Isolation and Characterization of High-Strength Phenol-Degrading Novel Bacterium of the Pantoea Genus

A new indigenous soil bacterium Pantoea strain NII-153 utilizing phenol as a sole carbon source was isolated and characterized. Phylogenetic analysis suggested its classification to the Enterobacteriaceae family, with 95.0% gene sequence similarity to Pantoea ananatis ATCC 33244. Biodegradation rates of phenol by NII-153 were found to be more effective at 64 h with initial concentration of 600 mg L^{? 1} of phenol and this is the first report of such activity in Pantoea species. Strain NII-153 has showed high tolerance to phenol concentration (900 mg L^{? 1}). Therefore, strain NII-153 could be used for biotreatment of high-strength phenol-containing industrial effluents and for bioremediation of phenol-contaminated soils.     

焦化废水中4株苯酚高效降解菌的分离及鉴定

目的:从焦化废水中筛选苯酚高效降解菌并进行鉴定.方法:在100～1000 mg/L的苯酚为惟-碳源的无机盐培养基上分离出单菌落,测定各菌株的生长曲线以及对苯酚的降解效牢;利用 16S rDNA序列分析结合菌株的形态特征确定各菌株的分类地位.结果:筛选获得4株苯酚降解菌,均能够以苯酚为惟一碳源,在30℃、pH7.0、摇床转速130 r/min、2%的接种量条件下,24h内能将1 000mg/L的苯酚降解91%以上;4株菌可初步鉴定为芽孢杆菌属(ZL1)、产碱杆菌属(ZL2、ZL4)、沙雷氏菌属(ZL3).其中,从焦化废水中分离出高效降解苯酚的沙雷氏菌未见报道.结论:从焦化废水中获得4株苯酚高效降解细菌,对高浓度含酚废水的生物降解具有潜在的应用前景.     

一株苯酚高效降解菌的分离及其分解能力的初步研究

为了寻找能高效降解苯酚的微生物 ,从武汉市某化工厂周围的下水道污泥中 ,筛选分离出一种具有很高苯酚降解能力的菌株PheD1。通过生理生化及外观鉴定[1～ 3] ,将其初步鉴定为假单胞菌 (Pseudomonassp .)。经驯化后发现 ,该菌生长的迟缓期随苯酚浓度的增大而延长 ,但比同类报道的苯酚降解菌要短 ;在 35℃对数生长期时的苯酚降解率超过同类报道[6 ] ,6 0 0mg·L- 1 苯酚浓度的完全降解时间在 2 4h之内 ,比同类报道[4～ 6 ] 苯酚降解菌的分解能力要高。该菌为好氧菌 ,在空气充足的条件下可提高降解能力。对该菌的继续研究可使其在苯酚的生物降解及污水处理等实际运用中起到重要的作用。     

Isolation and Characterization of Methanogenic Bacteria from Landfills

Methanogenic bacteria were isolated from landfill sites in the United Kingdom. Strains of Methanobacterium formicicum, Methanosarcina barkeri, several different immunotypes of Methanobacterium bryantii, and a coccoid methanogen distinct from the reference immunotypes were identified.     

Isolation and Characterization of Biosurfactant-Producing Bacteria from Oil-Contaminated Soil

Two biosurfactant-producing Pseudomonas aeruginosa strains (KISR C1 and KISR B1) were isolated from Kuwaiti oil-contaminated soil, which resulted from the Gulf War. The optimum environmental conditions that supported the growth and surfactant production of both isolates were examined. The two isolates differed in their biosurfactant-stimu-lating carbon source, nitrogen concentration, and the pH of the medium. C-1 isolate produced two types of rhamnolipids with a final concentration of 98.4?g/l after spiking the nitrogen-limited medium with 10?mg/ml olive oil. The other isolate (B-1) produced only one type of rhamnolipid (5.9?g/l) after spiking the medium with crude oil. The biosurfactant produced by this strain was found to be very effective in the emulsifica-tion of crude oil. The result suggests that this isolate can potentially be used to enhance bioremediation of oil-contamination and enhanced oil recovery.     

Isolation and Characterization of Cellulolytic Bacteria from Activated Sludge

Cellulolytic aerobic bacteria were isolated from activated sludge systems. Of the media tested for enumeration, only filter paper media gave reliable counts. Five isolates were studied further for characterization. It was found that one strain (DK) belonged to the genus Cellulomonas. The other four strains expressed similarity to the genus Pseudomonas. The different characteristics that were studied, however, do not permit them to be identified with any recognized species. Based on certain characters we believe that they are alcaligenes-like pseudomonads.     

Isolation and Characterization of Bacteria from the Baltic Sea

A bacteriological examination was done on samples of water and sediment from three localities in the Baltic. The highest numbers of bacteria were recovered from areas subjected to pollution. The isolates included members of the family Enterobacteria-ceae, the genus Pseudomonas and strains of Aeromonas hydrophila, Alteromonas putrefaciens and some Gram positive bacteria. It is suggested tentatively that H2S production in the black sediments was caused by Alt. putrefaciens. None of the isolates had an absolute requirement for NaCl, although all of them were salt-tolerant to varying degrees, and most were able to grow aerobically at salinities comparable with those found in seawater. Isolates belonging to the family Enterobacteriaceae were, however, unable to grow anaerobically under comparable conditions. Freshwater strains of several genera of the family Enterobacteriaceae and of Aeromonas hydrophila and Aer. sobria displayed salt tolerance identical with that of the Baltic isolates. One strain each of Escherichia coli, Klebsiella pneumoniae and Yersinia enterocolitica survived well during three weeks at 17°C in artificial seawater lacking both carbon and nitrogen sources. These results suggest the need for a re-evaluation of the persistence of potentially pathogenic bacteria in the sea.     

大豆内生细菌的分离及根腐病拮抗菌的筛选鉴定

内生细菌存在于健康植物体内,一些内生细菌具有促生长、抗病和固氮等生物学功能.本项研究采用化学药剂表面灭菌方法从黑龙江省大豆品种合丰25的根、茎、叶和种子中分离到大量内生细菌,其种群数量在根部最多,为3.4×103CFU/g,在叶部次之,为2.8×103CFU/g,在茎部和种子中最少,为2.9×102 CFU/g和1.4×102CFU/g.从121株内生细菌中筛选到31株对大豆根腐病菌Fusarium oxysporum f. sp.soybean具有较强抑制作用的拮抗内生细菌,其中菌株TF28抑菌谱广,抑菌率高,对不同植物的病原菌F. oxysporum的抑菌率为80.2%～96.7%.经形态、生理生化和16S rRNA鉴定为解淀粉芽孢杆菌(Bacillus amyloliquefaciens).     

Isolation and Characterization of Coumaphos-Metabolizing Bacteria from Cattle Dip

Coumaphos, an organophosphate insecticide, is used for tick control in cattle dipping vats along the U.S.-Mexican border. Recently, several vats (problem vats) have experienced a loss of efficacy because of microbial degradation. Three morphologically distinct bacteria (designated B-1, B-2, and B-3) that metabolized coumaphos were isolated from enrichment cultures that were initiated from problem vat dip material. In general, amino acids, pyrimidines, and acetate supported growth; carbohydrates were not utilized. Only B-2 required growth factors. In resting cell experiments, coumaphos was hydrolyzed to diethylthiophosphoric acid and chlorferon by all three isolates. Chlorferon was subsequently metabolized by B-1 and B-2 to α-chloro-β-methyl-2,3,4-trihydroxy-trans-cinnamic acid. Only B-1 produced additional metabolites. Experiments with [benzo ring-labeled U-¹⁴C]coumaphos or chlorferon demonstrated that B-1 was capable of both mineralizing and incorporating into biomass the aromatic portion of the molecule. The majority of label, however, was recovered in the form of soluble products, including α-chloro-β-methyl-2,3,4-trihydroxy-trans-cinnamic acid. Although B-1 had the capacity to use chlorferon as a carbon source at low concentrations (100 μg/ml), visible growth at higher concentrations (1,000 μg/ml) was not observed. The addition of 400 μg of chlorferon per ml to B-1 cells in the mid-log phase of growth resulted in complete inhibition of growth, while the addition of 100 to 200 μg of chlorferon per ml resulted in partial inhibition. The growth of B-2 and B-3 was inhibited by 100 μg of chlorferon per ml. These data suggest that, although B-1 and, to a lesser extent, B-2 and B-3 are responsible for the primary degradation of coumaphos, other organisms in the enrichment culture may play a secondary role in coumaphos degradation by removing inhibitory products of coumaphos metabolism.     

苯酚高效降解菌的筛选和降解特性的研究

从天津市煤气厂的活性污泥中筛选、分离得到一株高效苯酚降解菌。经BIOLOG细菌自动鉴定系统及16SrDNA鉴定,该菌株为粪产碱杆菌(Alcaligenesfaecalis)。苯酚降解实验证实,该菌能在76h内完全降解1600mg·L-1的苯酚,并且随着苯酚浓度的增加,底物抑制作用增强,细胞得率下降。     

Isolation and Characterization of Polymeric Galloyl-Ester-Degrading Bacteria from a Tannery Discharge Place

The culturable bacteria colonizing the rhizosphere of plants growing in the area of discharge of a tannery effluent were characterized. Relative proportions of aerobic, denitrifying, and sulfate-reducing bacteria were determined in the rhizosphere of Typha latifolia, Canna indica, and Phragmites australis. Aerobic bacteria were observed to be the most abundant group in the rhizosphere, and plant type did not seem to influence the abundance of the bacterial types analyzed. To isolate bacteria able to degrade polyphenols used in the tannery industry, enrichments were conducted under different conditions. Bacterial cultures were enriched with individual polyphenols (tannins Tara, Quebracho, or Mimosa) or with an undefined mixture of tannins present in the tannery effluent as carbon source. Cultures enriched with the effluent or Tara tannin were able to degrade tannic acid. Six bacterial isolates purified from these mixed cultures were able to use tannic acid as a sole carbon source in axenic culture. On the basis of 16S ribosomal DNA sequence analysis, these isolates were closely related to organisms belonging to the taxa Serratia, Stenotrophomonas maltophilia, Klebsiella oxytoca, Herbaspirillum chlorophenolicum, and Pseudomonas putida.     

一组优势菌对焦化废水中吲哚、吡啶的降解条件的实验研究

从普通变形杆菌BH、芽孢杆菌DC4 5、巨大芽孢杆菌F3、枯草芽孢杆菌DC4 2等 4种脱色菌中 ,经驯化、筛选出普通变形杆菌BH、芽孢杆菌DC4 5两株降解苯酚、吲哚、吡啶、喹啉的优势菌。其中普通变形杆菌BH降解吡啶的适宜条件是 :温度 30～ 4 5℃ ,pH 7～ 8,投加适量Pb2 + ;芽孢杆菌DC4 5降解吲哚的适宜条件是 :温度 2 0～ 5 0℃ ,pH 6～ 8,投加适量Zn2 + ,Fe2 + 。     

Isolation and Chemical Characterization of Lipid A from Gram-negative Bacteria

Lipopolysaccharide (LPS) is the major cell surface molecule of gram-negative bacteria, deposited on the outer leaflet of the outer membrane bilayer. LPS can be subdivided into three domains: the distal O-polysaccharide, a core oligosaccharide, and the lipid A domain consisting of a lipid A molecular species and 3-deoxy-D-manno-oct-2-ulosonic acid residues (Kdo). The lipid A domain is the only component essential for bacterial cell survival. Following its synthesis, lipid A is chemically modified in response to environmental stresses such as pH or temperature, to promote resistance to antibiotic compounds, and to evade recognition by mediators of the host innate immune response. The following protocol details the small- and large-scale isolation of lipid A from gram-negative bacteria. Isolated material is then chemically characterized by thin layer chromatography (TLC) or mass-spectrometry (MS). In addition to matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) MS, we also describe tandem MS protocols for analyzing lipid A molecular species using electrospray ionization (ESI) coupled to collision induced dissociation (CID) and newly employed ultraviolet photodissociation (UVPD) methods. Our MS protocols allow for unequivocal determination of chemical structure, paramount to characterization of lipid A molecules that contain unique or novel chemical modifications. We also describe the radioisotopic labeling, and subsequent isolation, of lipid A from bacterial cells for analysis by TLC. Relative to MS-based protocols, TLC provides a more economical and rapid characterization method, but cannot be used to unambiguously assign lipid A chemical structures without the use of standards of known chemical structure. Over the last two decades isolation and characterization of lipid A has led to numerous exciting discoveries that have improved our understanding of the physiology of gram-negative bacteria, mechanisms of antibiotic resistance, the human innate immune response, and have provided many new targets in the development of antibacterial compounds.     

Isolation and Characterization of Dehalogenases from 2,2-Dichloropropionate-Degrading Soil Bacteria

Five bacterial strains were isolated from polluted soils capable of degrading 2,2-dichloropropionate. In crude extracts, dehalogenase activity against haloacetates and longer-chained 2-haloalkanoic acids could be detected. Results from activity staining indicated that all bacterial strains expressed a single dehalogenase. In further biochemical characterization, two types of D,L-specific 2-haloalkanoic acid dehalogenases were described, which are different from each other not only in molecular weight and electrophoretic mobility, but also in sensitivity towards thiol reagents. Dehalogenases of these strains have been shown to be inducible and are catalyzing halide hydrolysis with inversion of product configuration. Received: 5 July 1996 / Accepted: 1 August 1996     

Isolation and Characterization of Bacteria from the Swedish West Coast

Most of the bacteria isolated from water and sediment samples from a locality off the west coast of Sweden had an absolute requirement for Na+. On the basis of phenotypic characterization and determination of DNA base composition, the strains could be assigned to the genera Beneckea, Alteromonas and Pseudomonas. Apart from a group of sulphide-forming alteromonads, none of the isolates appeared to be identical with organisms described previously.     

Isolation and Characterization of Diuron-degrading Bacteria from Lotic Surface Water

The bacterial community structure of a diuron-degrading enrichment culture from lotic surface water samples was analyzed and the diuron-degrading strains were selected using a series of techniques combining temporal temperature gradient gel electrophoresis (TTGE) of 16 S rDNA gene V1–V3 variable regions, isolation of strains on agar plates, colony hybridization methods, and biodegradation assays. The TTGE fingerprints revealed that diuron had a strong impact on bacterial community structure and highlighted both diuron-sensitive and diuron-adapted bacterial strains. Two bacterial strains, designated IB78 and IB93 and identified as belonging to Pseudomonas sp. and Stenotrophomonas sp., were isolated and shown to degrade diuron in pure resting cells in a first-order kinetic reaction during the first 24 h of incubation with no 3,4-DCA detected. The percentages of degradation varied from 25% to 60% for IB78 and 20% to 65% for IB93 and for a diuron concentration range from 20 mg/L to 2 mg/L, respectively. It is interesting to note that diuron was less degraded by single isolates than by mixed resting cells, thereby underlining a cumulative effect between these two strains. To the best of our knowledge, this is the first report of diuron-degrading strains isolated from lotic surface water.     

Isolation and Characterization of Extremely Thermophilic Bacteria from Hot Springs

In order to investigate the mechanism of microbial growth at elevated temperatures, it was tried to isolate different thermophilic microorganisms from wide origins, such as soils, composts, manure piles and hot spring waters. As the result, 5 strains of extremely thermophilic bacteria, the maximum, the optimum and the minimum temperatures for growth of which were 80, 70~75, and 40°C, respectively, were isolated from Izu-Atagawa hot spring and Beppu hot springs. These bacteria were gram-negative, yellow-pigmented, non-motile and non-sporulating rods of 0.5~0.7 μ in diameter and 2~5 μ in length. They were heterotrophs requiring several amino acids (such as glutamate, aspartate, et al.) and vitamins (such as biotin, folic acid and p-aminobenzoic acid) and grew well at neutral to slight alkali pH. The content of GC pairs of DNAs from the 5 strains was 69~70%, and this seemed to be one of the highest values in bacteria so far known. Among the 5 strains, strain AT–62 was named as Thermus flavus sp. n. AT–62 from its morphological and physiological characteristics. Comparison between Thermus flavus and other extremely thermophilic bacteria as Thermus aquaticus and Flavobacterium thermophilum is described and discussed in reference to classification of extremely thermophilic bacteria.     

Isolation and Characterization of Polyester-Based Plastics-Degrading Bacteria from Compost Soils

Four potential polyester-degrading bacterial strains were isolated from compost soils in Thailand. These bacteria exhibited strong degradation activity for polyester biodegradable plastics, such as polylactic acid (PLA), polycaprolactone (PCL), poly-(butylene succinate) (PBS) and polybutylene succinate-co-adipate (PBSA) as substrates. The strains, classified according to phenotypic characteristics and 16S rDNA sequence, belonging to the genera Actinomadura, Streptomyces and Laceyella, demonstrated the best polyester- degrading activities. All strains utilized polyesters as a carbon source, and yeast extract with ammonium sulphate was utilized as a nitrogen source for enzyme production. Optimization for polyester-degrading enzyme production by Actinomadura sp. S14, Actinomadura sp. TF1, Streptomyces sp. APL3 and Laceyella sp. TP4 revealed the highest polyester-degrading activity in culture broth when 1% (w/v) PCL (18 U/mL), 0.5% (w/v) PLA (22.3 U/mL), 1% (w/v) PBS (19.4 U/mL) and 0.5% (w/v) PBSA (6.3 U/mL) were used as carbon sources, respectively. All strains exhibited the highest depolymerase activities between pH 6.0–8.0 and temperature 40–60°C. Partial nucleotides of the polyester depolymerase gene from strain S14, TF1 and APL3 were studied. We determined the amino acids making up the depolymerase enzymes had a highly conserved pentapeptide catalytic triad (Gly-His-Ser-Met-Gly), which has been shown to be part of the esterase-lipase superfamily (serine hydrolase).