Study on the mechanism of biochar affecting the effectiveness of phosphate solubilizing bacteria.

Low phosphorus utilization and phosphorus fertilizer pollution are serious issues primarily affecting soil health. To investigate the effects of biochar on the growth, phosphorus solubilization, and metabolites of phosphorus-solubilizing bacteria (PSB), rice husk biochar (RH) and rice straw biochar (RS) were incubated with Bacillus megatherium (BM1) and Bacillus mucilaginosus (BM2), respectively. The highest phosphorus solubilization was observed in BM2 following the addition of RS. The dissolved amount of phosphorus was 244.99 mg/L, which was 43.86% higher than that of the control group. Hence, biochar can improve the phosphorus solubilization capacity of PSB by affecting the organic acid and polysaccharide contents, and phosphatase activity secreted by the PSB, as the porous structure and surface characteristics of biochar ensured the adsorption of PSB. This study can help improve the functional activity of PSB and provide basis for improving the utilization of soil phosphorus, which in turn, aid in the development of biochar-based microbial fertilizers.

Degradation of alkane hydrocarbons by Priestia megaterium ZS16 and sediments consortia with special reference to toxicity and oxidative stress induced by the sediments in the vicinity of an oil refinery.

Petroleum hydrocarbon is a critical ecological issue with impact on ecosystems through bioaccumulation. It poses significant risks to human health. Due to the extent of alkane hydrocarbon pollution in some environments, biosurfactants are considered as a new multifunctional technology for the efficient removal of petroleum-based contaminants. To this end, Yamuna river sediments were collected at different sites in the vicinity of Mathura oil refinery, UP (India). They were analysed by atomic absorption spectrophotometry and gas chromatography-mass spectrometry (GC-MS) for heavy metals and organic pollutants. Heptadecane, nonadecane, oleic acid ester and phthalic acid were detected. In total 107 bacteria were isolated from the sediments and screened for biosurfactant production. The most efficient biosurfactant producing strain was tested for its capability to degrade hexadecane efficiently at different time intervals (0 h, 7 d, 14 d and 21 d). FT-IR analysis defined the biosurfactant as lipopeptide. 16S rRNA gene sequencing identified the bacterium as Priestia megaterium. The strain lacks resistance to common antibiotics thus making it an important candidate for remediation. The microbial consortia present in the sediments were also investigated for their capability to degrade C16, C17 and C18 alkane hydrocarbons. By using gas chromatography-mass spectrophotometry the metabolites were identified as 1-docosanol, dodecanoic acid, 7-hexadecenal, (Z)-, hexadecanoic acid, docosanoic acid, 1-hexacosanal, 9-octadecenoic acid, 3-octanone, Z,Z-6,28-heptatriactontadien-2-one, heptacosyl pentafluoropropionate, 1,30-triacontanediol and decyl octadecyl ester. Oxidative stress in Vigna radiata L. roots was observed by using Confocal Laser Scanning Microscopy. A strong reduction in seed germination and radicle and plumule length was observed when Vigna radiata L. was treated with different concentrations of sediment extracts, possibly due to the toxic effects of the pollutants in the river sediments. Thus, this study is significant since it considers the toxicological effects of hydrocarbons and to degrade them in an environmentally friendly manner.

Engineering precursor and co-factor supply to enhance D-pantothenic acid production in Bacillus megaterium.

High-yielding chemical and chemo-enzymatic methods of D-pantothenic acid (DPA) synthesis are limited by using poisonous chemicals and DL-pantolactone racemic mixture formation. Alternatively, the safe microbial fermentative route of DPA production was found promising but suffered from low productivity and precursor supplementation. In this study, Bacillus megaterium was metabolically engineered to produce DPA without precursor supplementation. In order to provide a higher supply of precursor D-pantoic acid, key genes involved in its synthesis are overexpressed, resulting strain was produced 0.53 ± 0.08 g/L DPA was attained in shake flasks. Cofactor CH2-THF was found to be vital for DPA biosynthesis and was regenerated through the serine-glycine degradation pathway. Enhanced supply of another precursor, ß-alanine was achieved by codon optimization and dosing of the limiting L-asparate-1-decarboxylase (ADC). Co-expression of Pantoate-ß-alanine ligase, ADC, phosphoenolpyruvate carboxylase, aspartate aminotransferase and aspartate ammonia-lyase enhanced DPA concentration to 2.56 ± 0.05 g/L at shake flasks level. Fed-batch fermentation in a bioreactor with and without the supplementation of ß-alanine increased DPA concentration to 19.52 ± 0.26 and 4.78 ± 0.53 g/L, respectively. This present study successfully demonstrated a rational approach combining precursor supply engineering with cofactor regeneration for the enhancement of DPA titer in recombinant B. megaterium.

Bacillus megaterium-induced biocorrosion on mild steel and the effect of Artemisia pallens methanolic extract as a natural corrosion inhibitor.

Methanolic extract of Artemisia pallens (MEAP) (Asteraceae) was explored as greenbiocorrosion inhibitor for mild steel 1010 in 1.5% sodium chloride environment. Bacillus megaterium SKR7 induces the development of biofilm on the metal surface and forms the pitting corrosion. MEAP was showed (25 ppm) optimum inhibition effect of biocorrosion and further corrosion rate was highly reduced (0.3335 mm/year) than the control system (0.009 mm/year). The electrochemical study has supported the results with a higher value of total resistance (34 Ω cm2) when compared to control systems. It reveals the formation of a protective layer on the metal surface and reduces the adsorption of biofilm. This was due to the antimicrobial effect of MEAP. Overall, the results recognized that MEAP used as a green corrosion inhibitor for MS 1010 with 83% inhibition efficiency.

Responses of soil enzyme activities and plant growth in a eucalyptus seedling plantation amended with bacterial fertilizers.

Plant growth-promoting rhizobacteria (PGPR) are widely used to improve plant nutrient uptake and assimilation and soil physicochemical properties. We investigated the effects of bacterial (Bacillus megaterium strain DU07) fertilizer applications in a eucalyptus (clone DH32-29) plantation in Guangxi, China in February 2011. We used two types of organic matter, i.e., fermented tapioca residue ("FTR") and filtered sludge from a sugar factory ("FS"). The following treatments were evaluated: (1) no PGPR and no organic matter applied (control), (2) 3 × 109 CFU/g (colony forming unit per gram) PGPR plus FS (bacterial fertilizer 1, hereafter referred to as BF1), (3) 4 × 109 CFU/g plus FS (BF2), (4) 9 × 109 CFU/g plus FS (BF3), (5) 9 × 109 CFU/g broth plus FTR (BF4). Soil and plant samples were collected 3 months (M3) and 6 months (M6) after the seedlings were planted. In general, bacterial fertilizer amendments significantly increased plant foliar total nitrogen (TN) and soil catalase activity in the short term (month 3, M3); whereas, it significantly increased foliar TN, chlorophyll concentration (Chl-ab), proline; plant height, diameter, and volume of timber; and soil urease activity, STN, and available N (Avail N) concentrations in the long term (month 6, M6). Redundancy analysis showed that soil available phosphorus was significantly positively correlated with plant growth in M3, and soil Avail N was negatively correlated with plant growth in M6. In M3, soil catalase was more closely correlated with plant parameters than other enzyme activities and soil nutrients, and in M6, soil urease, polyphenol oxidase, and peroxidase were more closely correlated with plant parameters than other environmental factors and soil enzyme activities. PCA results showed that soil enzyme activities were significantly improved under all treatments relative to the control. Hence, photosynthesis, plant growth, and soil N retention were positively affected by bacterial fertilizer in M6, and bacterial fertilizer applications had positive and significant influence on soil enzyme activities during the trial period. Thus, bacterial fertilizer is attractive for use as an environmentally friendly fertilizer in Eucalyptus plantations following proper field evaluation.

Polyhydroxyalkanoate (PHA) biosynthesis from directly valorized ragi husk and sesame oil cake by Bacillus megaterium strain Ti3: Statistical optimization and characterization.

Polyhydroxyalkanoates (PHAs) signify the most promising biological substitute to petrochemical plastics. Renewable and inexpensive agro-industrial by-products can be used as potent fermentation feedstocks for sustainable PHA biosynthesis. This study aimed at using a wild type B. megaterium strain Ti3 innate hydrolytic enzyme/s for eco-friendly valorization of 16 lignocellulosic agrowastes to PHA without pretreatments. Initial hydrolytic screening PHA concentration of (0.04-0.17 g/L), highlighted the strain's metabolic versatility. Pareto ranking of Taguchi orthogonal array (TOA) established ragi husk (RH), sesame oil cake (SOC) and KH2PO4 as the most influential factors (p < 0.05). The optimized and validated Response surface methodology (RSM) model (R2, 0.979; desirability, 1) resulted in 3.8 and 3.6 fold increased PHA production, 4.3 and 3.25 fold increased PHA productivity. A positive correlation (r2, 0.5-0.97) was observed amid the producer innate hydrolytic enzymes (lipase, amylase and cellulase) and PHA production. The PHA was characterized by 1H and 13C NMR, GPC, TGA. The polymer was identified as a scl-mcl copolyester with 92% 3HB (3-hydroxybutyrate) and 8% 3HHp (3-hydroxyheptanoate) monomers by NMR. This the first report on B. megaterium self-enzyme reliant non-food agrowastes bioconversion to PHA with 3HHp (3-hydroxyheptanoate) monomers excluding precursor addition, commercial enzymes, pure carbon and nitrogen sources.

Maytenin Plays a Special Role in the Regulation of the Endophytic Bacillus megaterium in Peritassa campestris Adventitious Roots.

Peritassa campestris (Celastraceae) root bark accumulates potent antitumor quinonemethide triterpenes (QMTs). When grown in their natural habitat, plants of the family Celastraceae produce different QMTs such as celastrol (3) and pristimerin (4). However, when they are inserted in in vitro culture systems, they accumulate maytenin (1) as the main compound. Recently, Bacillus megaterium was detected as an endophytic microorganism (EM) living inside P. campestris roots cultured in vitro. We hypothesized that compound (1) controls EM growth more efficiently, and that the presence of EMs in the root culture causes compound (1) to accumulate. For the first time, this work has explored plant-microorganism interaction in a species of the family Celastraceae by co-culture with an EM. Live endophytic bacteria were used, and QMT accumulation in P. campestris adventitious roots was our main focus. The antimicrobial activity of the main QMTs against endophytic B. megaterium was also evaluated. Our results showed that compound (1) and maytenol (5) were more effective than their precursors QMTs (3) and (4) in controlling the EM. Co-culture of B. megaterium with roots significantly reduced bacterial growth whereas root development remained unaffected. Compound (1) production was 24 times higher after 48 hr in the presence of the highest B. megaterium concentration as compared to the control. Therefore, P. campestris adventitious roots affect the development of the endophyte B. megaterium through production of QMTs, which in turn can modulate production of compound (1).

Integrated Effects of Co-Inoculation with Phosphate-Solubilizing Bacteria and N2-Fixing Bacteria on Microbial Population and Soil Amendment Under C Deficiency.

The inoculation of beneficial microorganisms to improve plant growth and soil properties is a promising strategy in the soil amendment. However, the effects of co-inoculation with phosphate-solubilizing bacteria (PSB) and N2-fixing bacteria (NFB) on the soil properties of typical C-deficient soil remain unclear. Based on a controlled experiment and a pot experiment, we examined the effects of PSB (M: Bacillus megaterium and F: Pseudomonas fluorescens), NFB (C: Azotobacter chroococcum and B: Azospirillum brasilence), and combined PSB and NFB treatments on C, N, P availability, and enzyme activities in sterilized soil, as well as the growth of Cyclocarya Paliurus seedlings grow in unsterilized soil. During a 60-day culture, prominent increases in soil inorganic N and available P contents were detected after bacteria additions. Three patterns were observed for different additions according to the dynamic bacterial growth. Synergistic effects between NFB and PSB were obvious, co-inoculations with NFB enhanced the accumulation of available P. However, decreases in soil available P and N were observed on the 60th day, which was induced by the decreases in bacterial quantities under C deficiency. Besides, co-inoculations with PSB and NFB resulted in greater performance in plant growth promotion. Aimed at amending soil with a C supply shortage, combined PSB and NFB treatments are more appropriate for practical fertilization at intervals of 30-45 days. The results demonstrate that co-inoculations could have synergistic interactions during culture and application, which may help with understanding the possible mechanism of soil amendment driven by microorganisms under C deficiency, thereby providing an alternative option for amending such soil.

Biodegradation of hydrolyzed polyacrylamide by a Bacillus megaterium strain SZK-5: Functional enzymes and antioxidant defense mechanism.

Hydrolyzed polyacrylamide (HPAM) is the most widely used water-soluble linear polymer with high molecular weight in polymer flooding. Microbiological degradation is an environment-friendly and effective method of treating HPAM-containing oilfield produced water. In this study, a strain SZK-5 that could degrade HPAM was isolated from soil contaminated by oilfield produced water. Based on morphological, biochemical characteristics and 16S rDNA sequence homology analysis, the strain was identified as Bacillus megaterium. The biodegradation capability of strain SZK-5 was determined by incubation in a mineral salt medium (MSM) containing HPAM under different environmental conditions, showing 55.93% of the HPAM removed after 7 d of incubation under the optimum conditions ((NH4)2SO4 = 1667.9 mg L-1, temperature = 24.05 °C and pH = 8.19). Cytochrome P450 (CYP) and urease (URE) played significant roles in biological carbon and nitrogen removal, respectively. The strain SZK-5 could resist the damages caused by oxidative stress given by crude oil and HPAM. To our knowledge, this is the first report about the biodegradation of HPAM by B. megaterium. These results suggest that strain SZK-5 might be a new auxiliary microbiological resource for the biodegradation of HPAM residue in wastewater and soil.

Phosphorus Solubilization by Bacillus Species.

Microbial solubilization applies the natural ability of a microorganism to liberate phosphorus from unavailable structures. The main mechanism recognized to be responsible for the solubilization of phosphorus is the production of different types of organic acids. Three kinds of Bacillus species and three types of raw materials (poultry bones, fish bones, and ash) were tested for solubilization. The following parameters were compared for all discussed cases: pH, specific growth rate, solubilization factor, released phosphorus concentration, and total and individual concentration of organic acids. Utilization of ash brought about the highest specific and maximum specific growth rates. A decrease in pH was observed in most of the discussed cases with the exception of fish bones. At the same time, fish bones had the highest concentration of released P2O5 and the highest total concentration of produced organic acids (gluconic, lactic, acetic, succinic, and propionic) in all discussed cases. The tested Bacillus species produced the mentioned acids with the exception of B. megaterium, where propionic acid was not present. The lactic and acetic acids were those produced in the highest amount. The kind of raw materials and type of Bacillus species used in solubilization had a strong influence on the kind of organic acids that were detected in the broth culture and its total concentration, which had a direct influence on the amount of released phosphorus. The combination of Bacillus megaterium with the fish bones at 5 g/L is proposed as the pair that gives the highest concentration of released phosphorus (483 ± 5 mg/L).

Biosurfactant-biopolymer driven microbial enhanced oil recovery (MEOR) and its optimization by an ANN-GA hybrid technique.

A lipopeptide biosurfactant produced by marine Bacillus megaterium and a biopolymer produced by thermophilic Bacillus licheniformis were tested for their application potential in the enhanced oil recovery. The crude biosurfactant obtained after acid precipitation effectively reduced the surface tension of deionized water from 70.5 to 28.25mN/m and the interfacial tension between lube oil and water from 18.6 to 1.5mN/m at a concentration of 250mgL-1. The biosurfactant exhibited a maximum emulsification activity (E24) of 81.66% against lube oil. The lipopeptide micelles were stabilized by addition of Ca2+ ions to the biosurfactant solution. The oil recovery efficiency of Ca2+ conditioned lipopeptide solution from a sand-packed column was optimized by using artificial neural network (ANN) modelling coupled with genetic algorithm (GA) optimization. Three important parameters namely lipopeptide concentration, Ca2+ concentration and solution pH were considered for optimization studies. In order to further improve the recovery efficiency, a water soluble biopolymer produced by Bacillus licheniformis was used as a flooding agent after biosurfactant incubation. Upon ANN-GA optimization, 45% tertiary oil recovery was achieved, when biopolymer at a concentration of 3gL-1 was used as a flooding agent. Oil recovery was only 29% at optimal conditions predicted by ANN-GA, when only water was used as flooding solution. The important characteristics of biopolymers such as its viscosity, pore plugging capabilities and bio-cementing ability have also been tested. Thus, as a result of biosurfactant incubation and biopolymer flooding under the optimal process conditions, a maximum oil recovery of 45% was achieved. Therefore, this study is novel, timely and interesting for it showed the combined influence of biosurfactant and biopolymer on solubilisation and mobilization of oil from the soil.

Role of microbial inoculation and industrial by-product phosphogypsum in growth and nutrient uptake of maize (Zea mays L.) grown in calcareous soil.

BACKGROUND: Alkaline soils with high calcium carbonate and low organic matter are deficient in plant nutrient availability. Use of organic and bio-fertilizers has been suggested to improve their properties. Therefore, a greenhouse experiment was conducted to evaluate the integrative role of phosphogypsum (PG; added at 0.0, 10, 30, and 50 g PG kg-1 ), cow manure (CM; added at 50 g kg-1 ) and mixed microbial inoculation (Incl.; Azotobacter chroococcum, and phosphate-solubilizing bacteria Bacillus megaterium var. phosphaticum and Pseudomonas fluorescens) on growth and nutrients (N, P, K, Fe, Mn, Zn and Cu) uptake of maize (Zea mays L.) in calcareous soil. Treatment effects on soil chemical and biological properties and the Cd and Pb availability to maize plants were also investigated. RESULTS: Applying PG decreased soil pH. The soil available P increased when soil was inoculated and/or treated with CM, especially with PG. The total microbial count and dehydrogenase activity were enhanced with PG+CM+Incl. TREATMENTS: Inoculated soils treated with PG showed significant increases in NPK uptake and maize plant growth. However, the most investigated treatments showed significant decreases in shoot micronutrients. Cd and Pb were not detected in maize shoots. CONCLUSIONS: Applying PG with microbial inoculation improved macronutrient uptake and plant growth. © 2017 Society of Chemical Industry.

[Efficiency and Mechanism of Purifying High Iron-Manganese from Ground Water in the Cold Villages and Towns Based on The Coupling of Rice Husk and Iron-Manganese Oxidizing Bacteria].

Aiming at the special geo-climatic conditions and typical problems constraining the development of villages and towns, making villages and towns have the following characteristics:green and low-carbon, energy saving and environmental friendly, the coupling process of adsorption by carbonized rice husk particles and biological oxidation using Bacillus megaterium bacteria was developed for purifying high iron-manganese from groundwater in the cold villages and towns. The quick start-up method of biological filter at low temperature was studied. Based on the contact oxidation and biological method, iron removal efficiency and reaction rate in different filtration layers under different filtration conditions were comparatively analyzed, and iron removal mechanism of biological filter was determined. Filter materials and the solid samples in backwash water at different stages were characterized and the manganese removal mechanism of biological filter was analyzed through SEM, FTIR, XPS, Raman spectra and EPR. The results showed that the active biological membrane reached mature and stable only after 15 d with the operation mode of bacteria liquid cycle and low filtration rate in the filter column. The effluent concentration met the requirements of drinking water quality (GB 5749-2006) in the stable operation process. Fe, Mn and bacteria were lower than 0.3 mg·L-1, 0.1 mg·L-1, 100 CFU·mL-1, respectively. Iron removal mechanism relied mainly on the physical and chemical effects, supplemented by biological function. During the biofilm culturing stage and initial stable operation stage of filter column, manganese removal relied mainly on the biological effect. The physical chemistry was preferred at later stable operation stage. The study provides technical support for the applications of treating collectively Fe2+ and Mn2+ in groundwater.

Isolation and identification of Bacillus megaterium YB3 from an effluent contaminated site efficiently degrades pyrene.

Industrial effluents contaminated sites may serve as repositories of ecologically adapted efficient pyrene degrading bacteria. In the present study, six bacterial isolates from industrial effluents were purified using serial enrichment technique and their pyrene degrading potential on pyrene supplemented mineral salt medium was assessed. 16S rRNA sequence analysis showed that they belong to four bacterial genera, namely Acinetobacter, Bacillus, Microbacterium, and Ochrobactrum. Among these isolates, Bacillus megaterium YB3 showed considerably good growth and was further evaluated for its pyrene-degrading efficiency. B. megaterium YB3 could degrade 72.44% of 500 mg L(-1) pyrene within 7 days. GC-MS analysis of ethyl acetate extracted fractions detected two relatively less toxic metabolic intermediates of the pyrene degradation pathway. B. megaterium YB3 also tested positive for catechol 1, 2-dioxygenase and aromatic-ring-hydroxylating dioxygenase indole-indigo conversion assays. Considering the ability and efficiency of B. megaterium YB3 to degrade high pyrene content, the strain can be used as a tool to develop bioremediation technologies for the effective biodegradation of pyrene and possibly other PAHs in the environment.

Rock inhabiting potassium solubilizing bacteria from Kerala, India: characterization and possibility in chemical K fertilizer substitution.

The role of rock inhabiting bacteria in potassium (K) solubilization from feldspar and their application in crop nutrition through substitution of fertilizer K was explored through the isolation of 36 different bacteria from rocks of a major hill station at Ponmudi in Thiruvananthapuram, Kerala, India. A comprehensive characterization of K solubilization from feldspar was achieved with these isolates which indicated that the K solubilizing efficiency increases with decrease in pH and increase in viscosity and viable cell count. Based on the level of K solubilization, two potent isolates were selected and identified as Bacillus subtilis ANctcri3 and Bacillus megaterium ANctcri7. Exopolysaccharide production, scanning electron microscopic and fourier transform infrared spectroscopic studies with these efficient strains conclusively depicted the role of low pH, increase in viscosity, and bacterial attachment in K solubilization. They were also found to be efficient in phosphorus (P) solubilization, indole acetic acid production as well as tolerant to wide range of physiological conditions. Moreover, the applicability of K containing rock powder as a carrier for K solubilizing bacteria was demonstrated. A field level evaluation on the yield of a high K demanding tuberous vegetable crop, elephant foot yam (Amorphophallus paeoniifolius (dennst.) nicolson) established the possibility of substituting chemical K fertilizer with these biofertilizer candidates successfully.

Effects of interactions of auxin-producing bacteria and bacterial-feeding nematodes on regulation of peanut growths.

The influences of an IAA (indole-3-acetic acid)-producing bacterium (Bacillus megaterium) and two bacterial-feeding nematodes (Cephalobus sp. or Mesorhabditis sp.) on the growth of peanut (Arachis hypogaea L. cv. Haihua 1) after various durations of time were investigated in natural soils. The addition of bacteria and nematodes and incubation time all significantly affected plant growth, plant root growth, plant nutrient concentrations, soil nutrient concentrations, soil microorganisms and soil auxin concentration. The addition of nematodes caused greater increases in these indices than those of bacteria, while the addition of the combination of bacteria and nematodes caused further increases. After 42-day growth, the increases in soil respiration differed between the additions of two kinds of nematodes because of differences in their life strategies. The effects of the bacteria and nematodes on the nutrient and hormone concentrations were responsible for the increases in plant growth. These results indicate the potential for promoting plant growth via the addition of nematodes and bacteria to soil.

Dissolving mechanism of strain P17 on insoluble phosphorus of yellow-brown soil.

Strain P17 was a bacterial strain identified as Bacillus megaterium isolated from ground accumulating phosphate rock powder. The fermentation broth of strain P17 and the yellow-brown soil from Nanjing Agricultural University garden were collected to conduct this study. The simulation of fixed insoluble phosphorous forms after applying calcium superphosphate into yellow-brown soil was performed in pots, while available P and total P of soil were extremely positive correlative with those of groundwater. Then the dissolving effect of strain P17 on insoluble P of yellow-brown soil was studied. Results showed that Bacillus megaterium strain P17 had notable solubilizing effect on insoluble phosphates formed when too much water-soluble phosphorous fertilizer used. During 100 days after inoculation, strain P17 was dominant. Until the 120th day, compared with water addition, available P of strain P17 inoculation treated soil increased by 3 times with calcium superphosphate addition. Besides available P, pH, activity of acid and alkaline phosphatase and population of P-solubilizing microbes were detected respectively. P-solubilizing mechanism of P-solubilizing bacteria strain P17 seems to be a synergetic effect of pH decrease, organic acids, phosphatase, etc.

Platinum and rhenium extraction from a spent refinery catalyst using Bacillus megaterium as a cyanogenic bacterium: statistical modeling and process optimization.

The present study evaluated the potential of Bacillus megaterium as a cyanogenic bacterium to produce cyanide for solubilization of platinum and rhenium from a spent refinery catalyst. Response surface methodology was applied to study the effects and interaction between two main effective parameters including initial glycine concentration and pulp density. Maximum Pt and Re recovery was obtained 15.7% and 98%, respectively, under optimum conditions of 12.8 g/l initial glycine concentration and 4% (w/v) pulp density after 7 days. Increasing the free cyanide concentration to 3.6 mg/l, varying the pH from 6.7 to 9, and increasing the dissolved oxygen from 2 to 5mg/l demonstrated the growth characteristics of B. megaterium during bioleaching process. The modified shrinking core model was used to determine the rate limiting step of the process. It was found that diffusion through the product layer is the rate controlling step.

Dissolving mechanism of strain P17 on insoluble phosphorus of yellow-brown soil

Strain P17 was a bacterial strain identified as Bacillus megaterium isolated from ground accumulating phosphate rock powder. The fermentation broth of strain P17 and the yellow-brown soil from Nanjing Agricultural University garden were collected to conduct this study. The simulation of fixed insoluble phosphorous forms after applying calcium superphosphate into yellow-brown soil was performed in pots, while available P and total P of soil were extremely positive correlative with those of groundwater. Then the dissolving effect of strain P17 on insoluble P of yellow-brown soil was studied. Results showed that Bacillus megaterium strain P17 had notable solubilizing effect on insoluble phosphates formed when too much water-soluble phosphorous fertilizer used. During 100 days after inoculation, strain P17 was dominant. Until the 120th day, compared with water addition, available P of strain P17 inoculation treated soil increased by 3 times with calcium superphosphate addition. Besides available P, pH, activity of acid and alkaline phosphatase and population of P-solubilizing microbes were detected respectively. P-solubilizing mechanism of P-solubilizing bacteria strain P17 seems to be a synergetic effect of pH decrease, organic acids, phosphatase, etc.

Development of a two-step cultivation strategy for the production of vitamin B12 by Bacillus megaterium.

BACKGROUND: Vitamin B12 is a fascinating molecule which acts as a co-factor in the metabolism of many organisms, especially affecting DNA synthesis and regulation, fatty acid synthesis and energy production. The synthesis of vitamin B12 is limited to a few of bacteria and archaea. Therefore, industrial microbial fermentation is used to meet annual demands worldwide of vitamin B12 and as an alternative method to the chemical synthesis which requires at least 60 steps that is uneconomical. Bacillus megaterium is one of vitamin B12 producers and an ideal host for many biotechnology applications and being one of the best tools for the industrial production of several enzymes. Therefore, a two-step optimization strategy was established to produce high yield of vitamin B12 by B. megaterium through the provision of the production requirements and the suitable conditions for the biosynthesis of vitamin B12. RESULTS: We achieved the optimum conditions for the fermentation process of B. megaterium to produce high yield of vitamin B12 in a practical way based on statistical design and analysis which allowed vitamin B12 production to increase up to 759-fold (204.46 µg/l) as compared with control without parameters (0.26 µg/L). High performance liquid chromatography coupled to variable wavelength detector and mass spectrometry has been used to identify vitamin B12 forms and confirm the results. CONCLUSIONS: We developed the fermentation process of B. megaterium to enhance the production of vitamin B12 by providing the required supplements for the synthesis of vitamin B12 (CoCl2, δ-aminolevulinic acid (ALA) and 5,6-dimethylbenzimidazole (DMB)) and dividing the fermentation process into three stages. In addition, the optimum incubation times of the three fermentation stages were investigated and performed with reducing number of experimental and evaluated multiple parameters and their interactions by using statistical experimental design and analysis. All of these strategies has proven successful in enhancing the production of vitamin B12 up to 204.46 µg/l and demonstrated that B. megaterium could be a good candidate for the industrial production of vitamin B12.