Efficiency of Some Bacterial Strains in Potassium Release from Mica and Phosphate Solubilization under In Vitro Conditions

Phosphorus and potassium (K) are major essential macronutrients for biological growth and development. Application of beneficial microorganisms to soil is one approach to enhance crop growth. In this study, the ability of five bacterial strains, including four strains of Pseudomonas sp. (S10-3, S14-3, S19-1, and S21-1) and one strain of Azotobacter sp. SP16, to release K from muscovite and biotite was investigated. Furthermore, phosphate solubilization by these strains was measured when an insoluble source of P [Ca₃(PO₄)₂] was added to the medium. Among the bacterial strains, the highest average K release (73% higher than control) was observed with Pseudomonas sp. S14-3. The average amount of K released from biotite was 37% higher than that from muscovite in inoculated treatments. The enhanced release of mineral K might be attributed to the release of organic acids from the bacteria, a mechanism which plays a pivotal role in solubilizing phosphate from inorganic sources. The results confirmed the enhanced phosphate solubilization by the bacterial strains in the presence of muscovite. The highest P solubilizing activity (67% higher than control) was found in S21-1 and S14-3 strains. Concentrations of both K and P in the liquid phase were increased by increasing the time of experiment. X-ray diffraction analysis of muscovite specimens inoculated with S14-3 strain revealed a partial transformation of these minerals through the presence of 19.5 Å peak on the diffractogram of the magnesium-saturated sample. This may be due to the release of K from the interlayer space and subsequent filling with a number of bacterial metabolites. The findings of this research suggest K depletion from mica in the presence of bacteria, but further investigations are needed to clarify the mechanisms involved.     

Fermentation of n-Paraffins by Yeast

Screening test for obtaining microorganisms which produce l-amino acids or organic acids from n-paraffins were carried out. Fourteen strains of microorganisms which seemed to belong to the yeast showed strong ability to produce α-ketoglutaric acid. A representative strain of these microorganisms was identified as Candida lipolytica AJ 5004.

Optimal conditions for production of α-ketoglutarate using Candida lipolytica AJ 5004 were also studied. Under the condition thus obtained using a culture medium of 8 weight % of n-paraffins, the yeast accumulated 59% of α-ketoglutarate to the substrate added after three days culture.     

奶牛粪酸臭成分降解过程中的微生物群落变化

【目的】研究可降解成年泌乳奶牛粪中主要酸臭物的微生物群落的组成及动态变化。【方法】利用牛粪堆肥环境中的微生物进行了发酵优化、菌种驯化以及酸臭有机物降解规律的研究,结合r DNA高通量测序技术对有益微生物的组成及相对生物量进行了分析。【结果】实验发现,奶牛排泄物中的臭味来源主要为短链有机酸,堆肥自然环境中的微生物可以有效地对有机酸等污染物进行去除,经从低到高浓度的有机酸臭物(W/V,0.1%–0.2%)驯化发酵后,培养物中原核微生物以芽孢杆菌居多,而真核微生物主要由红曲霉及粉状毕赤酵母组成。【结论】进一步推测这几种微生物是耐受并降解有机酸臭物的优势微生物,可以应用于奶牛养殖过程中酸臭排泄物的生物控制。     

洋河浓香型白酒发酵过程酒醅微生物群落结构解析及其与有机酸合成的相关性

【目的】解析江苏洋河酒厂浓香型白酒窖内发酵过程酒醅微生物群落结构,建立酒醅微生物与主要有机酸合成的关联性。【方法】通过宏基因组测序获得白酒发酵过程中微生物群落结构变化规律,利用主成分分析和偏最小二乘回归分析寻找酒醅中影响主要有机酸合成的关键微生物。【结果】根据微生物组成结构变化和有机酸合成变化规律,可将白酒窖内发酵分为两个时期(0–14 d和15–60 d)。其中窖内发酵0–15 d与主要有机酸合成相关的微生物数量显著高于15–60 d的。窖内发酵过程与主要有机酸合成相关的微生物包括7个菌属,分别为乳杆菌属(Lactobacillus)、葡萄球菌属(Staphylococcus)、酵母属(Saccharomyces)、Naumovozyma、伊萨酵母属(Issatchenkia)、嗜冷芽孢杆菌属(Psychrobacillus)和根霉属(Rhizopus)。【结论】本研究识别了白酒窖内发酵过程中与主要有机酸合成相关的核心和关键微生物,可为阐明白酒窖内发酵产酸机理和保障白酒品质的稳定性奠定研究基础和理论依据。     

Effects of Mineral Structure and Microenvironment on K Release from Potassium Aluminosilicate Minerals by Cenococcum geophilum fr

K-bearing minerals with enormous reserve in the world, were not easy to provide the available K nutrient element for plant direct uptake because of the slow K release rate. With the potential potassium-solubilizing microorganisms, the slow K release rate from minerals would be improved significantly. In this work, Cenococcum geophilum Fr, one of the most common ectomycorrhizal fungi in boreal to temperate regions, was adopted to dissolve K-bearing minerals for K release. Five kinds of potassium aluminosilicate minerals were tested by bioleaching experiments in pure culture, including feldspar, nepheline, biotite, muscovite and illite. The available and unavailable potassium amounts in minerals before and after bioleaching were measured and compared with each other. The effect of mineral structure on the potassium solubilization efficiency by Cenococcum geophilum Fr. was discussed. Furthermore, the microenvironment formation between fungi and mineral surface to enhance the K release rate was investigated through detecting K, Al, Si concentrations and metabolites amounts (polysaccharide and organic acids) in microenvironment and external environment, respectively. Experimental results demonstrated that Cenococcum geophilum Fr. was a potential candidate of potassium solubilizing microorganisms, and both mineral structure and microenvironment have significant effects on the K release rate.     

Three New Lipases from Actinomycetes and Their Use in Organic Reactions

Three novel lipase-producing microorganisms have been isolated from 526 actinomycete strains by employing screening techniques on solid media. Time-course and scale-up of enzyme production were analyzed. The lipases, produced by microorganisms belonging to the Streptomyces genus, were tested in several reactions in organic medium using unnatural substrates. The lyophilized crude lipases are stable at least for 1 month at 4°C (100% recovered activity). The lipase activity per milliliter of cell culture broth was higher than described in the literature for other lipases from actinomycetes. The three selected lipases displayed better activity than commercial lipase from Candida rugosa in the resolution of chiral secondary alcohols. The lipase from S. halstedii also displayed very good activity in the synthesis of carbamates.     

Investigation of Microbes in the Rhizosphere of Selected Grasses for Rhizoremediation of Hydrocarbon-Contaminated Soils

Hydrocarbon-degrading microorganisms (HDM) associated with the rhizosphere of Paspalum vaginatum and Zoysia tenuifolia grown in bioremediated soil were isolated under controlled laboratory conditions. The isolation process was conducted at 30°C and 45°C to isolate mesophilic and thermotolerant microorganisms, respectively, under aerobic conditions. The isolated HDMs were identified using 16S rRNA gene sequencing and fatty acid methyl ester (FAME) analysis. Although differences in the genera of the isolated HDMs occurred between the two grasses, Arthrobacter spp and Bacillus spp were isolated from the rhizosphere of both plants. The efficiency of the isolated microorganisms in degrading a mixture of hydrocarbon compounds (HC) was also assessed. Among the bacterial isolates, Pseudomonas boreopolis was found to be the most effective HC degrader, while the only fungal isolate, Fusarium solani, demonstrated higher degradation rates than most of the bacterial isolates. A mixture of all the microbial isolates demonstrated a high degradation percent of HC. The isolated microorganisms thus appear to work synergistically to degrade efficiently all the tested organic compounds.     

Lithocholic Acid Side-chain Cleavage to Produce 17-Keto or 22-Aldehyde Steroids by Pseudomonas putida strain ST-491 Grown in the Presence of an Organic Solvent,Diphenyl Ether

We devised a method to screen for microorganisms capable of growing on bile acids in the presence of organic solvents and producing organic solvent-soluble derivatives. Pseudomonas putida biovar A strain ST-491 isolated in this study produced decarboxylated derivatives from the bile acids. Strain ST-491 grown on 0.5% lithocholic acid catabolized approximately 30% of the substrate as a carbon source, and transiently accumulated in the medium androsta-1,4-diene-3,17-dione in an amount of corresponding to 5% of the substrate added. When 20% (v/v) diphenyl ether was added to the medium, 60% of the substrate was converted to 17-keto steroids (androst-4-ene-3,17-dione-like steroid, androsta-1,4-diene-3,17-dione) or a 22-aldehyde steroid (pregna-1,4-dien-3-on-20-al). Amounts of the products were responsible for 45, 10, and 5% of the substrate, respectively. In the presence of the surfactant Triton X-100 instead of diphenyl ether, 40% of the substrate was converted exclusively to androsta-1,4-diene-3,17-dione.     

Isolation and investigation of anaerobic microorganisms involved in methanol transformation in an underground gas storage facility

High methanol and acetate concentrations (up to 12 and 14 g l⁻¹, respectively) were found in water samples collected at different objects of the North Stavropol underground gas storage facility (UGSF), and significant seasonal variations in the content of these compounds were revealed. The dominant anaerobic microorganisms isolated from these samples during the study belonged to acetogens, methanogens, and sulfate reducers. The results of 16S rRNA gene sequencing and analysis of the physiological properties showed that the isolates were close to the species of Eubacterium limosum, Sporomusa sphaeroides, Methanosarcina barkeri, Methanobacterium formicicum, and Desulfovibrio desulfuricans. The isolated organisms, except for Methanobacterium formicicum, were capable of methylotrophic growth. All strains were characterized by resistance to high methanol concentrations (up to 40–50 g l⁻¹). Their other energy substrate was hydrogen. The combination of the growth characteristics of these strains (pH, temperature, and salinity ranges) was shown to correspond to the ecological situation observed in the UGSF. The results of investigation of the isolated strains suggest that organic acids (acetate, butyrate) found in high concentrations in the initial samples are metabolic products of the revealed acetogens. Based on the established biological peculiarities of the isolated strains of methanogens, acetogens, and sulfate-reducing bacteria, these microorganisms may be considered as the main agents of anaerobic transformation of methanol and some other organic and inorganic compounds in UGSFs.     

Chemical and microbiological solubilization of silicates

The solubilization of silicates was investigated using kaolin and quartz sand as model substances. The mineral solubilization was studied in the concentration of solubilized Si and Al. The chemical leaching of the silicates was carried out using inorganic and organic acids as well as sodium hydroxide. The process was more effective in the alkine than in the acid pH range. In the acid medium, oxalic acid showed maximum acidity and a tendency to form complex structures, especially with aluminium, and was most effective in leaching. The microbiological influence on solubilization reactions was tested using a number of microorganisms among them acid, alkali and slime-forming species. The highest leaching activity was observed in the case of Thiobacillus thiooxidans, whereas the heterotrophic microorganisms (among them Bacillus mucilaginosus) did not exercise a sollubilizing effect on the silicates. X-ray phase analysis of leached kaolin samples did not show any differences from the non-leached mineral.     

Studies on Biosynthesis of Biotin by Microorganisms

During the course of the study on the production of biotin from desthiobiotin by microorganisms, the present authors have found that some strains of molds produced an unknown biotin-vitamer (BS-factor) from desthiobiotin. The present investigation was undertaken to clarify the characteristics of the unknown vitamer. The unknown vitamer produced from desthiobiotin was isolated in crystalline form from culture filtrate of Aspergillus oryzae. The compound isolated was identified as 4-methyl-5-(ω-carboxybutyl)-imidazolidone-2 by the physico-chemical procedures.

The biosynthesis of biotin-vitamers by resting cell system of Bacillus sphaericus was studied.

It was found that pimelic acid was essential substrate in biosynthesis of biotin-vitamers and that some amino acids and organic acids stimulated the biosynthesis of biotin-vitamers from pimelic acid. Alanine was found to be most effective. It was assumed that, in the presence of pimelic acid, some amino acids, especially alanine, and some organic acids play an important role in the biosynthesis of biotin-vitamers.

The main component of the biotin-vitamers synthesized by the resting cell system was identified as desthiobiotin. The existence of a small amount of unknown biotin-vitamer, an avidin-uncombinable substance, which was assumed to be 7-keto-8-amino-pelargonic acid, was also observed. True biotin was hardly observed in any conditions tested.     

Ammonia removal from livestock wastewater by ammonia-assimilating microorganisms immobilized in polyvinyl alcohol

We isolated ammonia-assimilating microorganisms from the livestock manure treatment systems and evaluated their ammonia-assimilating ability. Many isolates utilized ammonia at high rates when they were purely cultivated in a nitrogen-limited medium to which sterilized lagoon extract had been added. Some isolates that were immobilized in polyvinyl alcohol (PVA) utilized ammonia present in the media containing viable lagoon microorganisms. Staining with 4′,6′-diamidino-2-phenylindole (DAPI) indicated that the immobilized high ammonia-assimilating isolates grew dominantly within the PVA beads. High ammonia-assimilating isolates in the mixed culture containing viable lagoon microorganisms were identified as Pseudomonas spp. and member of Rhizobiaceae species by partial sequencing of the 16S ribosomal DNA.     

2-Deceno-δ-lactone-producing Fungi,Strains of Fusarium solani,Isolated by Using a Medium Containing Decano-δ-lactone as the Sole Carbon Source

A screening procedure for microorganisms which have an ability to produce a desired compound as their secondary metabolite is first proposed. In some cases, the target microorganisms can be expected to grow and be enriched in a medium containing the desired compound, namely one of the secondary metabolites of the microorganisms, as the sole source of carbon, degrading and assimilating the compound to the primary metabolites. This approach was applied to isolate alkano-δ-lactones producing fungi by using a medium containing alkano-δ-lactones as the sole source of carbon. We isolated Fusarium solani and Trichoderma viride that had the ability to biosynthesize 2-deceno-δ-lactone (massoialactone) and 2,4-decadieno-δ-lactone(6-pentyl-δ-pyrone), respectively, in a glucose medium.     

Presence of endophytic diazotrophs in sugarcane juice

Summary In this work we investigated the diazotrophs present in the juice of different varieties of sugarcane. Samples of the aerial part of sugarcane, between 3 and 5 months old, were assessed for the presence of diazotrophs. The isolated nitrogen-fixing microorganisms were identified as Gluconacetobacter diazotrophicus, Herbaspirillum seropedicae and Azospirillum brasilense. These microorganisms use organic acids found in the sugarcane juice as an energy source. They also use sugars, such as free glucose, that are present in all the organs of the aerial part. The distribution of endophytic microorganisms in sugarcane depends on the different organic acid and sugar concentrations present during development and growth. We concluded that in more mature regions of the sugarcane stem Gluconacetobacter diazotrophicus grows more abundantly than Herbaspirillum seropedicae or Azospirillum brasilense.     

Studies on the Utilization of Hydrocarbons by Microorganisms

It is well known that a good many microorganisms can utilize hydrocarbons as a sole carbon source. We have as yt little information, however, as to what kinds of intermediate or end products are produced by the microbial dissimilation of hydrocarbons. Above all, the formation of amino acids from hydrocarbons has not been reported. We have isolated many strains of microorganisms from soil samples by selective culture techniques in a medium containing kerosene and mineral salts, in order to examine if the products of economical value such as amino acids, organic acids, fatty acids, steroids, nucleic acids and their related compounds, can be produced by those microorganisms. Shaking cultures of those microorganisms were carried out in the medium containing 3.5% kerosene, 3.5% liquid paraffin and mineral salts (K₂HPO₄ 0.25%; MgSO₄·7H₂O 0.1%; NH₄MO₈ 0.5% or urea 0.3%); we found that 55 strains out of 127 strains tested produced amino acids in their broths, after 4 days cultivation at 26.5°C. The kinds and maximum yields of amino acids were as follows: alanine 50mg/l, aspartic acid 23mg/l, glutamic acid 485 mg/1, glycine 8.5mg/l, isoleucine 15 mg/1, lysine 23.9 mg/1, and valine 38 mg/1.     

Ethanol production by Saccharomyces cerevisiae in biofilm reactors

Biofilms are natural forms of cell immobilization in which microorganisms attach to solid supports. At ISU, we have developed plastic composite-supports (PCS) (agricultural material (soybean hulls or oat hulls), complex nutrients, and polypropylene) which stimulate biofilm formation and which supply nutrients to the attached microorganisms. Various PCS blends were initially evaluated in repeated-batch culture-tube fermentation with Saccharomyces cerevisiae (ATCC 24859) in low organic nitrogen medium. The selected PCS (40% soybean hull, 5% soybean flour, 5% yeast extract-salt and 50% polypropylene) was then used in continuous and repeated-batch fermentation in various media containing lowered nitrogen content with selected PCS. During continuous fermentation, S. cerevisiae demonstrated two to 10 times higher ethanol production in PCS bioreactors than polypropylene-alone support (PPS) control. S. cerevisiae produced 30 g L⁻¹ ethanol on PCS with ammonium sulfate medium in repeated batch fermentation, whereas PPS-control produced 5 g L⁻¹ ethanol. Overall, increased productivity in low cost medium can be achieved beyond conventional fermentations using this novel bioreactor design. Received 20 May 1997/ Accepted in revised form 29 August 1997     

Organic Acid Secretion and Phosphate Solubilizing Efficiency of Pseudomonas sp. PSB12: Effects of Phosphorus Forms and Carbon Sources

Secretion of organic acids is an important mechanism for phosphate solubilizing bacteria (PSB) to dissolve insoluble phosphorus in soil. However, the composition of organic acids produced by PSB in the presence of different phosphorus compounds is poorly known, and little is known about the ability of PSB to degrade pollutants in sediment. In this study, we isolated a strain Pseudomonas sp. PSB12 from the sediment of the Qihe River. PSB12 had maximum phosphate solubilization index (SI) of 3.86 on Pikovskaya's agar medium. The phosphate solubilizing activity was associated with the release of organic acids produced from glucose, while the composition of organic acids produced by PSB12 was dependent on the phosphorus forms. When initial soluble phosphorus was insufficient (in MP1 and MP2 media), gluconic acid was the predominant organic acid. In contrast, formic acid, butyrate, and propanedioic acid were the main organic acids produced when only soluble phosphorus (MP3) was supplied. RT-PCR indicated that the expression of glucose dehydrogenase gene (gcd) of PSB12 was two- to four-fold higher in MP1 than in MP3. PSB12 also possessed the phenol hydroxylase gene (phe) suggesting that phenol could be used as the carbon source to dissolve insoluble phosphorus. PSB12 is a potential candidate for in situ bioremediation and for promoting plant growth in soil contaminated by phenol with low levels of soluble phosphorus.     

南极乔治王岛冰锥洞微生物培养探索

【目的】南极洲具备独特的环境和相对的生物地理隔离,南极洲各类生境中蕴藏了大量尚未培养和难培养的微生物,也是新颖微生物物种的重要来源之一。本研究以南极冰锥洞这类特殊生境为研究对象,通过培养条件的多样化提升南极微生物的培养率和多样性,揭示南极冰锥洞可培养微生物类群多样性,为该环境可培养微生物功能研究奠定基础,也为南极极端环境未培养微生物的培养方法提供借鉴。【方法】通过采用不同培养基添加复苏促进因子(resuscitation promoting factor, Rpf)的方式,提高南极柯林斯冰盖冰锥洞生境中微生物的可培养率,探究该生境中微生物的多样性。采用4种不同营养水平的培养基,平行添加Rpf进行菌株培养,经分离纯化与16S rRNA基因鉴定,分析冰锥洞可培养微生物的多样性及培养条件对多样性的影响。【结果】本研究共分离培养细菌407株,涵盖5个门、18个科、29个属,其中：放线菌门(Actinomycetota)为优势门,占72.73%;微杆菌科(Microbacteriaceae)为优势科,占69.78%;Lacisediminihabitans属为优势属,占45.70%。从培养基效果...     

Phosphorus Recovery from Phosphate Rocks Using Phosphate-Solubilizing Bacteria

In this paper, we are presenting a biological process to recover phosphorus by solubilizing low-grade phosphate rocks. To this end, the efficiency of different phosphate-solubilizing microorganism (PSM) species for solubilizing P from phosphate rocks using both pure cultures and associations. Nutritional conditions, phosphate rock concentrations, and reactor designs were tested. The genus Bacillus, especially Bacillus megaterium (ATCC 14581), was found to be the most promising PSM for solubilizing P. Production of organic acids and acidic pH values were shown to be directly related to P solubilizing. However, associations between tested microorganisms did not significantly enhance process efficiency. We conclude that nutritional factors of the medium are important to solubilization, and lower phosphate rock concentrations lead to better solubilization. The Air Lift reactor was promising for B. megaterium (ATCC 14581), but adaptations are needed for further tests.     

Isolation and characterization of biogenic amine-producing bacteria in fermented soybean pastes

Biogenic amines (BAs) are produced primarily by microorganisms found in fermented foods and are often implicated in food poisoning. BA-producing bacteria found in fermented soybean pastes were isolated and characterized using a decarboxylating medium and multiplex PCR analysis. Two BA-producing bacteria were isolated from traditional soybean pastes: one was a histamine-producing Clostridium strain, and the other was a tyramine-producing Pseudomonas strain. The Clostridium strain was determined to be a potent histamine producer among the cultures tested. Synthesis of tyramine by Pseudomonas sp. T1 was observed for the first time in this study.