Cold-adapted yeasts from Antarctica and the Italian Alps—description of three novel species: Mrakia robertii sp. nov., Mrakia blollopis sp. nov. and Mrakiella niccombsii sp. nov.

Worldwide glaciers are annually retreating due to global overheating and this phenomenon determines the potential lost of microbial diversity represented by psychrophilic microbial population sharing these peculiar habitats. In this context, yeast strains, all unable to grow above 20°C, consisting of 42 strains from Antarctic soil and 14 strains isolated from Alpine Glacier, were isolated and grouped together based on similar morphological and physiological characteristics. Sequences of the D1/D2 and ITS regions of the ribosomal DNA confirmed the previous analyses and demonstrated that the strains belong to unknown species. Three new species are proposed: Mrakia robertii sp. nov. (type strain CBS 8912), Mrakia blollopis sp. nov. (type strain CBS 8921) and a related anamorphic species Mrakiella niccombsii sp. nov. (type strain CBS 8917). Phylogenetic analysis of the ITS region revealed that the new proposed species were closely related to each other within the Mrakia clade in the order Cystofilobasidiales, class Tremellomycetes. The Mrakia clade now contains 8 sub-clades. Teliospores were observed in all strains except CBS 8918 and for the Mrakiella niccombsii strains.     

Thermoactinospora rubra gen. nov., sp. nov., a thermophilic actinomycete isolated from Tengchong, Yunnan province, south–west China

Two novel Gram-positive, spore-forming, thermophilic actinomycetes, designated as strain YIM 77501^T and YIM 77570, were isolated from a sandy soil sample collected at Tengchong National Volcanic Geological Park, Yunnan province, south–west China. Phylogenetic analysis based on the 16S rRNA gene sequences suggested that the two isolates fell within the family Streptosporangiaceae. The strains formed extensively branched substrate and aerial mycelia which carried masses of long, straight or irregular spore chains composed of warty ornamented spores. Cell walls of the two strains contained meso-diaminopimelic acid and glucose, galactose, mannose and ribose were detected as whole-cell sugars. The predominant menaquinones were MK-9(H₄) and MK-9(H₆). The major polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, N-acetylglucosamine-containing phospholipids and phosphatidylinositol, phosphatidylinositolmannosides. The major cellular fatty acids were iso-C_16:0 and 10-methyl C_17:0. The DNA G+C content was 74–76 mol%. On the basis of the morphological and chemotaxonomic characteristics as well as the phylogenetic analysis, these strains represents a novel species of a new genus within the family Streptosporangiaceae, for which the name Thermoactinospora rubra gen. nov., sp. nov. is proposed. The type strain of T. rubra is YIM 77501^T (=DSM 45614^T = CCTCC AA 2011014^T).     

Fervidobacterium islandicum sp. nov., a new extremely thermophilic eubacterium belonging to the “Thermotogales”

An extremely thermophilic anaerobic fermentative eubacterium growing at temperatures between 50 and 80°C (opt.: 65°C) was isolated from an Icelandic hot spring. The cells were Gram-negative motile rods, about 1.8 m in length, and 0.6 m in width occurring singly and in pairs. About 50% of the cells formed large spheroids at one end similar to Fervidobacterium nodosum. The new isolate H 21 differed from Fervidobacterium nodosum by a 6 mol % higher GC-content of its DNA (41 mol %), its ability to grow on cellulose, and insignificant DNA homology. The lipids of isolate H 21 were similar to that of members of Thermotogales. 16S rRNA sequencing of isolate H 21 and Fervidobacterium nodosum indicated (a) that isolate H 21 represents a new species of the genus Fervidobacterium which we name Fervidobacterium islandicum and (b) that the genus Fervidobacterium belongs to the Thermotogales branch.     

Tabrizicola aquatica gen. nov. sp. nov., a novel alphaproteobacterium isolated from Qurugöl Lake nearby Tabriz city, Iran

A novel Gram-negative, aerobic, non-motile and rod-shaped bacterium was isolated from Qurugöl Lake near Tabriz city. The bacterium grew chemoorganolheterotrophically and chemolithoautotrophically. However, photo-organoheterotrophic, photo-lithoautotrophic and fermentative growth could not be demonstrated. The presence of photosynthesis genes pufL and pufM was not shown and photosynthesis pigments were not formed. Strain RCRI19^T grew without NaCl and tolerated up to 3 % NaCl. Growth occurred at pH 6–9 (optimum, pH 7) and 15–55 °C (optimum 40–45 °C). Vitamins were not required for growth. The major fatty acids are C_18:1 ω7C, 11-methyl C_18:1 ω7C, C_18:0 3-OH. The predominant respiratory quinone is ubiquinone Q-10. The G+C content of genomic DNA is 65.9 mol%. Analysis of 16S rRNA sequences showed that strain RCRI19^T has the highest similarities with uncultured environmental sequences followed by members of the genera Rhodobacter (≤95.75 %), Haematobacter (≤95.53 %), Gemmobacter (≤95.17 %) and Falsirhodobacter (94.60 %) in the family Rhodobacteraceae. DNA–DNA relatedness between strain RCRI19^T and the closest phylogenetically related strain, Rhodobacter blasticus LMG 4305^T, was 20 %. Based on its phenotypic and chemotaxonomic characteristics and considering that it does not form photosynthetic pigments and is unable to grow phototrophically, it is concluded that strain RCRI19^T cannot be included into the genus Rhodobacter and any of the other related genera. Therefore, we propose to place the new bacterium into a new genus and species for which the name Tabrizicola aquatica gen. nov. and sp. nov. is proposed. The type strain is RCRI19^T (=BCCM/LMG 25773^T = JCM 17277^T = KCTC 23724^T).     

Corynebacterium antarcticum sp. nov., Corynebacterium marambiense sp. nov., Corynebacterium meridianum sp. nov., and Corynebacterium pygosceleis sp. nov., isolated from Adélie penguins (Pygoscelis adeliae)

A taxonomic study was conducted on 16 bacterial strains isolated from wild Adélie penguins (Pygoscelis adeliae) from Seymour (Marambio) Island and James Ross Island. An initial screening by repetitive sequence-based PCR fingerprinting divided the strains studied into four coherent groups. Phylogenetic analysis based on 16S rRNA gene sequences assigned all groups to the genus Corynebacterium and showed that Corynebacterium glyciniphilum and Corynebacterium terpenotabidum were the closest species with 16S rRNA gene sequence similarities between 95.4 % and 96.5 %. Further examination of the strains studied with ribotyping, MALDI-TOF mass spectrometry, comprehensive biotyping and calculation of average nucleotide identity and digital DNA–DNA hybridisation values confirmed the separation of the four groups from each other and from the other Corynebacterium species. Chemotaxonomically, the four strains P5828^T, P5850^T, P6136^T, P7210^T representing the studied groups were characterised by C_16:0 and C_18:1 ω9c as the major fatty acids, by the presence of meso-diaminopimelic acid in the peptidoglycan, the presence of corynemycolic acids and a quinone system with the predominant menaquinone MK-9(H₂). The results of this study show that the strains studied represent four new species of the genus Corynebacterium, for which the names Corynebacterium antarcticum sp. nov. (type strain P5850^T = CCM 8835^T = LMG 30620^T), Corynebacterium marambiense sp. nov. (type strain P5828^T = CCM 8864^T = LMG 31626^T), Corynebacterium meridianum sp. nov. (type strain P6136^T = CCM 8863^T = LMG 31628^T) and Corynebacterium pygosceleis sp. nov. (type strain P7210^T = CCM 8836^T = LMG 30621^T) are proposed.     

Echinicola shivajiensis sp. nov., a novel bacterium of the family “Cyclobacteriaceae” isolated from brackish water pond

Strain AK12^T, an orange pigmented Gram-negative, rod shaped, non-motile bacterium, was isolated from a mud sample collected from a brackish water pond at Rampur of West Bengal, India. The strain was positive for oxidase, catalase and phosphatase. The predominant fatty acids were iso-C_15:0 (42.7%), iso-C_17:0 3OH (13.2%), C_16:1ω7c/C_16:1ω6c (summed feature 3) (8.0%), iso-C_17:1 I/anteiso-C_17:1 B (summed feature 4) (6.1%) and iso-C_17:1ω9c/C_16:0 10-methyl (summed feature 9) (9.4%). Strain AK12^T contained MK-7 as the major respiratory quinone and phosphatidylethanolamine, one unidentified aminophospholipid and six unidentified lipids as the polar lipids. The G + C content of DNA of the strain AK12^T was 46.2 mol%. The 16S rRNA gene sequence analysis indicated that strain AK12^T was member of the genus Echinicola and closely related to Echinicola vietnamensis, Echinicola pacifica and Echinicola jeungdonensis with pair-wise sequence similarity of 96.8, 96.3 and 96.0% respectively. Phylogenetic analyses indicated that the strain AK12^T clustered with E. vietnamensis and together with E. pacifica and E. jeungdonensis with a phylogenetic distance of 5.1, 6.3 and 6.6% (94.9, 93.7 and 93.4% similarity) respectively. Based on data from the current polyphasic study, strain AK12^T is proposed as a novel species of the genus Echinicola, for which the name Echinicola shivajiensis sp. nov. is proposed. The type strain of E. shivajiensis is AK12^T (= MTCC 11083^T = JCM 17847^T).     

Roseinatronobacter thiooxidans gen. nov., sp. nov., a new alkaliphilic aerobic bacteriochlorophylla—containing bacterium isolated from a soda lake

Several samples of microbial mat obtained from soda lakes of the Kunkurskaya steppe (Chita region) abundantly populated by purple bacteria were screened for the presence of heterotrophic alkaliphiles capable of oxidizing sulfur compounds to sulfate. This capacity was found in only one pigmented strain, ALG 1, isolated on medium with acetate and thiosulfate at pH 10. The strain was found to be a strictly aerobic and obligately heterotrophic alkaliphile. Growth on medium with acetate was possible within a narrow pH range from 8.5 to 10.4. The strain formed a reddish orange carotenoid and bacteriochlorophylla. Pigments were synthesized only at high concentrations of nitrogen-containing organic compounds (peptone or yeast extract). The production of bacteriochlorophylla was maximal under microaerobic conditions in darkness. Strain ALG 1 could oxidize sulfide, thiosulfate, sulfite, and elemental sulfur to sulfate. In heterotrophically growing culture (pH 10), thiosulfate was not oxidized until the late logarithmic phase. The sulfur-oxidizing activity was maximal at the most alkaline pH values. The notable increase in the efficiency of organic carbon utilization observed in the presence of thiosulfate suggested that the bacterium was a sulfur-oxidizing lithoheterotroph. The phylogenetic analysis of the 16S rRNA gene showed strain ALG 1 to be a member of the α-3 subgroup of Proteobacteria and to constitute a distinct branch located between nonsulfur purple bacteriaRhodobacter andRhodovulum. Based on the unique phenotypic properties and the results of phylogenetic analysis, the alkaliphilic isolate ALG 1 was assigned to a new genus and speciesRoseinatronobacter thiooxidans with the type strain DSM-13087     

Sphingomonas gimensis sp. nov., a novel Gram-negative bacterium isolated from abandoned lead–zinc ore mine

A novel bacterial strain designated 9PNM-6^T was isolated from an abandoned lead–zinc ore mine site in Meizhou, Guangdong Province, China. The isolate was found to be Gram-negative, rod-shaped, orange-pigmented, strictly aerobic, oxidase- and catalase-positive. Growth occurred at 0–4 % NaCl (w/v, optimum, 0 %), at pH 6.0–8.0 (optimum, pH 7.0) and at 15–32 °C (optimum, 28–30 °C). Phylogenetic analysis based on 16S rRNA gene sequence similarities showed that strain 9PNM-6^T belongs to the genus Sphingomonas, with the highest sequence similarities with Sphingomonas jejuensis NBRC 107775^T (99.7 %), Sphingomonas koreensis KCTC 2882^T (95.1 %) and Sphingomonas dokdonesis KCTC 12541^T (95.1 %). The chemotaxonomic characteristics of strain 9PNM-6^T were consistent with those of the genus Sphingomonas. The predominant respiratory quinone was identified as ubiquinone Q-10, the major polyamine as sym-homospermidine, and the major cellular fatty acids as C_18:1 ω7c, C_16:0, C_16:1 ω7c and/or C_16:1 ω6c and C_14:0 2-OH. The major polar lipids are sphingoglycolipid, phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, phosphatideylcholine, an unidentified phospholipid and four unidentified aminolipids. The genomic DNA G+C content of strain 9PNM-6^T was determined to be 69.2 ± 0.6 mol%. Based on comparative analyses of morphological, physiological and chemotaxonomic data, and levels of DNA–DNA relatedness values, strain 9PNM-6^T is considered to represent a novel species of the genus Sphingomonas, for which the name Sphingomonas gimensis sp. nov. (Type strain 9PNM-6^T = GIMCC 1.655^T = CGMCC 1.12671^T = DSM 27569^T) is proposed.     

Marinifaba aquimaris gen. nov., sp. nov., a novel chitin‐degrading gammaproteobacterium in the family Alteromonadaceae isolated from seawater of the Mariana Trench

Antonie van Leeuwenhoek - A novel Gram-negative, rod-shaped, aerobic, oxidase-positive and catalase-negative bacterium, designated strain SM1970T, was isolated from a seawater sample collected from...     

Chlorolepiota indica sp. nov.—A new species from India

A new species Chlorolepiota indica is reported from Patiala in Punjab, India. The fungus is characterized by convex brown pileus covered by concentrically arranged brown scales with yellow tinge around the umbo. The stipe of the fruiting body is cream to pale yellow, pinkish to reddish brown near the pileus, and yellowish to dark grey brown towards the base.     

Geobacillus thermodenitrificans subsp. calidus, subsp. nov., a thermophilic and α-glucosidase producing bacterium isolated from Kizilcahamam, Turkey

An α-glucosidase producing, thermophilic, facultatively anaerobic, and endospore-forming, motile, rod-shaped bacterial strain F84b(T) was isolated from a high temperature well-pipeline sediment sample in Kizilcahamam, Turkey. The growth occurred at temperatures, pH and salinities ranging from 45 to 69oC (optimum 60oC), 7.0 to 8.5 (optimum 8.0) and 0 to 5% (w/v) (optimum 3.5%), respectively. Strain F84b(T) was able to grow on a wide range of carbon sources. Starch and tyrosine utilization, amylase, catalase and oxidase activities, nitrate reduction, and gas production from nitrate were all positive. The G+C content of the genomic DNA was 49.6 mol%. The menaquinone content was MK-7. The dominant cellular fatty acids were iso-C17:0, iso-C15:0, and C16:0. In phylogenetic analysis of 16S rRNA gene sequence, strain F84b(T) showed high sequence similarity to Geobacillus thermodenitrificans (99.8%) and to Geobacillus subterraneus (99.3%) with DNA hybridization values of 74.3% and 29.1%, respectively. In addition, the Rep-PCR and the intergenic 16S-23S rRNA gene fingerprinting profiles differentiated strain F84b(T) from the Geobacillus species studied. The results obtained from the physiological and biochemical characters, the menaquinone contents, the borderline DNA-DNA hybridization homology, and the genomic fingerprinting patterns had allowed phenotypic, chemotaxonomic and genotypic differentiation of strain F84b(T) from G. thermodenitrificans. Therefore, strain F84b(T) is assigned to be a new subspecies of G. thermodenitrificans, for which the name Geobacillus thermodenitrificans subsp. calidus, subsp. nov. is proposed (The type strain F84b(T) = DSM 22629(T) = NCIMB 14582(T)).     

Purification and characterization of a liquefying α-amylase from alkalophilic thermophilic Bacillus sp. AAH-31

α-Amylase (EC 3.2.1.1) hydrolyzes an internal α-1,4-glucosidic linkage of starch and related glucans. Alkalophilic liquefying enzymes from Bacillus species are utilized as additives in dishwashing and laundry detergents. In this study, we found that Bacillus sp. AAH-31, isolated from soil, produced an alkalophilic liquefying α-amylase with high thermostability. Extracellular α-amylase from Bacillus sp. AAH-31 (AmyL) was purified in seven steps. The purified enzyme showed a single band of 91 kDa on SDS-PAGE. Its specific activity of hydrolysis of 0.5% soluble starch was 16.7 U/mg. Its optimum pH and temperature were 8.5 and 70 °C respectively. It was stable in a pH range of 6.4-10.3 and below 60 °C. The calcium ion did not affect its thermostability, unlike typical α-amylases. It showed 84.9% of residual activity after incubation in the presence of 0.1% w/v of EDTA at 60 °C for 1 h. Other chelating reagents (nitrilotriacetic acid and tripolyphosphate) did not affect the activity at all. AmyL was fully stable in 1% w/v of Tween 20, Tween 80, and Triton X-100, and 0.1% w/v of SDS and commercial detergents. It showed higher activity towards amylose than towards amylopectin or glycogen. Its hydrolytic activity towards γ-cyclodextin was as high as towards short-chain amylose. Maltotriose was its minimum substrate, and maltose and maltotriose accumulated in the hydrolysis of maltooligosaccharides longer than maltotriose and soluble starch.     

Anditalea andensis gen. nov., sp. nov., an alkaliphilic, halotolerant bacterium isolated from extreme alkali–saline soil

A novel alkalophilic salt-tolerant rod-shaped bacterium, designated ANESC-S^T, was isolated from an extremely alkali–saline soil in the rural area of Anda city in northeast China. Taxonomic study using a polyphasic approach revealed that this non-motile, orange colony-forming microbe was Gram-negative and obligately aerobic. Optimal growth of strain ANESC-S^T was achieved in the presence of NaCl with a concentration range of 0.5 to 4 % and pH between 7.5 and 9.2, and at temperatures ranging from 10 to 37 °C. Phylogenetic analysis of 16S rRNA sequences showed that of strain ANESC-S^T is most homologous to Mongoliicoccus roseus MIM28^T and Litoribacter ruber YIM CH208^T with sequence similarity of 95.1 and 93.2 %, respectively. The genomic DNA G+C content of strain ANESC-S^T was determined to be 39.1 mol%. The main isoprenoid quinone in ANESC-S^T was found to be menaquinone-7. The main fatty acids were found to be iso-C_15:0 (27.5 %), iso-C_17:03-OH (14.0 %), anteiso-C_15:0 (9.8 %), summed feature 9 (iso-C_17:1ω9c and/or 10-methyl C_16:0 10.6 %) and summed feature 3 (C_16:1w7c/C_16:1w6c, 9.78 %). Based on the phenotypic, chemotaxonomic and phylogenetic data, strain ANESC-S^T is considered to represent a new genus and species classified into the order Cytophagales, for which the name Anditalea andensis gen. nov., sp. nov. is proposed. The type strain is ANESC-S^T (=CICC 10485^T = NCCB 100412^T).     

Molecular cloning and characterization of a novel γ-CGTase from alkalophilic Bacillus sp.

We found a novel cyclodextrin glucanotransferase (CGTase) from alkalophilic Bacillus sp. G-825-6. The enzyme was expressed in the culture broth by recombinant Bacillus subtilis KN2 and was purified and characterized. The enzyme named CGTase825-6 showed 95% amino acid sequence identity with a known enzyme β-/γ-CGTase from Bacillus firmus/lentus 290-3. However, the product specificity of CGTase825-6 differed from that of β-/γ-CGTase. CGTase825-6 produced γ-cyclodextrin (CD) as the main product, but degradation of γ-CD was observed with prolonged reaction. The product specificity of the enzyme was positioned between γ-CGTase produced by Bacillus clarkii 7364 and B. firmus/lentus 290-3 β-/γ-CGTase. It showed that the difference of product specificity was dependent on only 28 amino acid residues in 671 residues in CGTase825-6. We compared the amino acid sequence of CGTase825-6 and those of other CGTases and constructed a protein structure model of CGTase825-6. The comparison suggested that the diminished loop (Val138-Asp142) should provide subsite -8 for γ-CD production and that Asp142 might have an important role in product specificity. CGTase825-6 should be a useful tool to produce γ-CD and to study the differences of producing mechanisms between γ-CD and β-CD.     

Rapid and simple purification of a novel extracellular β-amylase from Bacillus sp.

Bacillus sp. KYJ 963, a local isolate, produced an extracellular amylase with M _r=59 kDa. The amylase was easily purified by adsorption on soluble starch. The analyses of TLC and N-terminal amino acid sequence from the purified protein revealed that the enzyme was a novel -amylase which could not hydrolyze maltose or -cyclodextrin and its N-terminal amino acid sequence was A-V-N-G-Q-S-F-N-S-N-Y-K-T-Y-K-.     

Natranaerofaba carboxydovora gen. nov., sp. nov., an extremely haloalkaliphilic CO-utilizing acetogen from a hypersaline soda lake representing a novel deep phylogenetic lineage in the class ‘Natranaerobiia’

An anaerobic enrichment with CO from sediments of hypersaline soda lakes resulted in a methane-forming binary culture, whereby CO was utilized by a bacterium and not the methanogenic partner. The bacterial isolate ANCO1 forms a deep-branching phylogenetic lineage at the level of a new family within the class ‘Natranaerobiia’. It is an extreme haloalkaliphilic and moderate thermophilic acetogen utilizing CO, formate, pyruvate and lactate as electron donors and thiosulfate, nitrate (reduced to ammonia) and fumarate as electron acceptors. The genome of ANCO1 encodes a full Wood–Ljungdahl pathway allowing for CO oxidation and acetogenic conversion of pyruvate. A locus encoding Nap nitrate reductase/NrfA ammonifying nitrite reductase is also present. Thiosulfate respiration is encoded by a Phs/Psr-like operon. The organism obviously relies on Na-based bioenergetics, since the genome encodes for the Na⁺-Rnf complex, Na⁺-F1F0 ATPase and Na⁺-translocating decarboxylase. Glycine betaine serves as a compatible solute. ANCO1 has an unusual membrane polar lipid composition dominated by diethers, more common among archaea, probably a result of adaptation to multiple extremophilic conditions. Overall, ANCO1 represents a unique example of a triple extremophilic CO-oxidizing anaerobe and is classified as a novel genus and species Natranaerofaba carboxydovora in a novel family Natranaerofabacea.