Strain L7 Research Articles

Response surface optimization of poly-β-hydroxybutyrate synthesized by Bacillus cereus L17 using acetic acid as carbon source

A strain of Bacillus that can tolerate 10 g/L acetic acid and use the volatile fatty acids produced by the hydrolysis and acidification of activated sludge to produce polyhydroxyalkanoate was screened from the activated sludge of propylene oxide saponification wastewater. The strain was identified by 16S rRNA sequencing and phylogenetic tree analysis and was named Bacillus cereus L17. Various characterization methods showed that the polymer synthesized by strain L17 is poly-β-hydroxybutyrate, which has low crystallinity, good ductility and toughness, high thermal stability and a low polydispersity coefficient. It has wide thermoplastic material operating space as well as industrial and medicinal applications. The optimal fermentation conditions were determined by single factor optimization. Then, Plackett-Burman and Box-Behnken design experiments were carried out according to the single factor optimization results, and the response surface optimization was completed. The final results were: initial pH 6.7, temperature 25 °C, and loading volume 124 mL. The verification experiment showed that the yield of poly-β-hydroxybutyrate after optimization increased by 35.2 % compared to that before optimization.

Bacillus cereus strain L7 lyses Cylindrospermopsis raciborskii through intercellular contact

Cylindrospermopsis raciborskii is a new type of toxic filamentous cyanobacteria commonly found in tropical regions but has recently spread to temperate areas. It is considered an invasive species that poses a serious threat to drinking water sources and human health. Bacillus cereus strain L7, previously isolated and identified by our group, was found to have efficient algicidal activity against C. raciborskii FACHB-1503 in this study. The growth period and initial concentration of L7 had a significant impact on algicidal activity, with L7 culture in the decline phase exhibiting the highest algicidal activity (I = 99.62 %). The ideal environmental conditions for the coculture system were 30 °C, pH = 8, and a light-dark cycle (12 h light: 12 h dark). In experiments to confirm the algicidal mode of L7, the pellet resuspension with R2A medium had the highest algicidal efficiency (I = 98.54 %, t = 6 d), even higher than that of the L7 culture. In contrast, sterile filtrate and pellet resuspension with sterile water had no discernible impact on the algal growth, indicating that L7 performed algicidal activity through direct contact with algal cells. Following 6 days of coculture of L7 culture with C. raciborskii solution, the activity of the PSII system and the photosynthetic efficiency of algal cells were both zero. Following SEM and TEM results, it was discovered that the process of L7 lysis of algal cells begins with the disruption of their multicellular filamentous structure, rupture of the cell wall, efflux of the cell contents, and complete disappearance of the algal cell structure. This is the first research to demonstrate that L7 lyses C. raciborskii directly, which serves as an excellent strategic direction for launching innovative freshwater bloom control strategies.

Effective biodeterioration of a common endocrine disruptor 17β-estradiol using mixed microbial cultures isolated from waste water

Twelve bacteria were isolated from wastewater for the degradation of 17β-estradiol (strain K1, K2, K3, K4, L5, L6, L7, L8, M9, M10, M11 and M12) by enrichment method. Initial screening revealed maximum degradation (78 ± 1.1%, and 86 ± 4.2%) of 17β-estradiol by strains L7, and M10, respectively. The production of manganese peroxidase by bacteria gets involved in the degradation of various contaminants. Almost all strains isolated from the enriched medium produced manganese peroxidase. Based on initial screening on 17β-estradiol degradation and manganese peroxidase production, two bacteria (Flavobacterium longum L7 and Pseudomonas aeruginosa M10) were selected to analyze 17β-estradiol degradation in co-culture system. The co-cultured strains could rapidly remove 17β-estradiol (91.3%) from the culture medium at 10 mg/L 17β-estradiol initial concentration. Moreover, degradation of 17β-estradiol was only 82% and 75% when the strains L7 and M10 were treated individually. Initial 17β-estradiol concentration was 10 mg/L and 0.3% inoculum were optimum and achieved >90% degradation. Alkaline pH was optimum (pH 7.5) and reached 98.4 ± 7.5% degradation in co-culture. Glucose and trehalose improved the growth and degradation of 17β-estradiol. The organic nitrogen sources such as, yeast extract and peptone improved the removal of 17β-estradiol. The selected bacterial strains were resistant against penicillin, erythromycin, oxytetracycline and tetracycline. The present findings demonstrated the application of F. longum L7 and P. aeruginosa M10 co-culture for the removal of steroid hormones in the wastewater.

Use of a Potential Probiotic, Lactobacillus plantarum L7, for the Preparation of a Rice-Based Fermented Beverage.

This study aimed to isolate potential probiotic lactic acid bacteria from a traditional rice-based fermented beverage “bhaati jaanr” and to evaluate their role during preparation of the beverage. Among various isolates, Lactobacillus plantarum strain L7 exhibited satisfactory in vitro probiotic characteristics such as acid resistance and bile tolerance, cell surface hydrophobicity, auto-aggregation, antibiotic susceptibility, and antimicrobial activities. Therefore, performance of L7 as a starter culture in rice fermentation was determined during a 6-day rice fermentation study. L. plantarum L7 decreased the pH, associated with an increase in total titratable acidity and organic acid production up to the 4th day of fermentation. The highest concentrations of succinic acid (0.37 mg/g), lactic acid (4.95 mg/g), and acetic acid (0.36 mg/g) were recorded on the 3rd, 4th, and 5th days of fermentation, respectively. Saccharifying (148.13 μg/min g−1) and liquefying (89.47 μg/min g−1) activities were the highest on days 3 and 2, respectively, and thereafter, they decreased. Phytase activity and the cleavage of free minerals (sodium, calcium, magnesium, manganese, and ferrous) increased up to days 3–4. The concentration of various accumulated malto-oligosaccharides (glucose, fructose, maltotriose, and maltoterose) was noted to be the maximum on days 4 and 5. Furthermore, gas chromatography-mass spectrometry analysis indicated the presence of various volatile compounds. The fermented material also exhibited 1,1-diphenyl-2-picrylhydrazyl and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) radical scavenging activity. Therefore, the probiotic, L. plantarum L7, has a significant role in the fermentation of this beverage and enhances its functional properties.

MICROAEROPHILIC AND ANAEROBIC DECOLORIZATION OF AZO DYE IN RICH AND MINIMAL MEDIA BY CITROBACTER SP. STRAIN L17

This study aimed to investigate the ability of Citrobacter sp. strain L17 to decolourise azo dye in a rich medium (MP5) and three different minimal media (MMP5, MMGF11 and MM63) under microaerophilic and anaerobic conditions. Amaranth was used as the model dye in this investigation. Under microaerophilic condition, reactions were carried out at two different temperatures; 37°C and 45°C, whereas experiments with anaerobic condition were conducted only at 37°C. Results showed that, under microaerophilic condition, full decolourisation of Amaranth was achieved in all media tested at 37°C. However, at 45°C, complete decolourisation was observed only in MP5, MM63 and MMGF11 but no obvious decolourisation occurred in MMP5. On the other hand, complete decolourisation was observed in all media tested under anaerobic condition at 37°C, with the fastest decolourisation in MP5 (rich medium containing glucose and nutrient broth) and the slowest in MMP5. It was found that an inorganic buffer containing glucose at lower concentration was sufficient to achieve complete decolourisation under anaerobic condition. This finding is essential to identify a suitable medium for future study on biogas production by dye-degrading bacteria, which mostly requires anaerobic conditions.

Exogenous labile C application enhances Fe-P utilization for mycorrhizal plants through iron-reducing bacteria in subtropical soil

Abstract In a greenhouse experiment, Medicago sativa was grown in iron-rich soil inoculated with arbuscular mycorrhizal fungi ( Funelliformis mosseae , FM) and/or iron (Fe) -reducing bacteria (IRB, Klebsiella pneumoniae strain L17) with three exogenous labile C input forms (no C source, as a single dose, and as a pulse)at rates of P0 and P5 (0 and 5 mg P 2 O 5 kg -1 , KH 2 PO 4 ), which is to understand the role of IRB in enhancing the Fe-P utilization with exogenous labile C in subtropical soil. The results showed when inoculated with IRB, labile C as a pulsed input significantly improved the mycorrhiza colonization and hyphal length density, and increased the shoot P content, microbial biomass C and glomalin content at P5 rate; meanwhile, the labile C input also promoted the Fe(II) production, when inoculated with IRB, labile C as a pulsed input increased the C/A extractable Fe(II), oxalate-extractable P and microbial biomass P content. These facts suggested that application of labile C as a pulsed input enhanced the ability of IRB in improving the Fe reduction and Fe-P utilization in subtropical soil.

Interactions between Bacillus cereus strain L7 and Anabaena flos-aquae during cyanobacterial blooms

Interactions between Bacillus cereus strain L7 and Anabaena flos-aquae during cyanobacterial blooms

The characteristics of a novel heterotrophic nitrification–aerobic denitrification bacterium, Bacillus methylotrophicus strain L7

Bacillus methylotrophicus strain L7, exhibited efficient heterotrophic nitrification–aerobic denitrification ability, with maximum NH4+-N and NO2--N removal rate of 51.58mg/L/d and 5.81mg/L/d, respectively. Strain L7 showed different gaseous emitting patterns from those strains ever described. When 15NH4Cl, or Na15NO2, or K15NO3 was used, results of GC–MS indicated that N2O was emitted as the intermediate of heterotrophic nitrification or aerobic denitrification, while GC–IRMS results showed that N2 was produced as end product when nitrite was used. Single factor experiments suggested that the optimal conditions for heterotrophic nitrification were sodium succinate as carbon source, C/N 6, pH 7–8, 0g/L NaCl, 37°C and a wide range of NH4+-N from 80 to 1000mg/L. Orthogonal tests showed that the optimal conditions for aerobic denitrification were C/N 20, pH 7–8, 10g/L NaCl and DO 4.82mg/L (shaking speed 50r/min) when nitrite was served as substrate.

Stimulation and inhibition effects of algae-lytic products from Bacillus cereus strain L7 on Anabaena flos-aquae

Alga-bacterium relationships between a Bacillus cereus strain L7 and Anabaena flos-aquae were studied based on the effects of the algicidal substances on algal growth indicators such as enzyme activity and membrane lipid peroxidation. When exposed to algae-lytic products at a concentration of 0.05 mg mL−1, chlorophyll a (Chla), protein and phycobiliprotein contents increased significantly (p < 0.05); superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) activities and malondialdehyde (MDA) concentrations increased slightly to stimulate the algae growth. When exposed to algae-lytic products at a concentration of 0.5 mg mL−1, algae growth and composition were inhibited. Chla, protein and phycobiliprotein concentrations decreased significantly (p < 0.05 for protein, p < 0.01 for Chla and phycobiliprotein). MDA concentrations increased significantly (p < 0.05). POD and CAT activities increased by approximately six and three times, respectively, in 24 h compared with the control, then decreased to the initial level in 4 days. Algae-lytic products have not only inhibition but also stimulation effects on A. flos-aquae. Such effects are associated with antioxidative/oxidative reactions as indicated by the biomarkers SOD, POD, CAT, and MDA.

Screening of potential infants' lactobacilli isolates for amino acids production

A total of 9 lactic acid bacteria (LAB) isolated from Egyptian infants, belonging to lactobacilli strains, were screened and selected according to their amino acids production. The isolate strains produced different essential amino acids in fermentation of M17 medium. Lactobacillus strain L1 produced the highest yield of histidine 92.5 mg/l, L2 strain produced the highest amount of asparagine 59.61 mg/l, cysteine 32.96 mg/l, phenylalanine 17.49 mg/l and tyrosine 244.91 mg/l. On the other hand, Lactobacillus strain L3 is the most potential culture for the production of citrulline 61.12 mg/l and glutamic 68.78 mg/l. This strain also produced the highest amount of lysine, ornithine and proline, which were 57.04, 95.58 and 87.06 mg/l, respectively. While the strain L4 produced the highest amount of glycine 32.12 mg/l, strain L7 produced the highest amount of alanine 147.55 mg/l and strain L5 produced the highest amount of asparatic acid 218.09 mg/l. The present study concluded that the Lactobacillus strains have the potential to produce different amino acids with variance concentrations. Key words: Lactobacilli, Lactobacillus, amino acids, screening, production.

A study of electron-shuttle mechanism in Klebsiella pneumoniae based-microbial fuel cells

In microbial fuel cell (MFC), the rate of electron transfer to anode electrode is a key intrinsic limiting factor on the power output of MFCs. Using Klebsiella pneumoniae (K. pneumoniae) strain L17 as biocatalyst, we studied the mechanism of electron shuttle via self-producing mediator in a cubic air-chamber MFC. To eliminate the influence of biofilm mechanism, the anode electrode was coated with microfiltration membrane (0.22 μm). Data showed that the microfiltration membrane coated and uncoated MFCs achieved the maximum voltage outputs of 316.2 and 426.2 mV after 270 and 120 h, respectively. When the medium was replaced in MFCs that had the highest power generation, the power output dropped by 62.1% and 8.8%, and required 120 and 48 h to resume the original level in the coated and uncoated MFCs, respectively. The results suggested an electron-shuttle mechanism rather than biofilm mechanism was responsible for electricity generation in the membrane coated MFC. Cyclic voltammetric measurements demonstrated the presence of an electrochemical active compound produced by K. pneumoniae strain L17, which was identified to be 2,6-di-tert-butyl-p-benzoquinon (2,6-DTBBQ) by GC-MS. 2,6-DTBBQ, as a recyclable electron shuttle, could transfer electrons between K. pneumoniae L17 and the anode electrode.

Microbial fuel cell with an azo-dye-feeding cathode

Microbial fuel cells (MFCs) were constructed using azo dyes as the cathode oxidants to accept the electrons produced from the respiration of Klebsiella pneumoniae strain L17 in the anode. Experimental results showed that a methyl orange (MO)-feeding MFC produced a comparable performance against that of an air-based one at pH 3.0 and that azo dyes including MO, Orange I, and Orange II could be successfully degraded in such cathodes. The reaction rate constant (k) of azo dye reduction was positively correlated with the power output which was highly dependent on the catholyte pH and the dye molecular structure. When pH was varied from 3.0 to 9.0, the k value in relation to MO degradation decreased from 0.298 to 0.016 micromol min(-1), and the maximum power density decreased from 34.77 to 1.51 mW m(-2). The performances of the MFC fed with different azo dyes can be ranked from good to poor as MO>Orange I>Orange II. Furthermore, the cyclic voltammograms of azo dyes disclosed that the pH and the dye structure determined their redox potentials. A higher redox potential corresponded to a higher reaction rate.

Fe(III) oxide reduction and carbon tetrachloride dechlorination by a newly isolatedKlebsiella pneumoniaestrain L17

To isolate an iron-reducing bacterium and examine its ability of Fe(III) oxide reduction and dechlorination. A fermentative facultative anaerobe, strain L17 isolated from subterranean sediment, can reduce Fe(III) oxides and carbon tetrachloride (CT). It was identified as Klebsiella pneumoniae by 16S rRNA sequence analysis. Strain L17 can metabolize fermentable substrates such as citrate, glycerol, glucose and sucrose coupled with the reduction of hydrous ferric oxide, goethite, lepidocrocite and hematite. Fe(III) reduction was influenced by crystal structure of Fe(III) oxide, type of fermentable substrate, metabolic status of the strain, and significantly enhanced by addition of anthraquinone-2,6-disulfonate (AQDS). Strain L17 could dechlorinate CT to chloroform, and the rate was accelerated in the presence of Fe(III) oxide and AQDS. Biotic dechlorination by strain L17 and abiotic dechlorination by sorbed Fe(II) were proposed as the two main mechanisms. AQDS might accelerate the dechlorination by transferring electrons from strain L17 to Fe(III) oxide and CT. K. pneumoniae L17 can reduce Fe(III) oxides and CT. The two reductions can occur simultaneously, and be significantly promoted by AQDS. This is the first report of a strain of K. pneumoniae capable of reducing Fe(III) oxides and CT. As a strain of environmental origin, strain L17 may have the potential for bioremediation of chlorinated compound-contaminated sites.

Microbial fuel cell based on Klebsiella pneumoniae biofilm

In this paper we reported a novel microbial fuel cell (MFC) based on Klebsiella pneumoniae ( K. pneumoniae) strain L17 biofilm, which can utilize directly starch and glucose to generate electricity. The electrochemical activity of K. pneumoniae and the performance of the MFC were evaluated by cyclic voltammetry, scanning electron microscope (SEM) and polarization curve measurement. The results indicated that an established K. pneumoniae biofilm cells were responsible for the direct electron transfer from fuels to electrode during electricity production. The SEM observation proved the ability of K. pneumoniae to colonize on the electrode surface. This MFC generated power from the direct electrocatalysis by the K. pneumoniae strain L17 biofilm.

Isolation and characterization of a unicellular manganese-oxidizing bacterium from a freshwater lake in northwestern Russia

A unicellular manganese-oxidizing bacterium (strain L7), isolated from Lake Ladoga, is identified as "Siderocapsa" sp. according to its morphology. However, this bacterium belongs to the phylogenetic cluster of Pseudomonas putida. The physiological characteristics (utilization of sugars, polyatomic alcohols, organic acids, and phenolic substrates as carbon and energy sources) also indicate the similarity of strain L7 to representatives of the genus Pseudomonas. The growing culture oxidizes Mn(II); the rate of oxidation depends on the type of added organic substrate. Carbonate requirement for this process indicates mixotrophic metabolism. The relatedness of the isolated bacterium to the representatives of the genus Pseudonomas and their phenotypic similarity provide a basis for considering strain L7 not as "Siderocapsa" sp., but as a new species, Pseudomonas siderocapsa sp. nov., of the P. putida cluster.

Peroxisomicine A1 (plant toxin-514) affects normal peroxisome assembly in the yeast Hansenula polymorpha

Previously we demonstrated that peroxisomicine A1 (T-514), a plant toxin isolated from Karwinskia species, has a deteriorating effect on the integrity of peroxisomes of methylotrophic yeasts. Here we describe two strains of Hansenula polymorpha, affected in the normal utilization of methanol as sole source of carbon and energy due to peroxisomicine A1 treatment. The two strains isolated (L17 and RV31) grew poorly on methanol, apparently due to malfunctioning of their peroxisomes. Moreover, the cells displayed a high peroxisome turnover rate. We argue that the peroxisomicine A1 induced phenotype of both strains is due to a genomic mutation. Strain L17 was functionally complemented after transformation with a H. polymorpha genomic library. The complementing 2.8 kb DNA fragment did not contain a well-defined ORF and led us to speculate that it may contain regulatory sequences that, when present in multiple copies in the cell, result in a change of expression of specific genes, thus causing restoration of normal methylotrophic growth.

Specific nutrient uptake during initiation of somatic embryogenesis in coconut calluses

Changes in the concentration of major nutrients and sugars in the media were measured for two embryogenic strains of coconut calluses, L1 and L7, after 0, 14, 28, 42 and 56 days of culturing on multiplication medium and embryogenesis induction medium. A histological study was made on day 0, 28 and 56. The initation of somatic embryogenesis was achieved in these two strains through two different pathways: one (strain L1), of pluricellular origin, was obtained by decreasing the 2,4-D concentration in the medium, and the other (strain L7), of unicellular origin, by increasing the level of 2,4-D. In spite of these differences, both kinds of embryogenesis initiation were linked to specific requirements: higher uptake per g of dry matter of NH 4 +, Ca 2+, Mg 2+ and sucrose.

Anaerobic oxidation of ferrous iron by purple bacteria, a new type of phototrophic metabolism.

Anoxic iron-rich sediment samples that had been stored in the light showed development of brown, rusty patches. Subcultures in defined mineral media with ferrous iron (10 mmol/liter, mostly precipitated as FeCO3) yielded enrichments of anoxygenic phototrophic bacteria which used ferrous iron as the sole electron donor for photosynthesis. Two different types of purple bacteria, represented by strains L7 and SW2, were isolated which oxidized colorless ferrous iron under anoxic conditions in the light to brown ferric iron. Strain L7 had rod-shaped, nonmotile cells (1.3 by 2 to 3 microns) which frequently formed gas vesicles. In addition to ferrous iron, strain L7 used H2 + CO2, acetate, pyruvate, and glucose as substrate for phototrophic growth. Strain SW2 had small rod-shaped, nonmotile cells (0.5 by 1 to 1.5 microns). Besides ferrous iron, strain SW2 utilized H2 + CO2, monocarboxylic acids, glucose, and fructose. Neither strain utilized free sulfide; however, both strains grew on black ferrous sulfide (FeS) which was converted to ferric iron and sulfate. Strains L7 and SW2 grown photoheterotrophically without ferrous iron were purple to brownish red and yellowish brown, respectively; absorption spectra revealed peaks characteristic of bacteriochlorophyll a. The closest phototrophic relatives of strains L7 and SW2 so far examined on the basis of 16S rRNA sequences were species of the genera Chromatium (gamma subclass of proteobacteria) and Rhodobacter (alpha subclass), respectively. In mineral medium, the new isolates formed 7.6 g of cell dry mass per mol of Fe(II) oxidized, which is in good agreement with a photoautotrophic utilization of ferrous iron as electron donor for CO2 fixation. Dependence of ferrous iron oxidation on light and CO2 was also demonstrated in dense cell suspensions. In media containing both ferrous iron and an organic substrate (e.g., acetate, glucose), strain L7 utilized ferrous iron and the organic compound simultaneously; in contrast, strain SW2 started to oxidize ferrous iron only after consumption of the organic electron donor. Ferrous iron oxidation by anoxygenic phototrophs is understandable in terms of energetics. In contrast to the Fe3+/Fe2+ pair (E0 = +0.77 V) existing in acidic solutions, the relevant redox pair at pH 7 in bicarbonate-containing environments, Fe(OH)3 + HCO3-/FeCO3, has an E0' of +0.2 V. Ferrous iron at pH 7 can therefore donate electrons to the photosystem of anoxygenic phototrophs, which in purple bacteria has a midpoint potential around +0.45 V. The existence of ferrous iron-oxidizing anoxygenic phototrophs may offer an explanation for the deposition of early banded-iron formations in an assumed anoxic biosphere in Archean times.

Pseudomonas ficuserectae sp. nov., the Causal Agent of Bacterial Leaf Spot of Ficus erecta Thunb.

Pseudomonas ficuserectae, a new nonfluorescent, phytopathogenic pseudomonad species, is described. This bacterium produces dark brown, water-soaked spots on the leaves and stems of Ficus erecta Thunb., resulting either in defoliation or shoot blight on severely infected plants. The colonies on nutrient agar plates are white, circular, and 3 to 4 mm in diameter after 6 days. P. ficuserectae is similar to Pseudomonas amygdali in many properties. The differences between these two species include size of colonies on agar plates, hydrolysis of Tween 80, production of H2S, utilization of ribose, raffinose, mannitol, sorbitol, and malonate, and pathogenicity. The deoxyribonucleic acid base composition of the type strain of P. ficuserectae, strain L7, is 59 mol% guanine plus cytosine. The specific epithet of this new species reflects the pathogenicity of the bacterium on F. erecta Thunb.