Streptomyces pacificus sp. nov., a novel spongiicolazolicin-producing actinomycete isolated from a coastal sediment.

A polyphasic approach was used to determine the taxonomic position of a marine actinomycete, designated isolate CWH03T, which we previously reported to produce new linear azole-containing peptides spongiicolazolicins A and B. Strain CWH03T is mesophilic, neutrophilic, and halotolerant streptomycete that forms spiral spore chains on aerial mycelium. Comparative 16S rRNA gene sequencing showed that CWH03T was most closely related to Streptomyces tirandamycinicus HNM0039T (99.7%), Streptomyces spongiicola HNM0071T (99.4%), 'Streptomyces marianii' ICN19T (99.1%) and Streptomyces wuyuanensis CGMCC4.7042T (99.0%). The phylogenetic tree prepared using the 16S rRNA gene, as well as the phylogenomic tree using the genome BLAST distance phylogeny method and 81 core housekeeping genes, respectively, showed that the closest relative of strain CWH03T was S. spongiicola HNM0071T. The average nucleotide identity and digital DNA-DNA hybridization values between strains CWH03T and S. spongiicola HNM0071T were 91.46% and 44.2%, respectively, which were below the thresholds of 96% and 70% for prokaryotic conspecific assignation. The G+C content of the genomic DNA of strain CWH03T was 72.3%. Whole-cell hydrolysates of strain CWH03T contained LL-diaminopimelic acid. The predominant menaquinone was MK-9(H8) (88.3%), and the major fatty acids were iso-C16:0 (28.4%), anteiso-C15:0 (15.0%) and iso-C15:0 (12.9%). The major phospholipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and an unidentified phospholipid. Based on data obtained from phenotypic, phylogenetic, genomic, and chemotaxonomic analyses, strain CWH03T represents a novel species of the genus Streptomyces, for which the proposed name is Streptomyces pacificus sp. nov. The type strain is CWH03T ( = NBRC 114659T = TBRC 15780T).

Plant growth-promoting and antimicrobial chloropyrroles from a rare actinomycete of the genus Catellatospora.

Two new chloropyrroles, designated catellatopyrroles A (1) and B (2), along with 2-(2'-hydroxybenzoyl)pyrrole (3), were isolated from a culture extract of an actinomycete of the genus Catellatospora. The structures of 1-3 were elucidated through interpretation of NMR and MS data. Compounds 1 and 2 are the first chloropyrroles substituted by an aliphatic acyl group at the 5-position. Compounds 1-3 promoted root elongation of germinated lettuce seeds at 1-10 µM. While all compounds inhibited the growth of Gram-positive bacteria, activity against Gram-negative bacterium Rhizobium radiobacter and yeasts Candida albicans and Saccharomyces cerevisiae was varied. Compounds 1 and 2 were moderately cytotoxic against P388 cells.

Characterization of the Non-rhizobial Bacterial Community in the Nodule-Associated Soils of Akebono Soybeans and Isolation of Antibiotic Producing Amycolatopsis spp.

The unique Akebono soybeans are cultivated in Minobu Town, Yamanashi Prefecture, Japan. The biogeography of Akebono soybeans and the microbial diversity associated with their root nodules remain unexplored. This study investigated the nodule-associated microbial community of Akebono soybeans using molecular techniques. The results showed that the family Bradyrhizobiaceae was dominant in soybeans obtained from Minobu town Imata (MI), Minobu town IItomi (MS), and Minobu town Hirase (MN). In contrast, members of the family Pseudonocardiaceae were isolated from the nodule samples from Minobu town Yasaiku (MY). The community structure of MY was different from that of the other sites and the genus Amycolatopsis was dominant. The bacterial community in the nodule associated soil obtained from the Minobu area was different from that of soybeans cultivated in Kofu City (approximately 35 km away from Minobu Town). The MY1 strain of Amycolatopsis spp. was isolated from the nodule associated soil in MY and was found to produce antibiotics. This study showed that the bacterial community in the nodules and the adjacent rhizosphere may be a regional characteristic of Akebono soybeans and the specific Amycolatopsis spp. dominant in the nodule associated soil of MY was implicated in determining the bacterial community structure. Supplementary Information: The online version contains supplementary material available at 10.1007/s12088-022-00999-x.

Isolation and structure determination of new linear azole-containing peptides spongiicolazolicins A and B from Streptomyces sp. CWH03.

Linear azole-containing peptides are a class of ribosomally synthesized and post-translationally modified peptides. We performed a chemical investigation on marine actinomycetes, and new linear azole-containing peptides named spongiicolazolicins A and B were found in the MeOH extracts of a newly isolated strain Streptomyces sp. CWH03 (NBRC 114659) and two strains of S. spongiicola (strain HNM0071T: DSM 103383T and strain 531S: NBRC 113560). The strain Streptomyces sp. CWH03 was indicated to be a new species closely related to S. spongiicola by phylogenetic analysis using the genome sequence. The new peptides named spongiicolazolicins A and B were isolated from the cell of Streptomyces sp. CWH03. The partial structure of spongiicolazolicin A was determined by 2D NMR experiments. Based on data of MS/MS experiments, the chemical structures of spongiicolazolicins A and B were proposed using the amino acid sequence deduced from the precursor-encoding gene, which was found from whole-genome sequence data of Streptomyces sp. CWH03. The biosynthetic gene cluster of spongiicolazolicins was proposed based on comparative analysis with that of a known linear azole peptide goadsporin. KEY POINTS: • Streptomyces sp. CWH03 was a new species isolated from marine sediment. • New linear azole-containing peptides named spongiicolazolicins A and B were isolated. • Biosynthetic pathway of spongiicolazolicins was proposed.

Probiotic Bifidobacterium bifidum G9-1 ameliorates phytohemagglutinin-induced diarrhea caused by intestinal dysbiosis.

Diarrhea is largely caused by dysbiosis accompanying the hyperproliferation of Escherichia coli (E. coli). While current treatments can resolve the symptoms, they cannot suppress the proliferation of pathogenic bacteria in the intestine. Probiotics have numerous beneficial effects on host health, including restoring the balance of the intestinal microbiota. This study investigated the effect of the probiotic Bifidobacterium bifidum G9-1 (BBG9-1), which is active in intestinal dysbiosis, in the incidence of diarrhea, in the composition of the intestinal microbiota, and in the intestinal tissue of a rat model of phytohemagglutinin (PHA)-induced diarrhea. The rats were treated with PHA, with and without BBG9-1, and the microbiota composition throughout the intestine and stool was examined using high-throughput 16S rRNA sequencing. In line with previous reports, PHA administration caused diarrhea as well as dysbiosis due to E. coli hyperproliferation. Histological findings indicated that the jejunal villus length was shortened. Rats that received BBG9-1 showed clear improvements in dysbiosis, diarrhea symptoms, and jejunal villus length. Principal coordinates analysis demonstrated the microbiota profile to be more similar between the BBG9-1 and normal groups than between the PHA and normal groups. These results indicated that BBG9-1 suppresses the hyperproliferation of E. coli and restores the jejunal villus length, thereby improving dysbiosis, and in turn, alleviating the symptoms of diarrhea.

Cellulomonas algicola sp. nov., an actinobacterium isolated from a freshwater alga.

An actinomycete strain, TKZ-21T, was isolated from a freshwater alga (Chetophoraceae) collected from the Takizawa River, Yamanashi, Japan, and examined using a polyphasic taxonomic approach. Cells were Gram-stain positive, aerobic, non-sporulating, motile, and coccoid or short rod-shaped. The strain grew in the presence of 0-4 % (w/v) NaCl, between pH 6-9.4, and over a temperature range of 15-40 °C, with optimum growth at 30 °C. The peptidoglycan type of strain TKZ-21T was A4ß, containing l-ornithine as diagnostic diamino acid and d-glutamic acid as the interpeptide bridge. The predominant menaquinone was MK-9(H4). The polar lipids were diphosphatidylglycerol, phosphatidylglycerol, ninhydrin-positive glycolipid, and unidentified phospholipids. The major cellular fatty acids were anteiso-C15 : 0 and anteiso-C17 : 0, and the DNA G+C content was 75.6 mol%. On the basis of 16S rRNA gene sequence analysis, strain TKZ-21T was closely related to Cellulomonas fimi (98.5 % sequence similarity) and Cellulomonas biazotea (98.3 %). The genome orthoANI value between strain TKZ-21T and C. biazotea and C. fimi were 84.7 and 84.2 %, respectively. On the basis of fatty acid and MALDI-TOF MS profile analysis, phylogenetic analyses, genomic analysis, and phenotypic data, it is proposed that the isolate be classified as a representative of a novel species of the genus Cellulomonas, with the name Cellulomonas algicola sp. nov. The type strain is TKZ-21T (=NBRC 112905T=TBRC 8129T).

Alleviation of low-fiber diet-induced constipation by probiotic Bifidobacterium bifidum G9-1 is based on correction of gut microbiota dysbiosis.

Constipation, a functional disorder of the digestive system, is common in children and adults and may compromise patient quality of life. Because many patients are not satisfied with the efficacy of existing therapies, in this study, we investigated the efficacy of the probiotic Bifidobacterium bifidum G9-1 (BBG9-1) in constipation induced by a low-fiber diet. After inducing constipation in rats by feeding a low-fiber diet, rats were fed a low-fiber diet mixed with BBG9-1 in 14 days to determine the efficacy of BBG9-1 for alleviating constipation. BBG9-1 significantly alleviated the dysbiosis induced by a low-fiber diet and improved the fecal counts, fecal weights, and fecal water contents. Moreover, it also improved organic acid concentrations in the cecal contents. These results suggested that in low-fiber diet-induced constipation, BBG9-1 could alleviate dysbiosis and constipation and may improve the intestinal environment, supporting its potential application in the treatment of constipation.

Virgisporangium myanmarense sp. nov., a novel motile actinomycete isolated from an anthill soil in Myanmar.

An actinomycete strain, designated MM04-1133T, was isolated from an anthill soil sample collected in Bagan, Myanmar. To establish the taxonomic status of this strain, the isolate was subjected to a polyphasic approach. Strain MM04-1133T was Gram-staining positive, aerobic, motile and formed long and narrow sporangia directly above the surface of the substrate mycelium. Whole-cell hydrolysates of the strain contained 3-OH-diaminopimelic acid, arabinose, glucose, galactose, mannose, rhamnose and xylose. The predominant menaquinones were MK-10(H6) and MK-10(H8). The major cellular fatty acids were iso-C16:0 and anteiso-C17:0. The diagnostic phospholipid was phosphatidylethanolamine. The G+C content of the DNA was 69.1 mol%. Phylogenetic analysis based on 16S rRNA gene sequence revealed that strain MM04-1133T clustered within the genus Virgisporangium, with the sequence exhibiting highest similarity (98.5% identity) with Virgisporangium ochraceum NBRC 16418T. The strain grew in the presence of 0-1% (w/v) NaCl, at pH 5-8 and at 20-40 °C, with optimal growth at 30-37 °C. Based on phylogenetic analysis and chemotaxonomic and phenotypic data, we propose classifying this isolate as a novel species of the genus Virgisporangium, to be designated as Virgisporangium myanmarense sp. nov. The type strain is MM04-1133T (=NBRC 112733T=VTCC 910008T).

Development of intra-strain self-cloning procedure for breeding baker's yeast strains.

Previously reported self-cloning procedures for breeding of industrial yeast strains require DNA from other strains, plasmid DNA, or mutagenesis. Therefore, we aimed to construct a self-cloning baker's yeast strain that exhibits freeze tolerance via an improved self-cloning procedure. We first disrupted the URA3 gene of a prototrophic baker's yeast strain without the use of any marker gene, resulting in a Δura3 homozygous disruptant. Then, the URA3 gene of the parental baker's yeast strain was used as a selection marker to introduce the constitutive TDH3 promoter upstream of the PDE2 gene encoding high-affinity cyclic AMP phosphodiesterase. This self-cloning procedure was performed without using DNA from other Saccharomyces cerevisiae strains, plasmid DNA, or mutagenesis and was therefore designated an intra-strain self-cloning procedure. Using this self-cloning procedure, we succeeded in producing self-cloning baker's yeast strains that harbor the TDH3p-PDE2 gene heterozygously and homozygously, designated TDH3p-PDE2 hetero and TDH3p-PDE2 homo strains, respectively. These self-cloning strains expressed much higher levels of PDE2 mRNA than the parental strain and exhibited higher viability after freeze stress, as well as higher fermentation ability in frozen dough, when compared with the parental strain. The TDH3p-PDE2 homo strain was genetically more stable than the TDH3p-PDE2 hetero strain. These results indicate that both heterozygous and homozygous strains of self-cloning PDE2-overexpressing freeze-tolerant strains of industrial baker's yeast can be prepared using the intra-strain self-cloning procedure, and, from a practical viewpoint, the TDH3p-PDE2 homo strain constructed in this study is preferable to the TDH3p-PDE2 hetero strain for frozen dough baking.

Longitudinal study of effects of oral dosage of Bifidobacterium bifidum G9-1 on Japanese cedar pollen-induced allergic nasal symptoms in guinea pigs.

Previous studies using experimental animal models have reported the beneficial effects of probiotics on allergic responses; however, their long-term effects on allergic nasal symptoms in clinical settings have not yet been elucidated in detail. In the present study, a guinea pig allergic rhinitis model involving repeated inhalation challenges with a natural allergen, Japanese cedar pollen, was used to examine the longitudinal effects of Bifidobacterium bifidum G9-1 (BBG9-1) on allergic nasal symptoms. BBG9-1 was administered orally once a day. Amelioration of nasal blockage was consistently observed throughout the experimental period in the BBG9-1-treated group. Although challenge-induced sneezing was not significantly inhibited in the BBG9-1-treated group, prolonged treatment with BBG9-1 slightly reduced the frequency of sneezing. Antigen-specific IgE antibody production was also not inhibited in the BBG9-1-treated group. Increases in the numbers of eosinophils and neutrophils in nasal cavity lavage fluid collected after pollen challenge were almost completely suppressed by BBG9-1 treatment, whereas those in mast cell mediators, histamine and cysteinyl leukotrienes were not. In contrast, increases in the levels of nitric oxide metabolites were potently suppressed. Furthermore, prolonged BBG9-1 treatment markedly suppressed exogenous leukotriene D4 -induced nasal blockage. Thus, prolonged oral administration of BBG9-1 suppresses Japanese cedar pollen-induced allergic nasal symptoms. The inhibitory mechanisms responsible may involve reductions in the responsiveness of target organs, such as endothelial cells in nasal mucosal blood vessels, to chemical mediators.

Proposal of nine novel species of the genus Lysinimicrobium and emended description of the genus Lysinimicrobium.

Thirteen novel Gram-stain-positive bacteria were isolated from various samples collected from mangrove forests in Japan, and their taxonomic positions were investigated by a polyphasic approach. Phylogenetic analyses based on 16S rRNA gene sequence comparisons showed that the 13 isolates formed a single clade with Lysinimicrobium mangrovi HI08-69T, with a similarity range of 97.6-99.5 %. The peptidoglycan of the isolates was of the A4α type with an interpeptide bridge comprising Ser-Glu and an l-Ser residue at position 1 of the peptide subunit. The predominant menaquinone was demethylmenaquinone DMK-9(H4) and the major fatty acid was anteiso-C15 : 0. These chemotaxonomic characteristics corresponded to those of the genus Lysinimicrobium. On the basis of the phenotypic and phylogenetic data, along with average nucleotide identity values among the isolates, we concluded that the 13 isolates should be assigned to the following nine novel species of the genus Lysinimicrobium: Lysinimicrobium aestuarii sp. nov. (type strain HI12-104T = NBRC 109392T = DSM 28144T), Lysinimicrobium flavum sp. nov. (type strain HI12-45T = NBRC 109391T = DSM 28150T), Lysinimicrobium gelatinilyticum sp. nov. (type strain HI12-44T = NBRC 109390T = DSM 28149T), Lysinimicrobium iriomotense sp. nov. (type strain HI12-143T = NBRC 109399T = DSM 28146T), Lysinimicrobium luteum sp. nov. (type strain HI12-123T = NBRC 109395T = DSM 28147T), Lysinimicrobium pelophilum sp. nov. (type strain HI12-111T = NBRC 109393T = DSM 28148T), Lysinimicrobium rhizosphaerae sp. nov. (type strain HI12-135T = NBRC 109397T = DSM 28152T), Lysinimicrobium soli sp. nov. (type strain HI12-122T = NBRC 109394T = DSM 28151T) and Lysinimicrobium subtropicum sp. nov. (type strain HI12-128T = NBRC 109396T = DSM 28145T). In addition, an emended description of the genus Lysinimicrobium is proposed.

Total Synthesis and Structure Determination of JBIR-108-A 2-Hydroxy-2-(1-hydroxyethyl)-2,3-dihydro-3(2H)-furanone Isolated from Streptomyces gramineus IR087Pi-4.

The planar and stereostructures of JBIR-108 isolated from Streptomyces gramineus IR087Pi-4 were determined partly by spectral analysis, and these structural assignments were confirmed and completed by the total synthesis of both 1-epimers. The key stereocenters in JBIR-108 were constructed via a Corey-Bakshi-Shibata (CBS) reduction (C-1), vinylogous Mukaiyama aldol reaction (C-7), and Brown crotylation (C-14 and C-15). Although it was difficult to determine the stereochemistries at the C-1 and C-7 positions in the natural product using the modified Mosher's method, the synthesis of two possible C-1 diastereomers enabled the identification of the configurations at the hitherto unknown stereocenters.

Isolation of a novel plasmid from Couchioplanes caeruleus and construction of two plasmid vectors for gene expression in Actinoplanes missouriensis.

To date, no plasmid vector has been developed for the rare actinomycete Actinoplanes missouriensis. Moreover, no small circular plasmid has been reported to exist in the genus Actinoplanes. Here, a novel plasmid, designated pCAZ1, was isolated from Couchioplanes caeruleus subsp. azureus via screening for small circular plasmids in Actinoplanes (57 strains) and Couchioplanes (2 strains). Nucleotide sequencing revealed that pCAZ1 is a 5845-bp circular molecule with a G + C content of 67.5%. The pCAZ1 copy number was estimated at 30 per chromosome. pCAZ1 contains seven putative open reading frames, one of which encodes a protein containing three motifs conserved among plasmid-encoded replication proteins that are involved in the rolling-circle mechanism of replication. Detection of single-stranded DNA intermediates in C. caeruleus confirmed that pCAZ1 replicates by this mechanism. The ColE1 origin from pBluescript SK(+) and the oriT sequence with the apramycin resistance gene aac(3)IV from pIJ773 were inserted together into pCAZ1, to construct the Escherichia coli-A. missouriensis shuttle vectors, pCAM1 and pCAM2, in which the foreign DNA fragment was inserted into pCAZ1 in opposite directions. pCAM1 and pCAM2 were successfully transferred to A. missouriensis through the E. coli-mediated conjugative transfer system. The copy numbers of pCAM1 and pCAM2 in A. missouriensis were estimated to be one and four per chromosome, respectively. Thus, these vectors can be used as effective genetic tools for homologous and heterologous gene expression studies in A. missouriensis.

Forced expression of FLO11 confers pellicle-forming ability and furfural tolerance on Saccharomyces cerevisiae in ethanol production.

We constructed a plasmid that expresses FLO11 encoding a cell surface glycoprotein of Saccharomyces cerevisiae under the control of a constitutive promoter. This plasmid conferred pellicle-forming ability on the non-pellicle-forming industrial strain of S. cerevisiae at the air-liquid interface of the glucose-containing liquid medium. The induced pellicle-forming cells exhibited tolerance to furfural, which is a key toxin in lignocellulosic hydrolysates, in ethanol production.

Streptomyces hokutonensis sp. nov., a novel actinomycete isolated from the strawberry root rhizosphere.

A polyphasic approach was used to determine the taxonomic position of actinomycete strain R1-NS-10(T), which was isolated from a sample of strawberry root rhizosphere obtained from Hokuto, Yamanashi, Japan. Strain R1-NS-10(T) was Gram-staining-positive and aerobic, and formed brownish-white aerial mycelia and grayish-brown substrate mycelia on ISP-2 medium. The strain grew in the presence of 0-5% (w/v) NaCl and optimally grew without NaCl. The strain grew at pH 5-8, and the optimum for growth was pH 7. The optimal growth temperature was 30 °C, but the strain grew at 5-37 °C. Whole-cell hydrolysates of strain R1-NS-10(T) contained A2pm, galactose, mannose and rhamnose. The predominant menaquinones were MK-9(H6) and MK-9(H8). The major cellular fatty acids were anteiso-C15:0 and iso-C16:0. Comparative 16S rRNA gene sequence analysis revealed that strain R1-NS-10(T) was most closely related to Streptomyces prunicolor NBRC 13075(T) (99.4%). The draft genome sequences of both strains were determined for characterization of genome sequence-related parameters such as average nucleotide identity (ANI) and the diversity of secondary metabolite biosynthetic gene clusters. DNA-DNA hybridization (DDH) and ANI values for both strains were below the species delineation cutoff, and differences in physiological and biochemical characteristics differentiated strain R1-NS-10(T) from its closest phylogenetic relative. On the basis of these data, we propose that strain R1-NS-10(T) (=NBRC 108812(T)=KCTC 29186(T)) should be classified as the type strain of a novel Streptomyces species named Streptomyces hokutonensis sp. nov.

Agromyces iriomotensis sp. nov. and Agromyces subtropicus sp. nov., isolated from soil.

Three novel Gram-stain-positive bacteria, designated IY07-20(T), IY07-56(T) and IY07-113, were isolated from soil samples from Iriomote Island, Okinawa, Japan, and their taxonomic positions were investigated by a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequence comparisons showed that the three isolates were closely related to the members of the genus Agromyces, with similarity range of 95.6-98.7%. The isolates contained l-2,4-diaminobutylic acid, d-alanine, d-glutamic acid and glycine in their peptidoglycans. The predominant menaquinone was MK-12 and the major fatty acids were anteiso-C15:0 and anteiso-C17:0. The DNA G+C contents were 70.9-72.9 mol%. The chemotaxonomic characteristics of the isolates matched those described for members of the genus Agromyces. The results of phylogenetic analysis and DNA-DNA hybridization, along with differences in phenotypic characteristics between strains IY07-20(T), IY07-56(T) and IY07-113 and the species of the genus Agromyces with validly published names, indicate that the three isolates merit classification as representatives of two novel species of the genus Agromyces, for which the names Agromyces iriomotensis sp. nov. and Agromyces subtropica sp. nov. are proposed; the type strains are IY07-20(T) ( = NBRC 106452(T) = DSM 26155(T)) and IY07-56(T) ( = NBRC 106454(T) = DSM 26153(T)), respectively.

Janibacter cremeus sp. nov., an actinobacterium isolated from sea sediment.

A novel Gram-stain-positive actinobacterium, designated HR08-44(T), was isolated from a sea-sediment sample collected from the foreshore of Rishiri Island, Japan, and its taxonomic position was investigated by a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strain HR08-44(T) was closely related to the members of the genus Janibacter, with pairwise sequence similarities of 97.3-98.8 %. Strain HR08-44(T) had peptidoglycan type A1γ, with meso-diaminopimelic acid as the diagnostic diamino acid. The predominant menaquinone was MK-8(H4) and the major fatty acids were iso-C16 : 0, C17 : 1ω8c, C17 : 0 and C18 : 1ω9c. These data supported the affiliation of the novel strain to the genus Janibacter. Meanwhile, the results of DNA-DNA hybridization and physiological and biochemical tests indicated that strain HR08-44(T) can be distinguished from recognized species of the genus Janibacter. Therefore, strain HR08-44(T) represents a novel species of the genus Janibacter, for which the name Janibacter cremeus sp. nov. is proposed; the type strain is HR08-44(T) ( = NBRC 107693(T) = DSM 26154(T)).

A new simple method for isolating multistress-tolerant semidominant mutants of Saccharomyces cerevisiae by one-step selection under lethal hydrogen peroxide stress condition.

Tolerance of microorganisms to diverse stresses (i.e., multistress tolerance) is a very useful property with industrial applications. We have developed a simple method for isolating multistress-tolerant semidominant mutants of the budding yeast Saccharomyces cerevisiae by one-step selection under lethal hydrogen peroxide (H(2)O(2)) stress condition, which we named the lethal concentration of H(2)O(2) (LCH) method. This method involves simply isolating colonies after plating of mutagenized S. cerevisiae cells, which are cultivated overnight in liquid media, on agar plates containing a lethal concentration of H(2)O(2) for the wild-type strain. Phenotypic and genetic analyses of the ten strains isolated by this method revealed that two strains exhibiting stress tolerance to H(2)O(2), ethanol, heat shock, salt, organic solvent, freeze-thaw, chronological aging, and high concentrations of glucose possess semidominant and distinct single-gene mutations designated as MLT1-1 (multistress tolerance) and MLT2-1, which are responsible for multistress tolerance. From these results, we expect this method to confer multistress tolerance on industrial yeasts.

Paraoerskovia sediminicola sp. nov., an actinobacterium isolated from sea sediment, and emended description of the genus Paraoerskovia.

A novel Gram-stain-positive actinobacterium, designated H25-14(T), was isolated from a sea sediment sample, and its taxonomic position was investigated by a polyphasic approach. The peptidoglycan type of strain H25-14(T) was A4α and lysine was the diagnostic diamino acid of the peptidoglycan. The predominant menaquinone was MK-9(H4) and the major fatty acids were anteiso-C15 : 0 and iso-C15 : 0. The DNA G+C content was 73.2 mol%. Phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strain H25-14(T) was closely related to Paraoerskovia marina NBRC 104352(T) (98.3 %). However, DNA-DNA hybridization data and phenotypic characteristics revealed that strain H25-14(T) differed from P. marina NBRC 104352(T). Therefore, strain H25-14(T) represents a novel species of the genus Paraoerskovia, for which the name Paraoerskovia sediminicola sp. nov. is proposed. The type strain is H25-14(T) ( = NBRC 108565(T) = DSM 25477(T)). An emended description of the genus Paraoerskovia is also proposed.

Demequina flava sp. nov. and Demequina sediminicola sp. nov., isolated from sea sediment.

Two novel Gram-stain-positive bacteria, designated HR08-7(T) and HR08-43(T), were isolated from a sea sediment sample from Rishiri Island, Hokkaido, Japan, and their taxonomic positions were investigated by a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequence comparisons revealed that strains HR08-7(T) and HR08-43(T) and the members of the genus Demequina formed a monophyletic cluster with similarity range of 95.5-99.0 %. The peptidoglycan type of strains HR08-7(T) and HR08-43(T) was A4ß. The predominant menaquinone of both strains was demethylmenaquinone DMK-9(H(4)) and the major fatty acid was anteiso-C(15 : 0). The DNA G+C contents of strains HR08-7(T) and HR08-43(T) were 64.5 and 62.4 mol%, respectively. The results of phylogenetic analysis and DNA-DNA hybridization, along with differences of strains HR08-7(T) and HR08-43(T) from the recognized Demequina species in phenotypic characteristics, indicate that the two strains merit classification as representatives of two novel species of the genus Demequina, for which the names Demequina flava sp. nov. and Demequina sediminicola sp. nov. are proposed; the type strains are HR08-7(T) (= NBRC 105854(T) = DSM 24865(T)) and HR08-43(T) (= NBRC 105855(T) = DSM 24867(T)), respectively.