacity of cryopreserved and non-cryopreserved tissue was evaluated and compared. Experiment 3: 43 third molars were cryopreserved. After thawing, the dimension of the apical foramen was measured and the pulp was isolated and segmented horizontally. The different parts were cultured and growth capacity was evaluated and compared. Results of experiment 1 and 2 showed no significant difference in growth capacity between fibroblasts originating from different pulpal segments of the same tooth without cryopreservation and between fibroblasts originating from cryopreserved and non-cryopreserved isolated pulpal tissue. In experiment 3 it was demonstrated that the dimension of the apical foramen and pulpal viability after cryopreservation are positively correlated. A minimum dimension of 9.42 mm2 enables the cryoprotective agent to penetrate sufficiently and to protect the pulpal tissue from apex to crown. This study proved that cryopreservation of human pulpal tissue is possible if the cryoprotective agent can reach the entire pulp.     

Analysis of pristinamycin-resistant Staphylococcus epidermidis isolates in the Tunisian Bone Marrow Transplant Center

Aims: We report the analysis of genetic determinants conferring resistance to pristinamycin in Staphylococcus epidermidis strains and epidemiology typing of these strains by pulsed‐field gel electrophoresis. Methods and Results: Staphylococcus epidermidis (346 isolates) were searched for strains with pristinamycin resistance. Pristinamycin‐resistant strains (seven isolates) were isolated in five patients with haematological cancer in the Bone Marrow Transplant Centre of Tunisia in 2002. Resistance to pristinamycin was observed in 2% of isolates. The seven pristinamycin‐resistant strains shared resistance to oxacillin (MIC = 8–512 μg ml^?1), gentamicin (MIC = 16–512 μg ml^?1), erythromycin (MIC > 1024 μg ml^?1), lincomycin (MIC > 1024 μg ml^?1), pristinamycin (MIC = 4–16 μg ml^?1) and rifampin (MIC = 128–256 μg ml^?1). erm genes were amplified: ermA from six strains and ermC from one. vga gene encoding streptogramins A resistance (pristinamycin résistance) was amplified from all strains and typed as vgaA by analysis after electrophoresis of restriction profiles of vga amplicons (two fragments with Sau3A of 164 and 378 bp; one fragment with EcoRI). Pulsed‐field gel electrophoresis (PFGE) of SmaI chromosomal DNA digests of the seven S. epidermidis isolates divided them into two distinct pattern types: pulsed‐field type A (classified from A1 to A6 subtypes) and type B. The six strains harbouring ermA genes belonged to the PFGE type A while the strain harbouring ermC genes belonged to the PFGE type B. We characterized an epidemic strain carrying the vgaA and ermA genes responsible for the outbreak. Conclusions: Two clones of pristinamycin‐resistant S. epidermidis were isolated in our patients. One of them, isolated in all patients, had expanded over six months suggesting acquisition by cross‐contamination. Significance and Impact of the study: Increasing isolation of pristinamycin resistant S. epidermidis strains is an alarming indicator of nosocomial dissemination. The vector will be determined to establish a system of epidemiological surveillance.     

Survival and nutritional requirements of three bacteria isolated from ultrapure water

Bacteria isolated previously from ultrapure water (UPW) systems were examined for their ability to survive in UPW, with the ultimate goal of elucidating potential carbon and energy sources for the bacteria. Two strains of Ralstonia pickettii isolated from different areas within the UPW system (pretreatment and polishing loop, and referred to as strains 3A1 and MF254A, respectively) and a strain of Bradyrhizobium sp. were compared to increase our understanding of the fundamental behavior of bacteria contaminating UPW. R. pickettii (3A1) grew significantly slower in R2A medium, with a final cell yield much lower than the isolate from the polishing loop. In addition, R. pickettii MF254A showed a broader substrate range than either strain 3A1 or Bradyrhizobium sp. In UPW, there appears to be a threshold cell concentration (approximately 10⁶ colony-forming units/ml), whereby the cell numbers remain constant for a prolonged period of 6 months or more. Below this concentration, rapid proliferation is observed until the threshold concentration is attained. Preliminary experiments suggested that nitrogen gas (frequently added to UPW storage tanks) may contribute to growth of Bradyrhizobium sp. Above the threshold concentration, the strain of Ralstonia sp. isolated from the polishing loop was capable of cryptic growth with heat-killed cells in UPW. However, cryptic growth was not observed when the cells supplied as nutrients were killed using UV254 light. Furthermore, cryptic growth did not appear to contribute significantly to proliferation of Bradyrhizobium sp. or Ralstonia sp. 3A1 (isolated from the pretreatment loop). We believe that cryptic growth may aid survival of the bacteria in UPW, but further experiments are warranted to prove this phenomenon conclusively. Journal of Industrial Microbiology & Biotechnology (2002) 29, 75–82 doi:10.1038/sj.jim.7000273 Received 23 January 2002/ Accepted in revised form 29 April 2002     

Identification and Characteristics of Nisin Z-Producing <Emphasis Type="Italic">Lactococcus lactis</Emphasis> subsp. <Emphasis Type="Italic">lactis</Emphasis> Isolated from Kimchi

We isolated bacteriocin-producing Lactococcus lactis subsp. lactis from Kimchi. The bacteriocin inhibited strains of Clostridium perfringens, C. difficile, Listeria monocytogenes, vancomycin-resistant Enterococcus, and one out of four methicillin-resistant Staphylococcus aureus strains, as well as some closely related lactic acid bacteria. In tricine-SDS-PAGE, the bacteriocin migrated with an apparent molecular weight of about 4 kDa to the same location as nisin A and crude nisin Z. The gene encoding this bacteriocin was found to be identical to that of nisin Z with direct PCR sequence methods. The inhibitory activity was stable against heat and pH, but it was lost at 100°C for 1 h and at 121°C for 15 min. The bacteriocin was inactivated by proteolytic enzymes, but was not affected by lysozyme, lipase, catalase, or β-glucosidase. There were some differences in characteristics from those of nisins described previously. Received: 21 June 2002 / Accepted: 22 July 2002     

Homogeneity of the Morphological Groups of Bacteriophages Infecting <Emphasis Type="Italic">Bacteroides fragilis</Emphasis> Strain HSP40 and Strain RYC2056

Bacteriophages infecting Bacteroides fragilis strains RYC2056 and HSP40 have been proposed as indicators of water quality. To accomplish this function, homogeneity of the group of phages detected by these strains is necessary to ensure that the final results are not due to the different kinetics of inactivation of the phages. To evaluate homogeneity, we observed by electron microscopy bacteriophages isolated from sewage with two Bacteroides fragilis strains (HSP40 and RYC2056). A predominant group of phages was observed, Siphoviridae with slightly curved tails. Detection of other minority groups, which could be present in the sample, was done with neutralization experiments by using antiserum against the majority group and with host mutants resistant to infection with the predominant phage. Although two other minority groups were observed, results showed that bacteriophages infecting B. fragilis strain HSP40 and strain RYC2056 form a homogeneous group, Siphoviridae with slightly curved tails being the most predominant in sewage. Received: 7 March 2002 / Accepted: 5 August 2002     

Detection of <Emphasis Type="Italic">Actinobacillus pleuropneumoniae</Emphasis> by PCR on Field Strains from Healthy and Diseased Pigs

We investigated whether primers able to specifically amplify a 0.7-kb DNA fragment from the conserved cpx genes could be applied to analyze A. pleuropneumoniae field isolates. The specific cpx primers were tested on 120 strains of A. pleuropneumoniae and other NAD-dependent field isolates from healthy and diseased animals to analyze A. pleuropneumoniae isolates from pigs in Brazil. We found that PCR and hybridization were able to discriminate between isolates of A. pleuropneumoniae and other bacteria. The 0.7-kb cpx DNA fragments were amplified from all 63 A. pleuropneumoniae isolates from herds with clinical symptoms and were isolated from lesions of acute cases of swine pleuropneumonia, both serotypable and nonserotypable. The PCR was also applied to 57 field isolates obtained from animals of apparently healthy herds, and the amplified cpx product was present in four serotypable and only two out of eleven A. pleuropneumoniae nonserotypable isolates. All nonserotypable A. pleuropneumoniae isolates revealed the apxA amplification pattern compatible with previously known serotypes. Some nonserotypable isolates might represent a population of isolates that originally were serotypable but lost the ability to react with serotype-specific antisera or might belong to novel serotypes. The PCR method applied is highly sensitive for serotypable A. pleuropneumoniae strains and for nonserotypable strains isolated from acute cases of swine pleuropneumoniae in Brazil. Received: 13 June 2002 / Accepted: 5 August 2002     

Selection of Portuguese Rhizobium leguminosarum bv. trifolii strains for production of legume inoculants

From several native clover species, growing in six different soil types, 170 Rhizobium leguminosarum biovar trifolii strains were isolated, covering the central and southern regions of Portugal. The effectiveness of the strains varied from ineffective to highly effective on T. subterraneum cv. Clare and on T. fragiferum cv. Palestine, with a predominance of medium and high effectiveness on both host plants. The effectiveness was not influenced by provenence (soil or plant), except for the strains from the rankers soils and for the strains isolated from T. pratense, that were ineffective or medium effective on T. subterraneum.Selected strains were evaluated for effectiveness on T. subterraneum cv. Clare, using the commercial strain TA1 as reference. Several of the isolated strains were more effective than TA1, indicating that local strains may be used to produce better inoculants.     

Physiological and molecular detection of crystalliferous Bacillus thuringiensis strains from habitats in the South Central United States

Gram-positive, endospore-forming Bacillus thuringiensis-like strains were isolated from 95 of 413 samples collected at the 0–5 cm depth of noncultivated soils and stagnant or dried-up ponds as well as from dust from stored grain products in South Central United States. Based on the production of parasporal crystals, 25 isolates were identified as B. thuringiensis after examining 227 B. thuringiensis-like colonies. The greatest proportion of samples yielding B. thuringiensis were from the dust from grain storage. The sodium acetate selective medium, heat processing, and crystal staining used in the initial screening revealed diverse populations of B. thuringiensis, which were categorized into distinct crystal morphological groups. Sugar fermentation, antibiotic sensitivity, growth characteristics and PCR studies showed diversity among the isolates that were distributed among 25 of the 58 known strains. The most frequently isolated strains were kurstaki, aizawai, morrisoni, thuringiensis, sotto and kenyae that together represented more than 90% of the characterized isolates. PCR analysis using 30 family primer pairs for cry and cyt genes showed that the frequency of the cry1 gene (62%) was predominant followed by the cry2 genes (30%), and the rest (8%) were other cry gene types, such as cry3, cry4, cry10, cry11, cry14, cry15, cry20, cry24 and cry26. Both cyt1 and -2 genes were also detected. Several isolates showed PCR products on the gel that were not consistent with the expected sizes of nucleotides targeted by the primers. These were suggestive of nonspecific amplifications and were not used in the characterization process. Journal of Industrial Microbiology & Biotechnology (2002) 28, 284–290 DOI: 10.1038/sj/jim/7000244 Received 30 May 2001/ Accepted in revised form 10 January 2002     

Numerical Taxonomy of <Emphasis Type="Italic">Vibrionaceae</Emphasis> Isolated from Cultured Amberjack (<Emphasis Type="Italic">Seriola dumerili</Emphasis>) and Surrounding Water

A numerical taxonomic study was performed on 148 isolates of Gram-negative, heterotrophic, facultative anaerobic bacteria isolated from amberjack (Seriola dumerili) and its surrounding culture water. The study included 30 type and reference strains belonging to genera Vibrio, Listonella, and Photobacterium. The strains were characterized by 109 morphological, biochemical, physiological, and nutritional tests. Cluster analysis of similarity matrices obtained with S_SM and S_J coefficients was carried out. UPGMA (unweighted pair group mathematical average) analysis defined 11 phena at S_SM values ≥ 86%. Nine phena were identified as Vibrio alginolyticus, V. fischeri, V. harveyi, V. carchariae, V. mediterranei, V. splendidus, V. furnissii, V. parahaemolyticus, and Photobacterium damselae subsp. damselae. The two latter comprised strains isolated from diseased fish. Received: 27 March 2002 / Accepted: 24 May 2002     

Characterization of Cr(VI)-Resistant Bacteria Isolated from Chromium-Contaminated Soil by Tannery Activity

Bacterial strains, previously isolated from a chromium-polluted soil, were identified on the basis of Gram reaction and biochemical characteristics (Biolog system). Moreover, chromate MICs, chromate reduction capability, multiple heavy metal tolerance, and antibiotic susceptibility were tested for each isolate. All strains but one were Gram-positive and resistant to high concentrations of chromate. The most Cr(VI)-resistant isolate (22mM) was identified as Corynebacterium hoagii. All Cr(VI)-resistant strains except the isolate ChrC20 were capable of catalyzing the reduction of Cr(VI) to Cr(III), a less toxic and less water-soluble form of chromium. The only isolate Cr(VI)-sensitive, attributed to the Pseudomonas genus, also exhibited Cr(VI)-reduction. Isolates were also screened for the presence of plasmid DNA. The strains ChrC20 and ChrB20 harbored one and two plasmids of high molecular mass, respectively. This approach permitted selection of some bacterial strains, which could be used for bioremediation of Cr(VI)-polluted environments. Received: 21 February 2002 / Accepted: 27 March 2002     

Characterization of 4-nonylphenol-degrading bacterial consortium obtained from a textile wastewater pretreatment plant

4-Nonylphenol (4-NP) isomers are toxic and recalcitrant compounds often resulting, together with short-chain ethoxylated nonylphenol (NPnEO, where n is the number of ethylene oxide units), from NPnEO biodegradation in conventional activated sludge plants. In this work, a microbial consortium, defined as Consortium A, capable of removing 100 mg/L of 4-NP with no accumulation of metabolites with aromatic moiety was isolated from textile wastewaters after enrichment with 4-NP. The consortium showed remarkable degradation activities toward several short-chain NPnEO congeners. Culture-dependent techniques were used to isolate from the consortium twenty-six strains assigned to seven different amplified ribosomal DNA restriction analysis groups. Two- and three-member cocultures were prepared with the strains showing highest 4-NP-degrading capabilities, but neither the single strains nor the cocultures were as efficient in 4-NP degradation as Consortium A. FISH was used to characterize the microbial composition of Consortium A: it evidenced a strong occurrence of Proteobacteria and, in particular, of Gammaproteobacteria along with a relevant stability of the culture. Therefore, the isolated consortium has the potential of being used in the development of a biotechnological process for the tertiary treatment of effluents of activated sludge plants fed with NPnEO-contaminated wastewaters.     

Whole cell protein profiling reiterate phylogenetic relationships among strains of Yersinia enterocolitica biovar 1A as discerned earlier by different genotyping methods

Aim: To evaluate whole cell protein profiling vis‐à‐vis genotyping to discern phylogenetic relationships among strains of Y. enterocolitica biovar 1A. Methods and Results: Whole cell protein profiling of Y. enterocolitica biovar 1A strains was performed using SDS–PAGE. Twenty‐one distinct protein profile types were obtained among a collection of 81 strains isolated from clinical and nonclinical sources. Whole cell protein profiling exhibited discriminatory index (DI) of 0·80 and clustered the strains into two distinct clonal groups. The clinical and the aquatic serotype O:6,30–6,31 strains were clustered into two discrete subgroups. Conclusions: Whole cell protein profiling displayed sufficient diversity among strains of Y. enterocolitica biovar 1A, and the phylogenetic relationships obtained were in good agreement with those established earlier by genotyping techniques. Significance and impact of the study: Whole cell protein profiling was as discriminatory as some of the genotyping methods and has the potentiality to be used as an adjunct tool to study epidemiology of Y. enterocolitica.     

Isolation of bacterial consortia for degradation of p-nitrophenol from agricultural soil

bacterial consortium has been isolated containing Pseudomonas spp. strains S1 and S2, which was able to degrade p‐nitrophenol (PNP). The strains were isolated from agricultural soil contaminated with organophosphorus pesticides. Pseudomonas spp. strain S2 could convert p‐nitrophenol to 4‐nitrocatechol (4NC) after pre‐exposure to phenol, when PNP was used as the only carbon source in the medium. Pseudomonas spp. strain S2, when mixed with strain S1 in the ratio 1:5 respectively, decolorised PNP completely.     

药用植物青蒿不同种类的内生菌抑菌活性分析

为了研究青蒿不同种类的内生菌抑制细菌和抑制真菌的活性,该研究采用组织块法和研磨法从青蒿的根、茎、叶中分离内生细菌、放线菌和真菌,以大肠埃希菌(Escherichia coli)(CICC 23657)、枯草芽孢杆菌(Bacillus subtilis)(CICC 10275)、金黄色葡萄球菌(Staphylococcus aureus)(CICC 10384)、黑曲霉(Aspergillus niger)(CICC 2487)、酿酒酵母(Saccharomyces cerevisiae)(CICC 33032)为指示菌,采用琼脂块法和双层平板法检测内生菌的抑菌活性。结果表明:(1)从青蒿植株中共分离到76株内生菌,其中内生细菌19株、内生放线菌34株、内生真菌23株。从分离部位来看,56株来自于茎段、17株来自于根段、3株来自叶片。(2)内生细菌中抑菌活性菌株占总菌株的比例最高,为95%,内生放线菌和内生真菌中抑菌活性菌株的比例分别为41%、35%。(3)内生细菌的抗菌谱较广;虽然内生放线菌的抗菌谱较窄,但其中高抗菌株较多,尤其对酿酒酵母的抑菌效果好。综上结果显示,药用植物青蒿中存在着丰富的有抑菌活性的内生菌,且不同种类的内生菌抑菌活性不同。     

Characterization of a Yersinia enterocolitica biotype 1A strain harbouring an ail gene

Aims: The chromosomal ail gene (attachment and invasion locus) is commonly used as target gene for the detection of pathogenic Y. enterocolitica strains in food testing. The ail PCR does not detect strains of biotype 1A (BT1A), which are regarded as non‐pathogenic because BT1A strains lack the virulence plasmid and chromosomally encoded virulence genes. In some recent reports, however, BT1A strains were discovered that harboured the ail gene. We isolated an ail‐positive strain and characterized this strain with phenotypic and genotypic methods to study its possible relation to pathogenic Y. enterocolitica strains. Methods and Results: The ail region of the BT1A strain was sequenced and compared with the corresponding region of nonpathogenic BT1A strains and pathogenic strains. Pulsed field gel electrophoresis (PFGE) analysis was applied revealing no similarity of the PFGE pattern of this strain to the patterns of pathogenic strains. Virulence‐gene‐based PCR analyses showed the strain to be positive for ystB, but negative for virulence genes ystA, virF and yadA. Whole‐cell MALDI‐TOF MS combined with a shrinkage discriminant analysis approach was applied and clearly classified the ail‐positive biotype 1A strain within the cluster of BT1A strains. Conclusions: PCR detection of ail sequences in food matrices should be followed by the isolation of the responsible strain and its characterization using phenotypic or genotypic methods. Significance and Impact of the Study: The ail gene may be present in Y. enterocolitica BT1A strains, which are commonly considered as nonpathogenic. Efficient methods such as PCR typing of other virulence genes or rapid MALDI‐TOF MS‐based bacterial profiling allow a more comprehensive assessment of the pathogenicity potential of Yersinia strains.     

Identification of the basidiomycetous fungus isolated from butt rot of the Japanese cypress

 To identify a basidiomycetous fungus isolated from butt rot of Chamaecyparis obtusa, Japanese cypress, its cultural features were examined, and sequences of its nuclear ribosomal 18S and ITS1–5.8S–ITS2 regions were analyzed. In culture, this fungus is characterized by the occurrence of chlamydospores, blastoconidium-like cells, and clavate-to-spathulate hyphal ends at the tips of aerial hyphae, and production of a small basidioma on the mycelial mat after 3 months of incubation. The morphological features of the basidioma are identical to those of Phlebia brevispora. Furthermore, molecular data of the sequences of these strains and P. brevispora showed a high level of similarity. These results appear to justify determining the present fungus as P. brevispora. This is the first report of this species for Japan and outside of southeastern USA. Received: March 11, 2002 / Accepted: September 20, 2002 Acknowledgments We thank Dr. Karen K. Nakasone, Center for Forest Mycology Research, Forest Products Laboratory, USDA Forest Service, for providing the fungal strains used in this study. Correspondence to:R. Kondo