Biosorption of cadmium,cobalt, nickel,and strontium by aBacillus simplex strain isolated from the vadose zone

A subsurface Gram-positive, endospore-forming, filamentous bacterium, designated ZAN-044, was isolated from a depth of 96.2 m in the vadose zone of the Hanford Site in Washington State. A phylogenetic analysis of the 16S rRNA gene sequence of strain ZAN-044 revealed it to be 99.5% similar toBacillus simplex strain DSM 1321, indicating that they may be members of the same species.B. simplex ZAN-044 was studied along withBacillus subtilis 168, andEscherichia coli K-12 (AB264), two well-characterized metal-sorbing bacteria, for the binding of Cd²⁺, Co²⁺, Ni²⁺, and Sr²⁺. There was rapid (less than 1 h) uptake of 1 M metal by the three bacteria in the order Cd>NiCo>Sr. Binding followed a saturation isotherm at cation concentrations from 0.1 M to 1 mM. Cation binding was pH-dependent, with less binding at low pH.B. simplex ZAN-044 bound more metal thanB. subtilis orE. coli, demonstrating that subsurface microorganisms can remove significant quantities of metals from solution and may be able to influence radionuclide and metal transport in the subsurface.     

Effects of bioaugmentation strategies in UASB reactors with a methanogenic consortium for removal of phenolic compounds

The removal of phenol, ortho- (o-) and para- (p-)cresol was studied with two series of UASB reactors using unacclimatized granular sludges bioaugmented with a consortium enriched against these substances. The parameters studied were the amount of inoculum added to the sludges and the method of immobilization of the inoculum. Two methods were used, adsorption to the biomass or encapsulation within calcium alginate beads. In the bioaugmentation by adsorption experiment, and with a 10% inoculum, complete phenol removal was obtained after 36 d, while 178 d were required in the control reactor. For p-cresol, 95% removal was obtained in the bioaugmented reactor on day 48 while 60 d were required to achieve 90% removal in the control reactor. For o-cresol, the removals were only marginally better with the bioaugmented reactors. Tests performed with the reactors biomass under non-limiting substrate concentrations showed that the specific activities of the bioaugmented biomasses were larger than the original biomass for phenol, and p-cresol even after 276 of operations, showing that the inoculum bacteria successfully colonized the sludge granules. Immobilization of the inoculum by encapsulation in calcium alginate beads, was performed with 10% of the inoculum. Results showed that the best activities were obtained when the consortium was encapsulated alone and the beads added to the sludges. This reactor presented excellent activity and the highest removal of the various phenolic compounds a few days after start-up. After 90 d, a high-phenolic compounds removal was still observed, demonstrating the effectiveness of the encapsulation technique for the start-up and maintenance of high-removal activities.     

Application of actinomycetes to soil to ameliorate water repellency

AIMS: The aim of this study was to develop a novel isolation technique using a mixture of Bacillus and Streptomyces phages to selectively isolate wax-utilizing non-streptomycete actinomycetes effective in ameliorating water repellency in a problem soil. METHODS AND RESULTS: Phages added to a soil suspension reduced the dominance of Bacillus and Streptomyces isolates and significantly increased the number of non-streptomycete actinomycetes on isolation plates. Promising isolates, grown on a medium containing beeswax as sole carbon source, were selected for application to water repellent soil. Their addition significantly reduced water repellency. CONCLUSIONS: Phage application significantly increased the isolation of non-streptomycete actinomycetes. Wax-utilizing isolates were found to significantly reduce water repellency in a problem soil. SIGNIFICANCE AND IMPACT OF THE STUDY: The phage technique can be used for the routine isolation of non-streptomycete actinomycetes. Beeswax medium can be used to selectively isolate wax-utilizing micro-organisms with the potential to ameliorate water repellency in soil.     

Streptomyces sp.FIM-16-06产他克莫司摇瓶发酵条件的优化

采用单因素试验和正交试验研究发酵培养基组成,优化Streptomyces sp. FIM-16-06产他克莫司的发酵条件,探讨摇瓶发酵的主要影响因子初始p H、装液量、转速等发酵参数的影响。确定了适宜的发酵培养基和发酵参数:6. 0%玉米淀粉、2. 0%黄豆粉、2. 0%葡萄糖和0. 5%玉米浆,初始pH 7. 5,装液量50 m L/500 m L三角瓶,种子菌龄48 h,接种量10%,摇床转速250 r/min,发酵温度27℃,发酵周期7 d。优化后的发酵水平较原生产工艺提高60%以上。他克莫司的优化发酵工艺为其工业化生产奠定了基础。     

Factors affecting co-inoculation with antibiotic-producing bacteria to enhance rhizobial colonization and nodulation

Co-inoculation with antibiotic-producing bacteria and rhizobia resistant to those antibiotics has been proposed as a means of promoting colonization and nodulation of legumes by root-nodule bacteria. A study was conducted to establish some of the factors affecting co-inoculation with antibiotic-producing strains of Bacillus and Streptomyces griseus. The stimulation of Rhizobium meliloti and yield and N uptake by alfalfa was enhanced with increasing inoculum size of Bacillus sp. S. griseus and chitin added to soil increased nodulation of soybeans by Bradyrhizobium japonicum and increased nodulation, yield, and number of pods on a second crop grown in the same soil. Bacillus sp. persisted in soil in sufficient numbers for at least 51 days to increase colonization of soybean roots by B. japonicum. The populations of S. griseus, Bacillus sp., and antibiotic-resistant isolates of R. meliloti and B. japonicum fell after their addition to seeds. Nevertheless, a benefical effect by the antibiotic-producing bacteria was evident on R. meliloti colonization of the rhizosphere, nodulation, and yield of alfalfa grown from seeds stored 94 days and on B. japonicum colonization, nodule number, yield, and seed weight of soybeans grown from seeds stored 90 days. Because non-antibiotic-producing derivatives of Bacillus sp. and S. griseus did not promote colonization or nodulation of alfalfa roots by R. meliloti, the benefit of this co-inoculation is a result of antibiotic formation.     

Identification and determination of extracellular phytate-degrading activity in actinomycetes

In this study 97 soil samples from different soil ecosystems were collected. The initial screening was performed on modified glycerol arginine agar (MGAA) to isolate common actinomycetes and on modified MGA-SE (MMGA-SE) to isolate rare actinomycetes. Sixty-seven isolates potentially producing extracellular phytate-degrading activity were identified. The potential to dephosphorylate phytate was confirmed in liquid culture for 46.3 % of the isolates. 12 strains were selected for a direct determination of their phytate-degrading capacity. The results highlighted that the selected isolates produced extracellular phytate-degrading activity; however their capacity in InsP(6) degradation was different. In addition the fermentation medium had an effect on the extent of phytate degradation. Some enzymatic properties of the phytases from isolate No. 43 and isolate No. 63 were determined after obtaining phytase-enriched samples. The enzymes had maximum phytate-degrading capability at 55 °C and pH 5 (isolate No. 43) and 37 °C and pH 7 (isolates No. 63), respectively. Due to their properties, the phytase of isolate No. 43 behaves like a histidine acid phytase, whereas the phytase of No. 63 showed similar enzymatic properties to the phytase of lily. To our knowledge, the results from this study demonstrated for the first time that actinomycetes produce extracellular phytate-degrading activity. By 16SrRNA sequencing, the more closely studied phytase producers were identified as Streptomyces sp. Isolate No. 43 showed 98 % identity to Streptomyces alboniger and S. venezuelae, while isolate No. 63 exhibited 98 % sequence identity to S. ambofaciens and S. lienomycini.     

Microorganisms capable of metabolizing the herbicide metolachlor

We screened several strains of microorganisms and microbial populations for their ability to mineralize or transform the herbicide metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)-acetami de] because such cultures would potentially be useful in the cleanup of contaminated sites. Although we used various inocula and enrichment culture techniques, we were not able to isolate microorganisms that could mineralize metolachlor. However, strains of Bacillus circulans, Bacillus megaterium, Fusarium sp., Mucor racemosus, and an actinomycete were found to transform metolachlor. Several metabolites could be determined with high-performance liquid chromatography. The tolerance of the strains to high concentrations of metolachlor was also evaluated for the usefulness of the strains for decontamination. Tolerance of the actinomycete to metolachlor concentrations over 200 ppm (200 micrograms/ml) was low and could not be increased by doubling the sucrose concentration in the growth medium or by using a large biomass as inoculum. However, a Fusarium sp. could grow and transform metolachlor up to a concentration of 300 ppm.     

Microorganisms capable of metabolizing the herbicide metolachlor.

We screened several strains of microorganisms and microbial populations for their ability to mineralize or transform the herbicide metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)-acetami de] because such cultures would potentially be useful in the cleanup of contaminated sites. Although we used various inocula and enrichment culture techniques, we were not able to isolate microorganisms that could mineralize metolachlor. However, strains of Bacillus circulans, Bacillus megaterium, Fusarium sp., Mucor racemosus, and an actinomycete were found to transform metolachlor. Several metabolites could be determined with high-performance liquid chromatography. The tolerance of the strains to high concentrations of metolachlor was also evaluated for the usefulness of the strains for decontamination. Tolerance of the actinomycete to metolachlor concentrations over 200 ppm (200 micrograms/ml) was low and could not be increased by doubling the sucrose concentration in the growth medium or by using a large biomass as inoculum. However, a Fusarium sp. could grow and transform metolachlor up to a concentration of 300 ppm.     

Functional Establishment of Introduced Chlorobenzoate Degraders Following Bioaugmentation with Newly Activated Soil

Introduced degraders often do not survive when applied to polluted sites; however, the potential for successful bioaugmentation may be increased if newly activated soil (containing indigenous degrader populations recently exposed to the contaminant) or potentially active soil (containing indigenous degrader populations not previously exposed to the contaminant) is used as the inoculant. To investigate this concept, Madera and Oversite soils were amended with 0 or 500 micrograms of 2-, 3-, or 4-chlorobenzoate per gram soil. The Madera degraded 2-chlorobenzoate while the Oversite degraded 3- and 4-chlorobenzoate. After 22 days of incubation, non-active soils that had not degraded chlorobenzoate were bioaugmented with the appropriate activated soil that had been exposed to and degraded chlorobenzoate. Thus, Oversite soil that had not degraded 2-chlorobenzoate was bioaugmented with Madera soil that had degraded 2-chlorobenzoate. Likewise, Madera soil that had not degraded 3- or 4-chlorobenzoate was bioaugmented with the Oversite soil that had degraded 3- or 4-chlorobenzoate. Additionally, the non-active soils were bioaugmented with the corresponding potentially active soils. The Oversite soil amended with activated Madera soil degraded the 2-chlorobenzoate within 3 days of bioaugmentation. The Madera soil amended with activated Oversite soils degraded the 3- and 4-chlorobenzoate within 20 and 6 days, respectively. Large degrader populations developed in microcosms bioaugmented with activated soil, and shifts in the 3- and 4-CB degrader community structures occurred following bioaugmentation. In contrast, bioaugmentation with potentially active soil did not impact degradation. The results indicate the potential for bioaugmentation with newly activated soil to enhance contaminant degradation.     

Detoxification of Toxic Heavy Metals by Marine Bacteria Highly Resistant to Mercury

Pollution in industrial areas is a serious environmental concern, and interest in bacterial resistance to heavy metals is of practical significance. Mercury (Hg), Cadmium (Cd), and lead (Pb) are known to cause damage to living organisms, including human beings. Several marine bacteria highly resistant to mercury (BHRM) capable of growing at 25 ppm (mg L(-1)) or higher concentrations of mercury were tested during this study to evaluate their potential to detoxify Cd and Pb. Results indicate their potential of detoxification not only of Hg, but also Cd and Pb. Through biochemical and 16S rRNA gene sequence analyses, these bacteria were identified to belong to Alcaligenes faecalis (seven isolates), Bacillus pumilus (three isolates), Bacillus sp. (one isolate), Pseudomonas aeruginosa (one isolate), and Brevibacterium iodinium (one isolate). The mechanisms of heavy metal detoxification were through volatilization (for Hg), putative entrapment in the extracellular polymeric substance (for Hg, Cd and Pb) as revealed by the scanning electron microscopy and energy dispersive x-ray spectroscopy, and/or precipitation as sulfide (for Pb). These bacteria removed more than 70% of Cd and 98% of Pb within 72 and 96 h, respectively, from growth medium that had initial metal concentrations of 100 ppm. Their detoxification efficiency for Hg, Cd and Pb indicates good potential for application in bioremediation of toxic heavy metals.     

Metal tolerance and biosorption potential of filamentous fungi isolated from metal contaminated agricultural soil

Heavy metal analysis of agricultural field soil receiving long-term (>20 years) application of municipal and industrial wastewater showed two- to five-fold accumulation of certain heavy metals as compared to untreated soil. Metal-resistant fungi isolated from wastewater-treated soil belonged to genera Aspergillus, Penicillium, Alternaria, Geotrichum, Fusarium, Rhizopus, Monilia and Trichoderma. Minimum inhibitory concentrations (MIC) for Cd, Ni, Cr, Cu, and Co were determined. The MIC ranged from 0.2 to 5 mg ml(-1) for Cd, followed by Ni (0.1-4 mg ml(-1)), Cr (0.3-7 mg ml(-1)), Cu (0.6-9 mg ml(-1)) and for Co (0.1-5 mg ml(-1)) depending on the isolate. Aspergillus and Rhizopus isolates were tested for their metal biosorption potential for Cr and Cd in vitro. Biosorption experiments were conducted with initial metal concentrations of 2, 4, 6 and 8 mM with a contact time of 4 h and wet fungal biomass (1-5 g) at 25 degrees C. Maximum biosorption of Cr and Cd ions was found at 6 mM initial metal concentration. Aspergillus sp.1 accumulated 1.20 mg of Cr and 2.72 mg of Cd per gram of biomass. Accumulation of these two metals by very tolerant Aspergillus sp.2 isolate was at par with relatively less tolerant Aspergillus sp.1 isolate. Rhizopus sp. accumulated 4.33 mg of Cr and 2.72 mg of Cd per g of biomass. The findings indicated promising biosorption of cadmium and chromium by the Rhizopus and Aspergillus spp. from aqueous solution. There is little, if any, correlation between metal tolerance and biosorption properties of the test fungi.     

Enhanced uptake of Cd by biofilm forming Cd resistant plant growth promoting bacteria bioaugmented to the rhizosphere of Vetiveria zizanioides

Abstract

Heavy metals are the major cause of pollution and cadmium is one among the highly toxic metals discharged into the environment from various industries. The current study was focused on the bioremoval of cadmium by phyto and rhizoremediation approach using Vetiveria zizanioides. The bacterial strains were isolated from wetland paddy rhizosphere soil and the isolate VITJAN13 was found to be a biofilm forming Cd resistant plant growth promoting rhizobacteria (PGPR). The 16S rRNA gene sequencing revealed VITJAN13 to be the closest neighbor of Aeromonas sp. and was submitted to Genbank with the accession number KX770741. Further, pot culture studies indicated that the treatments bioaugmented with VITJAN13 increased the root length and shoot height by 21.4 and 17.36%, respectively as compared to the non-augmented plants. Hence, bioaugmentation of Aeromonas sp. in the rhizosphere of Vetiveria zizanioides enhanced the uptake of cadmium by 67.7% in the soil treated with 15?mg/kg of Cd to that of the phytoremediation setup.     

Do detached root-cap cells influence bacteria associated with maize roots?

Abstract. A model rhizosphere has been used which consisted of detached root-cap cells of maize in their surrounding root-cap mucilage on the surface of Noble agar. These cells were co-cultured for periods up to 32 d with eight different bacterial isolates from soil-grown roots and surrounding soil and two laboratory cultures. Cap cells were unaffected by the bacteria. There were five different type-specific responses of the bacteria in proximity to the cap cells. There were, strong growth inhibition ( Rhizobium sp. and Escherichia coli ), strong stimulation ( Pseudomonas fluorescens , laboratory strain), mixed weak inhibition or stimulation ( Pseudomonas fluorescens , field isolate), early inhibition followed by strong stimulation then spore formation ( Bacillus spp.), no effect ( Streptomyces sp. and Cytophaga sp.). It is concluded that detached root-cap cells are actively involved in the establishment of characteristic rhizosphere bacterial microflora.     

Influence of Temperature on the Virulence of Two Races of Meloidogyne chitwoodi on Wheat and Barley

Races of the Columbia root-knot nematode, Meloidogyne chitzooodi, from Idaho (R1) and Utah (R2) suppressed (P < 0.05) tillering of Dusty winter wheat, Fielder spring wheat, Luther winter barley, and Steptoe spring barley at 15-30 C. Nematode inoculum density was negatively correlated with tillering (r = -0.79). Inoculum densities of both nematode races were negatively correlated with heads per plant (r = -0.83), head length (r = -0.87), and head dry weight (r = 0.73) of Fielder spring wheat and Steptoe spring barley at all temperatures; the greatest growth restrictions occurred at Pi 20 eggs/cm³ soil. Both nematode races were most damaging at 25-30 C. Fielder spring wheat and Steptoe spring barley inoculated with R2 produced fewer heads than R1 when inoculated at 15 C, whereas the same cultivars inoculated with R1 produced fewer heads than R2 at 30 C. No differences were observed between root growth of winter and spring wheat or between winter and spring barley. Nematode reproduction was positively correlated to temperature (r = 0.87) and negatively correlated with inoculum density (r = -0.86). Reproductive rates were greatest with Pi = 2 eggs/cm³ soil at 25 C and lowest with Pi = 20 eggs/cm³ soil at 15 C for both nematode races.     

Cadmium and zinc sensitivity and tolerance in Bacillus subtilis subsp. niger and in a Pseudomonas sp.

The action of Cd2+ and Zn2+ on Bacillus subtilis subsp. niger ATCC 9372, and on a Pseudomonas sp. (possibly Pseudomonas fluorescens), isolated from cadmium-polluted soil has been determined and compared with results obtained previously with Klebsiella aerogenes. In liquid medium the lag and the mean generation time of Bacillus subtilis subsp, niger increased with increasing Cd2+ or Zn2+ concentrations whereas only the total biomass of the Pseudomonas sp. was affected. Nevertheless, the responses of both species indicated a specific action at low concentrations and a more general toxic action at high concentrations. The survival on Cd2+ - or Zn2+ - agar depended on the state of the metal ions with regard to chelating ligands and on the nutritional stage of the organisms. In admixture, the metal ions acted synergistically, particularly on the Pseudomonas sp. Resistance to both metal ions developed. It was graded to the training concentration and reciprocal cross-resistance occurred with Bacillus subtilis. subsp. niger but not with the Pseudomonas sp.     

Role of inoculum preparation and density on the bioremediation of 2,4-D-contaminated soil by bioaugmentation

The effect of inoculum preparation and density on the efficiency of remediation of 2,4-dichlorophenoxyacetic acid (2,4-D) by bioaugmentation was studied in non-sterile soil. A 2,4-D-degrading Pseudomonas cepacia strain (designated BRI6001) was used initially in liquid culture to determine the effects of pre-growth induction and of inoculum density. The time for complete 2,4-D degradation was reduced by 0.5 day for each log increase of inoculum density. In mixed (BRI6001 and soil bacteria) liquid cultures, a competition effect for 2,4-D became apparent at low inoculum levels (less than 10 10⁵ cfu/ml BRI6001 for 10⁸ cfu/ml soil bacteria) but only when the soil bacteria included indigenous 2,4-D degraders. In static non-sterile soil, the effect of inoculum density on 2,4-D degradation was comparable to that in liquid culture but only at high inoculation levels. At lower levels, a biological effect for 2,4-D degradation became apparent, as was observed in mixed liquid cultures, whereas at intermediate levels, a combination of biological, physical and chemical factors decreased the efficiency of bioaugmentation. The acclimation period for 2,4-D degradation in soil bioaugmented with BRI6001 reflected mainly the time required for cell induction and, presumably, for overcoming the physical limitation of diffusion of both 2,4-D and added bacteria in the soil matrix. Correspondence to: R. SamsonISSUED AS NRCC 33848     

Production of extracellular alkaline alpha-amylase by solid state fermentation with a newly isolated Bacillus sp

Production of alkaline alpha-amylase employing our laboratory isolate, Bacillus sp., under solid state fermentation, was optimized. The effect of wheat bran and lentil husk was examined. Lentil husk exhibited the highest enzyme production. The appropriate incubation time, inoculum size, moisture level, and buffer solution level were determined. Maximum yields of 216,000 and 172,800 U/g were achieved by employing lentil husk and wheat bran as substrates in 0.1 M carbonate/bicarbonate buffer at pH 10.0 with 30% initial moisture level at 24 h. Inoculum size and buffer solution level were found to be 20% and 1:0.5 for two solid substrates.     

The Bioadsorption of Cadmium and Lead by Bacteria in Root Exudates Culture

Microorganisms are important for phytoremediation of soil contaminated with heavy metals. In the present study, bacteria Bacillus sp., Pseudomonas sp., Alcaligenes sp., and Flavobacterium sp. isolated from the Zhangshi Irrigation Area were applied to bioadsorbed Cd and Pb in liquid cultures with root exudates of sunflower as a sole carbon source. The experimental data demonstrated that these bacteria had a high potential of enrichment of Cd and Pb, and Bacillus sp. and Alcaligenes sp. had better ability to accumulate Cd or Pb than the others; the distinct bioadsorption of Cd and Pb by bacteria might depend on the physiology of bacteria, categories of heavy metals, and environmental factors (such as pH). In addition, root exudates of sunflower could not only support the growth of these bacteria, but also influence the toxicity and bioavailability of Cd and Pb. Our results indicated that amendment with bacteria isolated from heavy-metal-polluted soil and root exudates could be considered as a potential approach to enhance the phytoremediation of Cd- or Pb-contaminated soil.     

连作花生田根际土壤优势微生物的分离和鉴定

【目的】从不同连作年限的花生田根际土壤中分离优势微生物并进行鉴定,为研究花生连作后优势微生物的变化奠定基础。【方法】采用土壤稀释分离法从不同连作年限花生根际土壤中分离优势细菌、真菌和放线菌,结合菌株形态特征、培养性状、生理生化特征及16S rDNA序列分析对细菌、放线菌进行鉴定,通过形态特征、培养特征和分子鉴定方法对优势真菌进行鉴定。【结果】从连作花生田根际土壤中分离鉴定出7种优势细菌、7种优势真菌和7种优势放线菌。7种优势细菌分别为Leifsonia xyli、氯酚节杆菌(Arthrobacterchlorophenolicus)、黄色微杆菌(Microbacterium flavescens)、鞘氨醇单胞菌属(Sphingomonas sp.)、巴斯德菌属(Pasteurella sp.)、简单芽孢杆菌(Bacillus simplex)和巨大芽孢杆菌(Bacillus megaterium)。7种优势真菌分别为枝状枝孢菌(Cladosporium cladosporioides)、产紫青霉(Penicillium purpurogenum)、哈茨木霉有性型(Hypocrea lixii)、Exophiala pisciphila、微紫青霉(Penicillium janthinellum)、曲霉(Aspergillus sp.)和大丽轮枝菌(Verticillium dahliae)。7种优势放线菌分别为紫红链霉菌(Streptomyces violaceoruber)、华丽黄链霉菌(Streptomyces flaveus)、Streptomyces panaciterrae、不产色链霉菌(Streptomyces achromogenes)、假浅灰链霉菌(Streptomyces pseudogriseolus)、纤维素链霉菌(Streptomyces cellulosae)和金色链霉菌(Streptomyces aureus)。【结论】本研究是第一次系统的从连作花生根际土中分离鉴定优势微生物,种植花生后根际土壤中优势微生物的种类发生了明显变化,但变化没有规律。     

Absolute configuration of hydroxycitric acid produced by microorganisms

Optical resolution for (2S,3R) and (2R,3S)-hydroxycitric acid (HCA) enantiomers was developed using chiral column chromatography. HCA from Bacillus megaterium G45C and Streptomyces sp. U121, newly isolated in our previous study, was analyzed to determine the absolute configuration. These results indicate that both strains generate optically pure (2S,3R)-hibiscus type HCA enantiomer.