Isolation and Characterization of Acetate-Utilizing Anaerobes from a Freshwater Sediment

Acetate-degrading anaerobic microorganisms in freshwater sediment were quantified by the most probable number technique. From the highest dilutions a methanogenic, a sulfate-reducing, and a nitrate-reducing microorganism were isolated with acetate as substrate. The methanogen (culture AMPB-Zg) was non-motile and rod-shaped with blunted ends (0.5–1 μm × 3–4 μm long). Doubling times with acetate at 30–35°C were 5.6–8.1 days. The methanogen grew only on acetate. Analysis of the 16S rRNA sequence showed that AMPB-Zg is closely related toMethanosaeta concilii. The isolated sulfate-reducing bacterium (strain ASRB-Zg) was rod-shaped with pointed ends (0.5–0.7 μm × 1.5–3.5 μm long), weakly motile, spore forming, and gram positive. At the optimum growth temperature of 30°C the doubling times with acetate were 3.9–5.3 days. The bacterium grew on a range of organic acids, such as acetate, butyrate, fumarate, and benzoate, but did not grow autotrophically with H₂, CO₂, and sulfate. The closest relative of strain ASRB-Zg isDesulfotomaculum acetoxidans. The nitrate-reducing bacterium (strain ANRB-Zg) was rod-shaped (0.5–0.7 μm × 0.7–1 μm long), weakly motile, and gram negative. Optimum growth with acetate occurred at 20–25°C. The bacterium grew on a range of organic substrates, such as acetate, butyrate, lactate, and glucose, and did grow autotrophically with H₂, CO₂, and oxygen but not with nitrate. In the presence of acetate and nitrate, thiosulfate was oxidized to sulfate. Phylogenetically, the closest relative of strain ANRB-Zg isVariovorax paradoxus.     

Purification and characterization of thermostable pectate-lyases from a newly isolated thermophilic bacterium, Thermoanaerobacter italicus sp. nov.

A novel thermophilic spore-forming anaerobic microorganism (strain Ab9) able to grow on citrus pectin and polygalacturonic acid (pectate) was isolated from a thermal spa in Italy. The newly isolated strain grows optimally at 70°C with a growth rate of 0.23 h⁻¹ with pectin and 0.12 h⁻¹ with pectate as substrates. Xylan, starch, and glycogen are also utilized as carbon sources and thermoactive xylanolytic (highest activity at 70°–75°C), amylolytic as well as pullulolytic enzymes (highest activity at 80°–85°C) are formed. Two thermoactive pectate lyases were isolated from the supernatant of a 300-l culture of isolate Ab9 after growth on citrus pectin. The two enzymes (lyases a and b) were purified to homogeneity by ammonium sulfate treatment, anion exchange chromatography, hydrophobic chromatography and finally by preparative gel electrophoresis. After sodium dodecylsulfate (SDS) gel electrophoresis, lyase a appeared as a single polypeptide with a molecular mass of 135 000 Da whereas lyase b consisted of two subunits with molecular masses of 93 000 Da and 158 000 Da. Both enzymes displayed similar catalytic properties with optimal activity at pH 9.0 and 80°C. The enzymes were very stable at 70°C and at 80°C with a half-life of more than 60 min. The maximal activity of the purified lyases was observed with orange pectate (100%) and pectate-sodium salt (90%), whereas pectin was attacked to a much lesser extent (50%). The K _m values of both lyases for pectate and citrus pectin were 0.5 g·l⁻¹ and 5.0 g·l⁻¹, respectively. After incubation with polygalacturonic acid, mono-, di-, and tri-galacturonate were detected as final products. A 2.5-fold increase of activity was obtained when pectate lyases were incubated in the presence of 1 mM Ca²⁺. The addition of 1 mM ethylenediaminetetraacetic acid (EDTA) resulted in complete inhibition of the enzymes. These heat-stable enzymes represent the first pectate-lyases isolated and characterized from a thermophilic anaerobic bacterium. On the basis of the results of the 16S rRNA sequence comparisons and the observed phenotypic differences, we propose strain Ab9 as a new species of Thermoanaerobacter, namely Thermoanaerobacter italicus sp. nov. Received: May 25, 1997 / Accepted: June 5, 1997     

Obligately and facultatively autotrophic,sulfur- and hydrogen-oxidizing thermophilic bacteria isolated from hot composts

A variety of autotrophic, sulfur- and hydrogen-oxidizing thermophilic bacteria were isolated from thermogenic composts at temperatures of 60–80° C. All were penicillin G sensitive, which proves that they belong to the Bacteria domain. The obligately autotrophic, non-spore-forming strains were gram-negative rods growing at 60–80°C, with an optimum at 70–75°C, but only under microaerophilic conditions (5 kPa oxygen). These strains had similar DNA G+C content (34.7–37.6 mol%) and showed a high DNA:DNA homology (70–87%) with Hydrogenobacter strains isolated from geothermal areas. The facultatively autotrophic strains isolated from hot composts were gram-variable rods that formed spherical and terminal endospores, except for one strain. The strains grew at 55–75° C, with an optimum at 65–70° C. These bacteria were able to grow heterotrophically, or autotrophically with hydrogen; however, they oxidized thiosulfate under mixotrophic growth conditions (e.g. pyruvate or hydrogen plus thiosulfate). These strains had similar DNA G+C content (60–64 mol%) to and high DNA:DNA homology (> 75%) with the reference strain of Bacillus schlegelii. This is the first report of thermogenic composts as habitats of thermophilic sulfur- and hydrogen-oxidizing bacteria, which to date have been known only from geothermal manifestations. This contrasts with the generally held belief that thermogenic composts at temperatures above 60° C support only a very low diversity of obligatory heterotrophic thermophiles related to Bacillus stearothermophilus. Received: 20 July 1995 / Accepted: 25 September 1995     

Thermococcus siculi sp. nov., a novel hyperthermophilic archaeon isolated from a deep-sea hydrothermal vent at the Mid-Okinawa Trough

A novel coccoid-shaped, hyperthermophilic, anaerobic archaeon, strain RG-20, was isolated from a deep-sea hydrothermal vent fluid sample taken at 1394-m depth at the Mid-Okinawa Trough (27°32.7′N, 126°58.5′E). Cells of this isolate occur singly or in pairs and are about 0.8 to 2 μm in diameter. Growth was observed at temperatures between 50° and 93°C, with an optimum at 85°C. The pH range for growth is 5.0–9.0, with an optimum around 7.0. Strain RG-20 requires 1%–4% of NaCl for growth, and cell lysis occurs at concentrations below 1%. The newly isolated strain grows preferentially in the presence of elemental sulfur on proteinaceous substrates such as yeast extract, peptone, or tryptone, and no growth was observed on carbohydrates, carboxylic acids, alcohols, or lipids. This microorganism is resistant to streptomycin, chloramphenicol, ampicillin, and kanamycin at concentrations up to 150 μg/ml, but is susceptible to rifampicin. Analysis of the hydrolyzed core lipids by thin-layer chromatography (TLC) revealed the presence of archaeol and caldarchaeol. The mol% G+C content of the DNA is 55.8. Partial sequencing of the 16S rDNA indicates that strain RG-20 belongs to the genus Thermococcus. Considering these data and on the basis of the results from DNA-DNA hybridization studies, we propose that this strain should be classified as a new species named Thermococcus siculi (si′cu.li. L. gen. n. siculi, of the deep-sea [siculum, deep-sea in literature of Ovid], referring to the location of the sample site, a deep-sea hydrothermal vent). The type strain is isolate RG-20 (DSM No. 12349). Received: May 11, 1998 / Accepted: July 24, 1998     

<Emphasis Type="Italic">Ancylobacter abiegnus</Emphasis> sp. nov., an oligotrophic member of the xylotrophic mycobacterial community

A new species of the genus Ancylobacter, exemplified by strain Z-0056, was isolated from dystrophic humified waters formed by xylotrophic fungi grown on decaying spruce wood. The cells of strain Z-0056 (0.65–0.9 μm) are coccoid, gram-negative, with fimbriae, and nonmotile. The strain is pleomorphic and reproduces by nonuniform division. Strain Z-0056 is an aerobic organoheterotroph and a mesophilic and moderately acidophilic oligotrophic microorganism. As an inhabitant of dystrophic ultrafresh waters, strain Z-0056 is sensitive to NaCl. The bacterium utilizes organic acids (acetate, pyruvate, oxalate, gluconate, malate, succinate, and citrate), as well as xylose and xylan. The microorganism grows in a pH range of 4.0–8.0, with an optimum at pH 5.5. The temperature range for growth is 15–25°C, with an optimum at 20°C. According to its ecophysiological properties, strain Z-0056 belongs to the group of ombrophilic dissipotrophs. The DNA G+C base content is 66.8 mol %. According to phylogenetic analysis, strain Z-0056 belongs to the genus Ancylobacter. Strain Z-0056 showed the highest similarity (98.3%) with the type strain of the species A. oerskovii. The phenotypic and phylogenetic properties of strain Z-0056 support classification of this microorganism within the genus Ancylobacter as the novel species Ancylobacter abiegnus sp. nov.     

Elevated content of hsp 70 does not induce tolerance of sporulation to higher temperature

The temperature permissive for sporulation (up to 42°C) inBacillus megaterium is by 4–5°C lower than that for its growth (up to 46–47°C). The ability ofB. megaterium cells to synthesize and degrade stress proteins under incubation in the sporulation medium was therefore investigated. The higher level of hsp 70, a typical stress protein induced by a temperature shock in postexponential growth phase, did not increase the permissive temperature of sporulation. The hsp 70 protein did not undergo a rapid turnover and its portion in the soluble protein fraction did not drop for at least 6 h at a temperature that was nonpermissive for sporulation (43.5°C). On the other hand, the elevated level of hsp 70 could not bring about the inhibition of sporulation as it was retained in the cells even after a shift of the temperature to 35°C, permitting sporulation of the culture.     

Specific characteristics of the strains isolated from a thermoacidophilic microbial community oxidizing antimony sulfide ore

Investigation of the phenotypic properties of three mixotrophic bacteria, strains Sb-K, Sb-F, and Sb-S, isolated from an aboriginal thermoacidophilic microbial community participating in biooxidation of ore with high antimony content (26%) and ore concentrates from the Olympiadinskoe deposit under semicontinuous cultivation conditions at 46 ± 1°C, revealed the differentiating characteristics of these strains. The isolated cultures grew lithotrophically through different numbers of transfers: strains Sb-F and Sb-K grew through seven and eight transfers, respectively, and strain Sb-S grew through two or three transfers. Strains Sb-K and Sb-S utilized a wide range of organic substrates for active organotrophic growth during nine or ten transfers, while strain Sb-F was less tolerant to organic compounds. Strain Sb-K grew on a medium with the ore and sulfide ore concentrates in the pH range of 1.0–3.0. Growth of strains Sb-F and Sb-S occurred in the pH ranges of 1.0–2.5 and 1.5–5.5 on media with Fe²⁺ and S⁰, respectively.. The optimal initial pH values of the media, corresponding to the maximum specific growth rates, were 1.6–1.7, 1.9, and 2.0–3.0 for strains Sb-K, Sb-F, and Sb-S, respectively. All three strains were able to grow within a broad temperature range, 20–65°C, with an optimum at 46°C (Sb-K), 40–46°C (Sb-F), and 48–50°C (Sb-S). According to the results of DNA-DNA hybridization and phylogenetic analysis, as well as their phenotypic characteristics, the isolates can be classified as novel strains of species of the genus Sulfobacillus. Strains Sb-K, Sb-F, and Sb-S, isolated as predominant cultures on the media with sulfide compounds, iron, or sulfur, respectively, were affiliated to the species S. thermotolerans, S. sibiricus, and S. thermosulfidooxidans.     

Thermovorax subterraneus, gen. nov., sp. nov., a thermophilic hydrogen-producing bacterium isolated from geothermally active underground mine

A thermophilic, rod-shaped, motile, Gram-positive, spore-forming bacterium strain 70B^T was isolated from a geothermally active underground mine in Japan. The temperature and pH range for growth was 50–81°C (optimum 71°C) and 6.2–9.8 (optimum pH 7–7.5), respectively. Growth occurred in the presence 0–2% NaCl (optimum 1% NaCl). Strain 70B^T could utilize glucose, fructose, mannose, mannitol, pyruvate, cellobiose and tryptone as substrates. Thiosulfate was used as electron acceptor. Major whole-cell fatty acids were iso-C_15:0, C_16:0 DMA (dimethyl acetal), C_16:0 and anteiso-C_15:0. The G+C mol% of the DNA was 44.2%. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that the closest relatives of strain 70B^T were Thermosediminibacter oceani DSM 16646^T (94% similarity) and Thermosediminibacter litoriperuensis DSM 16647 (93% similarity). The phenotypic, chemotaxonomic and phylogenetic properties suggest that strain 70B^T represents a novel species in a new genus, for which the name Thermovorax subterraneus gen. nov., sp. nov. is proposed. The type strain of Thermovorax subterraneus is 70B^T (=DSM 21563 = JCM 15541).     

Characterization of Halanaerobaculum tunisiense gen. nov., sp. nov., a new halophilic fermentative, strictly anaerobic bacterium isolated from a hypersaline lake in Tunisia

A new halophilic anaerobe was isolated from the hypersaline surface sediments of El-Djerid Chott, Tunisia. The isolate, designated as strain 6SANG, grew at NaCl concentrations ranging from 14 to 30%, with an optimum at 20–22%. Strain 6SANG was a non-spore-forming, non-motile, rod-shaped bacterium, appearing singly, in pairs, or occasionally as long chains (0.7–1 × 4–13 μm) and showed a Gram-negative-like cell wall pattern. It grew optimally at pH values between 7.2 and 7.4, but had a very broad pH range for growth (5.9–8.4). Optimum temperature for growth was 42°C (range 30–50°C). Strain 6SANG required yeast extract for growth on sugars. Glucose, sucrose, galactose, mannose, maltose, cellobiose, pyruvate, and starch were fermented. The end products from glucose fermentation were acetate, butyrate, lactate, H₂, and CO₂. The G + C ratio of the DNA was 34.3 mol%. Strain 6SANG exhibited 16S rRNA gene sequence similarity values of 91–92% with members of the genus Halobacteroides, H. halobius being its closest phylogenetic relative. Based on phenotypic and phylogenetic characteristics, we propose that this bacterium be classified as a novel species of a novel genus, Halanaerobaculum tunisiense gen. nov., sp. nov. The type strain is 6SANG^T (=DSM 19997^T = JCM 15060^T).     

Sunfleck dynamics and canopy structure in a Phaseolus vulgaris L. canopy

 Photosynthetic photon flux density (PPFD) fluctuations were quantified in crops of beans (Phaseolus vulgaris L.) in the field as the canopy developed between July and October. Two different methods were used to select sunflecks and shadeflecks. Four ranges of zenith angles (60–70°, 50–60°, 40–50° and 30–40°) were selected for analysing PPFD fluctuations. At the base of the canopy, sunflecks contributed 18%, 53%, 10% and 4% during the 1st, 3rd, 5th and 7th week of growth, respectively. At a height of 20 cm above the soil surface, the respective contributions were 28% and 21% during the 6th and 7th weeks. Sunfleck lengths of 0–5 s were the most frequent, with the greatest number being found with smaller zenith angles. The proportion of short duration sunflecks increased as the growth period advanced. The number of long sunflecks decreased with time, with very few longer than 100 s by the 5th and 7th weeks. The distributions of sunfleck irradiance were similar to normal distributions and irradiance ranged in μmol m⁻² s⁻¹ from 600–900, 800–1500 and 1000–1600 respectively at zenith angles of 50–60°, 40–50° and 30–40°. A multiple regression showed that short sunflecks (<100 s) depended on zenith angle, plant height, and leaf and stem area index (L _s ), whereas long sunflecks (>100 s) depended on zenith angle and L _s. Shadefleck distributions were similar to those for sunflecks but there were fewer of the shortest examples and more of the longest. The best statistical distribution to describe sunflecks and shadeflecks was the gamma distribution, which could provide the basis for the future development of a good model for sunfleck and shadefleck distributions. Received: 10 November 1997 / Accepted: 2 April 1998     

Thermophilic protease-producing Geobacillus from Buranga hot springs in Western Uganda

Two proteolytic thermophilic aerobic bacterial strains, PA-9 and PA-5, were isolated from Buranga hot springs in western Uganda. The cells were rods, approximately 10–12 μm in length and 3 μm in width. Isolate PA-9 grew at between 38°C and 68°C (optimum, 62°C), and PA-5 grew at between 37°C and 72°C (optimum, 60°C). Both isolates grew optimally at pH 7.5–8.5. Their 16S rRNA gene sequences indicated that they belong to the newly described genus Geobacillus. Zymogram analysis of the crude enzyme extracts revealed the presence of two extracellular proteases for isolate PA-5, and at least eight for isolate PA-9. The optimum temperature and pH for casein-degrading activity were 70°C, pH 6.5 for isolate PA-9, but caseinolytic activity could also be observed at 2°C. In the case of isolate PA-5, optimal activity was observed over a temperature and pH range of 50–70°C and pH 5–10, respectively. Received: 26 November 2001 / Accepted: 12 December 2001     

Characterization of Microaerobacter geothermalis gen. nov., sp. nov., a novel microaerophilic, nitrate- and nitrite-reducing thermophilic bacterium isolated from a terrestrial hot spring in Tunisia

A novel thermophilic anaerobic and microaerophilic bacterium (optimal growth in the presence of 5–10% O₂), strain Nad S1^T was isolated from the terrestrial hot spring of Hammam Sidi Jdidi, Nabeul, Tunisia. Cells were motile rods having a Gram-positive cell wall structure. Strain Nad S1^T grew optimally at 55°C (range 37–70°C). Optimum pH for growth was 6.5–7.0. It was halotolerant growing with NaCl up to 7% (optimum concentration 1.5–3.0%). It grew chemoorganotrophically on various carbohydrates, organic-acids and amino-acids as energy sources, or chemolithotrophically on H₂ using nitrate, as terminal electron acceptor. Beside oxygen (under microaerobic conditions) and nitrate, nitrite was also used. Nitrate was completely reduced to N₂. No fermentation occurred. The genomic DNA G + C content was 41.8 mol%. Based on 16S rRNA gene sequence analysis, strain Nad S1^T belongs to the Bacillaceae family within the class ‘Bacilli’. Because of its phylogenetic and phenotypic characteristics, we propose this isolate to be assigned as a novel genus and a novel species within the domain Bacteria, Microaerobacter geothermalis gen. nov., sp. nov. The type strain is Nad S1^T (=DSM 22679^T =JCM 16213^T).     

Isolation and characterization of a new CO-utilizing strain, Thermoanaerobacter thermohydrosulfuricus subsp. carboxydovorans, isolated from a geothermal spring in Turkey

A novel anaerobic, thermophilic, Gram-positive, spore-forming, and sugar-fermenting bacterium (strain TLO) was isolated from a geothermal spring in Ayaş, Turkey. The cells were straight to curved rods, 0.4–0.6 μm in diameter and 3.5–10 μm in length. Spores were terminal and round. The temperature range for growth was 40–80°C, with an optimum at 70°C. The pH optimum was between 6.3 and 6.8. Strain TLO has the capability to ferment a wide variety of mono-, di-, and polysaccharides and proteinaceous substrates, producing mainly lactate, next to acetate, ethanol, alanine, H₂, and CO₂. Remarkably, the bacterium was able to grow in an atmosphere of up to 25% of CO as sole electron donor. CO oxidation was coupled to H₂ and CO₂ formation. The G + C content of the genomic DNA was 35.1 mol%. Based on 16S rRNA gene sequence analysis and the DNA–DNA hybridization data, this bacterium is most closely related to Thermoanaerobacter thermohydrosulfuricus and Thermoanaerobacter siderophilus (99% similarity for both). However, strain TLO differs from Thermoanaerobacter thermohydrosulfuricus in important aspects, such as CO-utilization and lipid composition. These differences led us to propose that strain TLO represents a subspecies of Thermoanaerobacter thermohydrosulfuricus, and we therefore name it Thermoanaerobacter thermohydrosulfuricus subsp. carboxydovorans.     

Thermococcus peptonophilus sp. nov., a fast-growing,extremely thermophilic archaebacterium isolated from deep-sea hydrothermal vents

Two extremely thermophilic archaebacteria, strains OG-1 and SM-2, were isolated from newly discovered deep-sea hydrothermal vent areas in the western Pacific ocean. These strains were cocci, obligately anaerobic Archaea about 0.7–2 μm in diameter. Optimum growth conditions for OG-1 and SM-2 were at 85–90°C (range 60–100°C), pH 6 (range pH 4–8), a NaCl concentration of 3% (range 1–5%), and a nutrient concentration (tryptone plus yeast extract) of 0.2% (range 0.005–5%). Elemental sulfur stimulated the growth rate fourfold. Ammonium slightly stimulated growth. Both tryptone and yeast extract allowed growth as sole carbon sources; these isolates were not able to utilize or grow exclusively on sucrose, glucose, maltose, succinate, pyruvate, propionate, acetate, or free amino acids. OG-1 showed the fastest growth rate within the genus Thermococcus. Growth was inhibited by rifampicin. The DNA G+C content was 52 mol%. Sequencing of their 16S rDNA gene fragment indicated that these isolates belonged to the genus Thermococcus. OG-1 and SM-2 were different than the described Thermococcus species. We propose that OG-1 belongs to a new species: Thermococcus peptonophilus. Received: 8 March 1995 / Accepted: 24 May 1995     

The time-profile of the PBMC HSP70 response to in vitro heat shock appears temperature-dependent

Summary. Heat shock proteins (HSPs) are synthesised by cells subsequent to a stress exposure and are known to confer protection to the cell in response to a second challenge. HSP induction and decay are correlated to thermotolerance and may therefore be used as a biomarker of thermal history. The current study tested the temperature-dependent nature of the heat shock response and characterised its time profile of induction. Whole blood from 6 healthy males (Age: 26 ± (SD) 2 yrs; Body mass 74.2 ± 3.8 kgs; VO_2max: 49.1 ± 4.0 ml·kg⁻¹·min⁻¹) were isolated and exposed to in vitro heat shock (HS) at 37, 38, 39, 40, and 41 °C for a period of 90 min. After HS the temperature was returned to 37 °C and intracellular HSP70 was quantified from the leukocytes at 0, 2, 4, and 6 h after heat treatment. The concentration of HSP70 was not different between temperatures (P > 0.05), but the time-profile of HSP70 synthesis appeared temperature-dependent. At control (37 °C) and lower temperatures (38–39 °C) the mean HSP70 concentration increased up to 4 h post HS (P < 0.05) and then returned towards baseline values by 6 h post HS. With in vitro hyperthermic conditions (40–41 °C), the time-profile was characterised by a sharp rise in HSP70 levels immediately after treatment (P < 0.05 for 40 °C at 0 h), followed by a progressive decline over time. The results suggest a temperature-dependent time-profile of HSP70 synthesis. In addition, the temperature at which HSP70 is inducted might be lower than 37 °C.     

The bifunctional enzyme chitosanase-cellulase produced by the gram-negative microorganism Myxobacter sp. AL-1 is highly similar to Bacillus subtilis endoglucanases

The gram-negative bacterium Myxobacter sp. AL-1 produces chitosanase-cellulase activity that is maximally excreted during the stationary phase of growth. Carboxymethylcellulase zymogram analysis revealed that the enzymatic activity was correlated with two bands of 32 and 35 kDa. Ion-exchange-chromatography-enriched preparations of the 32-kDa enzyme were capable of degrading the cellulose fluorescent derivatives 4-methylumbelliferyl-β-d-cellobioside and 4-methylumbelliferyl-β-d-cellotrioside. These enzymatic preparations also showed a greater capacity at 70° C than at 42° C to degrade chitosan oligomers of a minimum size of six units. Conversely, the β-1,4 glucanolytic activity was more efficient at attacking carboxymethylcellulose and methylumbelliferyl-cellotrioside at 42° C than at 70° C. The 32-kDa enzyme was purified more than 800-fold to apparent homogeneity by a combination of ion-exchange and molecular-exclusion chromatography. Amino-terminal sequencing indicated that mature chitosanase-cellulase shares more than 70% identity with endocellulases produced by strains DLG, PAP115, and 168 of the gram-positive microorganism Bacillus subtilis. Received: 6 January 1997 / Accepted: 29 May 1997     

Isolation and characterization of a novel thermophilic Bacillus strain degrading long-chain n-alkanes

A thermophilic Bacillus strain NG80-2 growing within the temperature range of 45–73°C (optimum at 65°C) was isolated from a deep subterranean oil-reservoir in northern China. The strain was able to utilize crude oil and liquid paraffin as the sole carbon sources for growth, and the growth with crude oil was accompanied by the production of an unknown emulsifying agent. Further examination showed that NG80-2 degraded and utilized only long-chain (C15–C36) n-alkanes, but not short-chain (C8–C14) n-alkanes and those longer than C40. Based on phenotypic and phylogenic analyses, NG80-2 was identified as Geobacillus thermodenitrificans. The strain NG80-2 may be potentially used for oily-waste treatment at elevated temperature, a condition which greatly accelerates the biodegradation rate, and for microbial enhancing oil recovery process.Lei Wang, Yun Tang and Shuo Wang contributed equally to this study.     

Control and possible applications of a novel carrot-spoilage basidiomycete, Fibulorhizoctonia psychrophila

A novel cold-tolerant fungus, Fibulorhizoctonia psychrophila, was isolated from a refrigerated carrot storage facility and identified as an anamorph of Athelia, often classified in Rhizoctonia s.l. Growth of this fungus was observed between 0 and 20°C with an optimum at 9–12°C, while incubation of mycelium grown at 15–32°C resulted in absence of growth even after the fungus was transferred back to 15°C. Growth was inhibited in the presence of the antifungals sorbic acid or natamycin, in particular when the fungus was incubated at 18°C. F. psychrophila produces polysaccharide degrading enzymes that, when compared to enzymes from the ascomycete fungus Aspergillus niger, retain a larger proportion of their activity at lower temperatures. This indicates that F. psychrophila could be used as a source for novel industrial enzymes that are active at 4–15°C.     

Expression and characterization of a novel heterologous moderately thermostable lipase derived from metagenomics in <Emphasis Type="Italic">Streptomyces lividans</Emphasis>

Seven lipolytic genes were isolated and sequenced from a metagenomic library that was constructed following biomass enrichment in a fed-batch bioreactor submitted to high temperature (50–70°C) and alkaline pH (7–8.5). Among those sequences, lipIAF1-6 was chosen for further study and cloned in Streptomyces lividans 10–164. The G+C content within the sequence was 64.3%. The encoded protein, LipIAF1-6, was related to various putative lipases previously identified in different genome sequences. Homology of LipIAF-6 with the different lipases did not exceed 31%. The optimum pH (8.5) and temperature (60°C) of the purified enzyme were in agreement with the enrichment conditions. Furthermore, the enzyme was thermostable for as long as 30 min at 70°C. The maximum activity of the purified lipase was 4,287 IU/mg towards p-nitrophenyl (p-NP) butyrate (60°C; pH 8.5). LipIAF1-6 does not seem to need the presence of metal ions for its activity. The enzyme was slightly inhibited by 10 mM CoCl₂ (14%), HgCl₂ (12%), and dithiothreitol (DTT) (15%). The serine protease inhibitor phenylmethylsulphonyl fluoride (PMSF) reduced activity by 39% and 71% when incubated at concentrations of 1 and 10 mM, respectively. Finally, LipIAF1-6 was stable in different organic solvents, and against several surfactants and oxidative agents commonly found in detergent formulations. These results are quite encouraging for further use of this enzyme in different industrial processes.