Search results for: Bacillus licheniformis LHH100

Authors: Laribi-Habchi Hasiba, Bouanane-Darenfed Amel, Drouiche Nadjib, Pausse André, Mameri Nabil

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Chitin, a linear 1, 4-linked N-acetyl-d-glucosamine (GlcNAc) polysaccharide is the major structural component of fungal cell walls, insect exoskeletons and shells of crustaceans. It is one of the most abundant naturally occurring polysaccharides and has attracted tremendous attention in the fields of agriculture, pharmacology and biotechnology. Each year, a vast amount of chitin waste is released from the aquatic food industry, where crustaceans (prawn, crab, Shrimp and lobster) constitute one of the main agricultural products. This creates a serious environmental problem. This linear polymer can be hydrolyzed by bases, acids or enzymes such as chitinase. In this context an extracellular chitinase (ChiA-65) was produced and purified from a newly isolated LHH100. Pure protein was obtained after heat treatment and ammonium sulphate precipitation followed by Sephacryl S-200 chromatography. Based on matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/MS) analysis, the purified enzyme is a monomer with a molecular mass of 65,195.13 Da. The sequence of the 27 N-terminal residues of the mature ChiA-65 showed high homology with family-18 chitinases. Optimal activity was achieved at pH 4 and 75◦C. Among the inhibitors and metals tested p-chloromercuribenzoic acid, N-ethylmaleimide, Hg2+ and Hg + completelyinhibited enzyme activity. Chitinase activity was high on colloidal chitin, glycol chitin, glycol chitosane, chitotriose and chitooligosaccharide. Chitinase activity towards synthetic substrates in the order of p-NP-(GlcNAc) n (n = 2–4) was p-NP-(GlcNAc)2> p-NP-(GlcNAc)4> p-NP-(GlcNAc)3. Our results suggest that ChiA-65 preferentially hydrolyzed the second glycosidic link from the non-reducing end of (GlcNAc) n. ChiA-65 obeyed Michaelis Menten kinetics the Km and kcat values being 0.385 mg, colloidal chitin/ml and5000 s−1, respectively. ChiA-65 exhibited remarkable biochemical properties suggesting that this enzyme is suitable for bioconversion of chitin waste.

Keywords: Bacillus licheniformis LHH100, characterization, extracellular chitinase, purification

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Authors: Padmanabhan D, Kavitha S

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The purpose of the study is to investigate arsenic resistance ability of indigenous microflora and its ability to utilize arsenic species form containing water source. PS-6 potential arsenic tolerance bacterium was screened from thirty isolates from Pichavaram Mangroves of India having tolerance to grow up to 1000 mg/l of As (V) and 800 mg/l of As (III) and arsenic utilization ability of 98 % of As (V) and 97% of As (III) with initial concentration of 3-5 mg/l within 48 hrs. Optimum pH and temperature was found to be ~7-7.4 and 37°C. Active growth of PS-6 in minimal salt media (MSB) helps in cost effective biomass production. Dry weight analysis of PS-6 has shown significant difference in biomass when exposed to As (III) and As (V). Protein level study of PS-6 after exposing to As (V) and As (III) shown modification in total protein concentration and variation in SDS-PAGE pattern. PS-6 was identified as Bacillus licheniformis based on partially sequenced of 16S rRNA using NCBI Blast. Further investigation will help in using this potential bacterium as a well-grounded source for urgency.

Keywords: arsenite, arsenate, Bacillus licheniformis, utilization

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Authors: Sahira Nsayef Muslim, Israa M. S. Al-Kadmy, Nadheema Hammood Hussein, Alaa Naseer Mohammed Ali, Buthainah Mohammed Taha, Rayim Sabah Abbood, Sarah Naji Aziz

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A novel strain of Bacillus licheniformis isolated from spoilt cucumber and pepper samples have the ability to produce the chitosanase enzyme when grown on chitosan substrate. Chitosanase was purified to homogeneity with a recovery yield of 35.71% and 5.5 fold of purification by using ammonium sulfate at 45% saturation followed by ion exchange chromatography on DEAE-cellulose column and gel filtration chromatography on Sephadex G-100 column. The purified chitosanase inhibited the biofilm formation ability for all Gram-negative and Gram-positive biofilm-forming bacteria (biofilm producers) after using Congo Red agar and Microtiter plates methods. Highly antibiofilm of chitosanase recorded against Pseudomonas aeruginosa followed by Klebsiella pneumoniae with reduction of biofilm formation ratio to 22 and 29%, respectively compared with (100)% of control. Thus, chitosanase has promising benefit as antibiofilm agent against biofilm forming pathogenic bacteria and has promising application as alternative antibiofilm agents to combat the growing number of multidrug-resistant pathogen-associated infections, especially in situation where biofilms are involved.

Keywords: chitosanase, Bacillus licheniformis, vegetables, biofilm

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Authors: Patience Orobosa Olajide

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Five hydrocarbon degrading bacterial strains isolated from contaminated environment were investigated with respect to polyhydroxybutyrate (PHB) biosynthesis. Screening for bioplastic production was done on assay mineral salts agar medium containing 0.2% poly (3-hydroxybutyrate) as the sole carbon source. Two of the test bacteria were positive for PHB biosynthesis and were identified based on gram staining, biochemical tests, 16S rRNA gene sequence analysis as Pseudomonas aeruginosa and Bacillus licheniformis which grew at 37 and up to 65 °C respectively, thus suggesting the later to be thermotolerant. In this study, the effects of different carbon and nitrogen sources on PHB production in these strains were investigated. Maximum PHB production was obtained in 48 hr for the two strains and amounted to yields of 72.86 and 62.22 percentages for Bacillus licheniformis and Pseudomonas aeruginosa respectively. In these strains, glycine was the most efficient carbon sources for the production of PHB compared with other carbon (glucose, lactose, sucrose, Arabinose) and nitrogen (L- glycine, L-cysteine, DL-Tryptophan, and Potassium Nitrate) sources. The screening of microbial strains for industrial PHB production should be based on several factors including the cell’s capability to mineralize an inexpensive substrate, rate of growth and the extent of polymer accumulation.

Keywords: bacteria, poly-3-hydroxybutyrate (PHB), hydrocarbon, thermotolerant

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Authors: Saba Pajuhan, Afshin Farahbakhsh, S. M. M. Dastgheib

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Biosurfactants and bioemulsifiers are a structurally diverse group of surface-active molecules synthesized by microorganisms, they are amphipathic molecules which reduce surface and interfacial tensions and widely used in pharmaceutical, cosmetic, food and petroleum industries. In this paper, several methods of bioemulsifer synthesis and purification by Bacillus licheniformis strains (namely ACO1, PTCC 1595 and ACO4) were investigated. Strains were grown in nutrient broth with different conditions in order to get maximum production of bioemulsifer. The purification of bio emulsifier and the quality evaluation of the product was done by adding sulfuric acid (H₂SO₄) (98%), Ethanol or HCl to the solution followed by centrifuging. To determine the optimal conditions yielding the highest bioemulsifier production, the effect of various carbon and nitrogen sources, temperature, NaCl concentration, pH, O₂ levels, incubation time are indispensable and all of them were highly effective in bioemulsifiers production.

Keywords: biosurfactant, bioemulsifier, purification, surface tension, interfacial tension

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Authors: Rashida Rahmat Zohra, Afsheen Aman, Shah Ali Ul Qader

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Commercially important biopolymer, dextran, is enzymatically degraded into lower molecular weight fractions of vast industrial potential. Various organisms are associated with dextranase production, among which fungal, yeast and bacterial origins are used for commercial production. Dextranases are used to remove contaminating dextran in sugar processing industry and also used in oral care products for efficient removal of dental plaque. Among the hydrolytic products of dextran, isomaltooligosaccharides have prebiotic effect in humans and reduces the cariogenic effect of sucrose in oral cavity. Dextran derivatives produced by hydrolysis of high molecular polymer are also conjugated with other chemical and metallic compounds for usage in pharmaceutical, fine chemical industry, cosmetics, and food industry. Owing to the vast application of dextran and dextranases, current study focused on purification and analysis of kinetic parameters of dextranase from a newly isolated strain of Bacillus licheniformis KIBGE-IB25. Dextranase was purified up to 35.75 folds with specific activity of 1405 U/mg and molecular weight of 158 kDa. Analysis of kinetic parameters revealed that dextranase performs optimum cleavage of low molecular weight dextran (5000 Da, 0.5%) at 35ºC in 15 min at pH 4.5 with a Km and Vmax of 0.3738 mg/ml and 182.0 µmol/min, respectively. Thermal stability profiling of dextranase showed that it retained 80% activity up to 6 hours at 30-35ºC and remains 90% active at pH 4.5. In short, the dextranase reported here performs rapid cleavage of substrate at mild operational conditions which makes it an ideal candidate for dextran removal in sugar processing industry and for commercial production of low molecular weight oligosaccharides.

Keywords: Bacillus licheniformis, dextranase, gel permeation chromatograpy, enzyme purification, enzyme kinetics

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Authors: Cheba Ben Amar

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Bacillus and Paenibacillus are Gram-positive aerobic endospore-forming bacteria (AEFB) and most abundant in the rhizosphere, they mediated plant growth promotion and disease protection by several complex and interrelated processes involving direct and indirect mechanisms that include nitrogen fixation, phosphate solubilization, siderophores production, phytohormones production and plant diseases control. In addition to their multiple PGPR properties, high secretory capacity, spore forming ability and spore resistance to unfavorable conditions enabling their extended commercial applications for long shelf-life. Due to these unique advantages, Bacillus species were the most an ideal candidate for developing efficient PGPR products such as biopesticides, fungicides and fertilizers. This review list all studied and reported plant growth promoting Bacillus species and strains, discuss their capacities to enhance plant growth and protection with special focusing on the most frequent species Bacillus subtilis, B. pumilus ,B. megaterium, B. amyloliquefaciens , B. licheniformis and B. sphaericus, furthermore we recapitulate the beneficial activities and mechanisms of several species and strains of the genus Paenibacillus involved in plant growth stimulation and plant disease control.

Keywords: bacillus, paenibacillus, PGPR, beneficial activities, mechanisms, growth promotion, disease control, agrobiotechnology

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Authors: A. Ribeiro, N. Valério, C. Vilarinho, J. Araujo, J. Carvalho

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Res2ValHUM project involves institutions from the Spanish Autonomous Region of Galicia and the north of Portugal (districts of Porto and Braga) and has as overall objectives of promotion of composting as an process for the correct managing of organic waste, valorization of compost in different fields or applications for the constitution of products with high added value, reducing of raw materials losses, and reduction of the amount of waste throw in landfills. Three main actions were designed to achieve the objectives: development of a management tool to improve collection and residue channeling for composting, sensibilization of the population for composting and characterization of the chemical and biological properties of compost and humic and fulvic substances to envisage high-value applications of compost. Here we present the cooperative activity of Galician and northern Portuguese institutions to valorize organic waste in both regions with common socio-economic characteristics and residue management problems. Results from the creation of the resource manage tool proved the existence of a large number of agricultural wastes that could be valorized. In the North of Portugal, the wastes from maize, oats, potato, apple, grape pomace, rye, and olive pomace can be highlighted. In the Autonomous Region of Galicia the wastes from maize, wheat, potato, apple, and chestnuts can be emphasized. Regarding the isolation and identification of microbial consortia from compost samples, results proved microorganisms belong mainly to the genus Bacillus spp. Among all the species identified in compost samples, Bacillus licheniformis can be highlighted in the production of humic and fulvic acids.

Keywords: agricultural wastes, Bacillus licheniformis, Bacillus spp., humic-acids, fulvic-acids

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Authors: S. V. Ukwungwu, A. J. Abbas, G. G. Nasr

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The increasing high price of natural gas and oil with attendant increase in energy demand on world markets in recent years has stimulated interest in recovering residual oil saturation across the globe. In order to meet the energy security, efforts have been made in developing new technologies of enhancing the recovery of oil and gas, utilizing techniques like CO2 flooding, water injection, hydraulic fracturing, surfactant flooding etc. Surfactant flooding however optimizes production but poses risk to the environment due to their toxic nature. Amongst proven records that have utilized other type of bacterial in producing biosurfactants for enhancing oil recovery, this research uses a technique to combine biosurfactants that will achieve a scale of EOR through lowering interfacial tension/contact angle. In this study, three biosurfactants were produced from three Bacillus species from freeze dried cultures using sucrose 3 % (w/v) as their carbon source. Two of these produced biosurfactants were screened with the TEMCO Pendant Drop Image Analysis for reduction in IFT and contact angle. Interfacial tension was greatly reduced from 56.95 mN.m-1 to 1.41 mN.m-1 when biosurfactants in cell-free culture (Bacillus licheniformis) were used compared to 4. 83mN.m-1 cell-free culture of Bacillus subtilis. As a result, cell-free culture of (Bacillus licheniformis) changes the wettability of the biosurfactant treatment for contact angle measurement to more water-wet as the angle decreased from 130.75o to 65.17o. The influence of microbial treatment on crushed rock samples was also observed by qualitative wettability experiments. Treated samples with biosurfactants remained in the aqueous phase, indicating a water-wet system. These results could prove that biosurfactants can effectively change the chemistry of the wetting conditions against diverse surfaces, providing a desirable condition for efficient oil transport in this way serving as a mechanism for EOR. The environmental friendly effect of biosurfactants applications for industrial purposes play important advantages over chemically synthesized surfactants, with various possible structures, low toxicity, eco-friendly and biodegradability.

Keywords: bacillus, biosurfactant, enhanced oil recovery, residual oil, wettability

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Authors: Muhammad Shahzad Tufail, Iram Liaqat, Umer Sohail Meer, Muhammad Ishtaiq, Muhammad Sattar

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Nanotechnology is a vibrant field with numerous applications in many different branches of science and technology. Several methods are used to synthesize nanoparticles (NPs), which have multiple range of applications. Comparatively, the biogenic synthesis of NPs is a more economical and environmentally favourable method than the traditional chemical method. The current study aims to synthesize biogenically silver nanoparticles (AgNPs) using bacterial isolates. Four bacterial strains Escherichia coli (MT448673), Pseudomonas aeruginosa (MN900691), Bacillus subtilis (MN900684) and Bacillus licheniformis (MN900686) were used for the synthesis of AgNPs from silver nitrate (AgNO3) solution. The biofilm time kinetics of four bacterial isolates (P. aeruginosa, E. coli, B. licheniformis and B. subtilis) was analysed by incubating bacterial cultures at 37◦C in test tubes over a period of different time intervals i.e., 2, 3, 5 and 7 days following crystal violet staining method. All the four strains had ability to form strong biofilms between 48 to 72 hours of incubation. Two strains (B. subtilis and B. licheniformis) formed significant (p < 0.05) biofilm after 3 days of incubation period. The other two strains (E. coli and P. aeruginosa) showed strong biofilm formation after 2 days of incubation. Next, the antibiofilm activity of biogenically synthesized AgNPs (10 - 100 µgmL-1) was analysed against biofilm forming human pathogenic bacteria. Findings of the work revealed that 60-90% inhibition was observed at 60 µgmL-1 of AgNPs, while maximum inhibition (i.e.,100%) was found at highest concentration (90 µgmL-1). It was evident that highly significant (p < 0.05) decrease in biofilm formation was observed with increasing concentration of AgNPs.

Keywords: antibiofilm, biofilm formation, nanotechnology, pathogenic bacteria, silver nanoparticles

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Authors: I. Savitskaya, A. Kistaubayeva, A. Zhubanova, I. Blavachinskaiya, D. Ibrayeva, M. Abdulzhanova, A. Otarbay, A.Isabekova

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This study was conducted for the investigation of number of cellulolytic bacteria and their ability in decomposition. Seven samples surface soil were collected on cellulose Zailiskii Alatau slopes. Cellulolitic activity of new strains of Bacillus, isolated from soil is determined. Isolated cellulose degrading bacteria were screened for determination of the highest cellulose activity by quantitative assay using Congo red, gravimetric assay and colorimetric DNS method trough of the determination of the parameters of sugar reduction. Strains are assigned to: B.subtilis, B.licheniformis, B. cereus and, В. megaterium. Bacillus strains consisting of several different types of cellulases have broad substrate specificity of cellulase complexes formed by them. Cellulolitic bacteria were recorded to have highest cellulase activity and selected for optimization of cellulase enzyme production.

Keywords: cellulose-degrading bacteria, cellulase complex, foothills soil, screening

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Authors: Ayoub Azizi-Shotorkhoft, Tahereh Mohammadabadi, Hosein Motamedi, Morteza Chaji, Hasan Fazaeli

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This study was conducted to investigate nutritive value of wheat straw processed with termite gut symbiotic bacteria with lignocellulosic-degrading potential including Bacillus licheniformis, Ochrobactrum intermedium and Microbacterium paludicola in vitro. These bacteria were isolated by culturing termite guts contents in different culture media containing different lignin and lignocellulosic materials that had been prepared from water-extracted sawdust and wheat straw. Results showed that incubating wheat straw with all of three isolated bacteria increased (P<0.05) acid-precipitable polymeric lignin (APPL) compared to control, and highest amount of APPL observed following treatment with B. licheniformis. Highest and lowest (P<0.05) in vitro gas production and ruminal organic matter digestibility were obtained when treating wheat straw with B. licheniformis and control, respectively. However, other fermentation parameters such as b (i.e., gas production from the insoluble fermentable fractions at 144h), c (i.e., rate of gas production during incubation), ruminal dry matter digestibility, metabolizable energy, partitioning factor, pH and ammonia nitrogen concentration were similar between experimental treatments (P>0.05). It is concluded that processing wheat straw with isolated bacteria improved its nutritive value as ruminants feed.

Keywords: termite gut bacteria, wheat straw, nutritive value, ruminant

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Authors: Ifueko Oghogho Ukponmwan, Mike O. Orji

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This study investigated the antifungal activity of Bacilluss subtilis in the control of postharvest fungal rot of white yam (Dioscorea spp). Bacillus subtilis was isolated from the soil and fungi (Aspergillus spp, Mucor and yeasts) were isolated from rotten yam. The organisms were paired in yam nutrient agar (YNA) and yam Sabourraud dextrose agar media. In the yam dextrose agar media (YSDA) plates, the Bacillus grew rapidly and established itself and restricted the growth of the fungi organisms, but there was no zone of inhibition. This behaviour of Bacillus on the plates of YSDA was also observed in the yams where the fungi caused rot but the rot was suppressed by the presence of the Bacillus as compared to the degree of rot observed in the control that had only spoilage fungi. The control yam showed greater rot than other yams that contained a combination of Bacillus and fungi. The t-Test analysis showed that the difference in the rot between the treated samples and the control sample is significant and this implies that the presence of Bacillus significantly reduced the growth of fungi in the samples (yams). It was revealed from this study that Bacillus subtilis treatment can be successfully used to preserve white yams in storage. Its fast growth and early establishment in the sample accounts for its antifungal strength.

Keywords: Bacillus subtilis, rot, fungi, yam

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Authors: Wayde Veldman, Ozlem T. Bishop, Igor Polikarpov

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In bacteria, mono and disaccharides are phosphorylated during uptake into the cell via the widely used phosphoenolpyruvate (PEP)-dependent phosphotransferase transport system. As an initial step in the phosphorylated disaccharide metabolism pathway, certain glycoside hydrolase family 1 (GH1) enzymes play a crucial role in releasing phosphorylated and non-phosphorylated monosaccharides. However, structural determinants for the specificity of these enzymes still need to be clarified. GH1 enzymes are known to have a wide array of functions. According to the CAZy database, there are twenty-one different enzymatic activities in the GH1 family. Here, the structure and substrate specificity of a GH1 enzyme from Bacillus licheniformis, hereafter known as BlBglH, was investigated. The sequence of the enzyme BlBglH was compared to the sequences of other characterized GH1 enzymes using sequence alignment, sequence identity calculations, phylogenetic analysis, and motif discovery. Through these various analyses, BlBglH was found to have sequence features characteristic of the 6-phospho-β-glucosidase activity enzymes. Additionally, motif and structure comparisons of the three most commonly studied GH1 enzyme-activities revealed a shared loop amongst the different structures that consist of different sequence motifs – this loop is thought to guide specific substrates (depending on activity) towards the active-site. To further affirm BlBglH enzyme activity, molecular docking and molecular dynamics simulations were performed. Docking was carried out using 6-phospho-β-glucosidase enzyme-activity positive (p-Nitrophenyl-beta-D-glucoside-6-phosphate) and negative (p-Nitrophenyl-beta-D-galactoside-6-phosphate) control ligands, followed by 400 ns molecular dynamics simulations. The positive-control ligand maintained favourable interactions within the active site until the end of the simulation. The negative-control ligand was observed exiting the enzyme at 287 ns. Binding free energy calculations showed that the positive-control complex had a substantially more favourable binding energy compared to the negative-control complex. Jointly, the findings of this study suggest that the BlBglH enzyme possesses 6-phospho-β-glucosidase enzymatic activity.

Keywords: 6-P-β-glucosidase, glycoside hydrolase 1, molecular dynamics, sequence analysis, substrate specificity

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Authors: Murada Alholy, Mengshi Lin, Omar Alhaj, Mahmoud Abugoush

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Alicyclobacillus is a causative agent of spoilage in pasteurized and heat-treated apple juice products. Differentiating between this genus and the closely related Bacillus is crucially important. In this study, Fourier transform infrared spectroscopy (FT-IR) was used to identify and discriminate between four Alicyclobacillus strains and four Bacillus isolates inoculated individually into apple juice. Loading plots over the range of 1350 and 1700 cm-1 reflected the most distinctive biochemical features of Bacillus and Alicyclobacillus. Multivariate statistical methods (e.g. principal component analysis (PCA) and soft independent modeling of class analogy (SIMCA)) were used to analyze the spectral data. Distinctive separation of spectral samples was observed. This study demonstrates that FT-IR spectroscopy in combination with multivariate analysis could serve as a rapid and effective tool for fruit juice industry to differentiate between Bacillus and Alicyclobacillus and to distinguish between species belonging to these two genera.

Keywords: alicyclobacillus, bacillus, FT-IR, spectroscopy, PCA

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Authors: Abdelaziz Sheba Abdelrahman

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Compost enriched with actinomyces and Bacillus polymyxa algae as a Partial Substitute for Mineral N in Ewaise Mango Orchards Organic fertiliser, compost enriched with actinomyces, and the biofertilizer Bacillus polymyxa algae were used as a partial replacement for mineral N fertiliser in Ewaise mango orchards during the 2019 and 2020 seasons. When compared to using mineral N alone, the results showed that reducing the percentage of mineral N fertiliser from 100 to 50% and using compost enriched with actinomyces at 25 to 50% and Bacillus polymyxa had an announced promotion on leaf area, total chlorophylls, leaf N, P, and K, yield, and fruit quality. The use of compost enriched with actinomyces and Bacillus polymyxa, as well as mineral N, resulted in a significant decrease in nitrite in the pulp. Reducing mineral N to 25% of the suitable N had a negative impact on yield. The application of appropriate N via 50% inorganic N + compost enriched with actinomyces at 50% + Bacillus polymyxa algae increased yield quantitatively and qualitatively in Ewaise mango orchards. This promised treatment significantly reduced nitrite levels in the pulp fruit.

Keywords: bacillus polymyxa algae, fertiliser, biofertilizer, ewaise mango

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Authors: Han-Seul Lee, Min-Koo Kang, Sang-Ill Lee

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This study was conducted to reduce runoff by rainwater infiltration facility using attached growth with Bacillus sp., which are reported to remove nitrogen and phosphorus, as well as organic matter effectively. This study was investigated non-point pollutants removal efficiency of organic, nitrogen, and phosphorus in column using the media attached growth with Bacillus sp. To compare attached growth with bacillus sp. and detached media, two columns filled with perlite, zeolite, vermiculite, pumice, peat-moss was installed. In A column (attached growth with bacillus sp.), in case of infiltration velocity 30 mm/hr in high concentration of influent, it showed the removal efficiency (after aging term) is SS (suspended solid) 85.8±1.2 %, T-P (total phosphorus) 67.0±8.1 %, T-N (total nitrogen) 66.0±4.9 %, COD (chemical oxygen demand) 73.6±2.9 %, NH4+-N 72.7±3.0 %. In B column (detached media), in case of infiltration velocity 30 mm/hr in high concentration of influent, it showed the removal efficiency (after aging term) is SS 86.0±2.2 %, T-P 62.5±11.3 %, T-N 53.3±3.9 %, COD 34.6±3.7 %, NH4+-N 61.5±2.8 %. Removal efficiency of A column is better than B column. As the result from this study, using media with Bacillus sp. can improve an effective removal of non-point source pollutants.

Keywords: non-point source pollutants, Bacillus sp., rainwater, infiltration facility

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Authors: Jerry John T. M., Sylas V. P., Shijo Joy

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Hydrogen is the most essential gas that can be used for many purposes. During the production of hydrogen using raw materials like Soil and leftover cooked rice water (kanjivellam), the major by-product formed is water. Soil is collected from three different places in kottayam district: Kallara, Meenachilar, and Athirampuzha. Collected samples are mixed with rice water and tested for traces of hydrogen using a biohydrogen sensor after 72 hours. The result was the presence of hydrogen in all the 3 samples. After streaking, PCR and gel electrophoresis detected the bacteria which produced the hydrogen. RGCB Thiruvananthapuram conducted the sequencing of the PCR resultant. And identified the bacterial strains. Five variants of Bacillus bacteria ( (1) Bacillus cereus strain JTM GenBank: OP278839.1 (2) Bacillus toyonensis strain JTM2 GenBank: OP278841.1 (3) Bacillus anthracis strain JTM_SR2989-3-R_H08 GenBank: OP278960.1 (4) Bacillus thuringiensis strain JRY1 GenBank: OP278976.1 (5) Bacillus anthracis strain JTM_SR2989-3-F_H07 GenBank: OP278959.1 ) are identified and successfully registered in NCBI Gen bank. These Bacillus bacteria are major types of Rhizobacteria that can form spores and can survive in the soil for a long time period under harsh environmental conditions. Also, plant growth is enhanced by PGPR (Plant growth promoting rhizobacteria) through the induction of systemic resistance, antibiosis, and competitive omission. The molecular sequencing was submitted to the NCBI Gen Bank, and the accession numbers were allotted for the bacterial cultures.

Keywords: bio hydrogen production, bacterial bio hydrogen production, plant related to bacillus bacteria., bacillus bacteria study

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Authors: Ben Amar Cheba, Taha Ibrahim Zaghloul, Mohamad Hisham El-Massry, Ahmad Rafik El-Mahdy

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Seventy two bacterial strains with ability to degrade chitin were isolated during a screening program. One of the most potent isolates (strain R2) was identified as Bacillus sp. using conventional methods as well as 16S rRNA technique and submitted in the Gen Bank sequence database as Bacillus sp. R2 with a given accession number DQ 923161. This strain was able to produce high levels of extracellular chitinase. The chitinase of Bacillus sp. R2 hydrolyzed several chitinous substrates preferentially and showed a maximum activity toward the β chitin such as Calmar pen and squid bone chitins with the folds 1.47 and 1.23 respectively. The enzyme also exhibited a substrate binding capacity of more than 70% for squid chitin, shrimp shell colloidal chitin, chitosan and prawn shell chitin. The chitinase showed a moderate antifungal activity against many phytopathogenic fungi such as Aspergillus niger, A. flavus, Penicillium degitatum and Fusarium calmorum.This strain could be a suitable candidate for chitinase production on an industrial scale for using as promising antifungal biopestecide.

Keywords: antifungal activity, Bacillus sp. R2, chitinase, substrate specificity

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Authors: Adebowale T. Odeyemi, Oluwafemi A. Ajenifuja

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The leachate samples collected from Erifun dumpsite along Federal Polythenic road, Ado-Ekiti, Ekiti State, were subjected to bacteriological and mineral analyses. The bacteriological estimation and isolation were done using serial dilution and pour plating techniques. Antibiotic susceptibility test was done using agar disc diffusion technique. Atomic Absorption Spectophotometry method was used to analyze the heavy metal contents in the leachate samples. The bacterial and coliform counts ranged from 4.2 × 105 CFU/ml to 2.97 × 106 CFU/ml and 5.0 × 104 CFU/ml to 2.45 x 106 CFU/ml, respectively. The isolated bacteria and percentage of occurrence include Bacillus cereus (22%), Enterobacter aerogenes (18%), Staphylococcus aureus (16%), Proteus vulgaris (14%), Escherichia coli (14%), Bacillus licheniformis (12%) and Klebsiella aerogenes (4%). The mineral value ranged as follow; iron (21.30mg/L - 25.60mg/L), zinc (1.80mg/L - 5.60mg/L), copper (1.00mg/L - 2.60mg/L), chromium (0.50mg/L - 1.30mg/L), candium (0.20mg/L - 1.30mg/L), nickel (0.20mg/L - 0.80mg/L), lead (0.05mg/L-0.30mg/L), cobalt (0.03mg/L - 0.30mg/L) and in all samples manganese was not detected. The entire organisms isolated exhibited a high level of resistance to most of the antibiotics used. There is an urgent need for awareness to be created about the present situation of the leachate in Erifun, on the need for treatment of the nearby stream and other water sources before they can be used for drinking and other domestic use. In conclusion, a good method of waste disposal is required in those communities to prevent leachate formation, percolation, and runoff into water bodies during the raining season.

Keywords: antibiotic susceptibility, dumpsite, bacteriological analysis, heavy metal

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Authors: S. Hannah Elizabeth, A. Panneerselvam

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Objective: The main objective was to evaluate the degradative properties of Bacillus cereus sp TUHP2 isolated from TNT-Polluted soils in the Vellore District, Tamil Nadu, India. Methods: Among the 3 bacterial genera isolated from different soil samples, one potent TNT degrading strain Bacillus cereus sp TUHP2 was identified. The morphological, physiological and the biochemical properties of the strain Bacillus cereus sp TUHP2 was confirmed by conventional methods and genotypic characterization was carried out using 16S r-DNA partial gene amplification and sequencing. The broken down by products of DNT in the extract was determined by Gas Chromatogram- Mass spectrometry (GC-MS). Supernatant samples from the broth studied at 24 h interval were analyzed by HPLC analysis and the effect on various nutritional and environmental factors were analysed and optimized for the isolate. Results: Out of three isolates one strain TUHP2 were found to have potent efficiency to degrade TNT and revealed the genus Bacillus. 16S rDNA gene sequence analysis showed highest homology (98%) with Bacillus cereus and was assigned as Bacillus cereus sp TUHP2. Based on the energy of the predicted models, the secondary structure predicted by MFE showed the more stable structure with a minimum energy. Products of TNT Transformation showed colour change in the medium during cultivation. TNT derivates such as 2HADNT and 4HADNT were detected by HPLC chromatogram and 2ADNT, 4ADNT by GC/MS analysis. Conclusion: Hence this study presents the clear evidence for the biodegradation process of TNT by strain Bacillus cereus sp TUHP2.

Keywords: bioremediation, biodegradation, biotransformation, sequencing

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Authors: Najat El-Kurdi, Sherif Hammad, Mohamed Ghazi, Sahar El-Shatoury, Khaled Zakaria

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The increasing number of plastic production and its importance to humanity in daily life made it a headache to the planet earth. The persistence of plastic wastes in the environment formed a serious problem. They are prominent with their capability to resist microbial degradation for decades. Thus, it was crucial to find ways to eliminate the plastics without depending on conventional recycling methods, which causes the formation of more hazardous compounds and doubles the problem. In this paper, mealworms were fed with a mixture of plastic wastes such as plastic bags, Styrofoam, PE foam, and plastic tarpaulins film as the sole food source for a month. Frass was collected at the end of the test and examined using FTIR analysis. Also, the gut bacteria were isolated and identified using 16S rRNA. The results show the mineralization of plastic in the frass of plastic-fed worms when compared to control. The 16S rRNA and the BLAST analysis showed that the obtained isolate belongs to the genus Bacillus Sp especially Bacillus subtilis. Phylogenetic analysis showed their relatedness to the other Bacillus species in the NCBI database.

Keywords: mealworm, waste management, plastic-degrading bacteria, gut microbiome, Bacillus sp

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Authors: Sugandha Asthana, Geetika Vajpayee, Pratibha Kumari, Shanthy Sundaram

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An economically important disease of wheat in North Western region of India is Karnal Bunt caused by smut fungus Tilletia indica. This fungal pathogen spreads by air, soil and seed borne sporodia at the time of flowering, which ultimately leads to partial bunting of wheat kernels with fishy odor and taste to wheat flour. It has very serious effects due to quarantine measures which have to be applied for grain exports. Chemical fungicides such as mercurial compounds and Propiconazole applied to the control of Karnal bunt have been only partially successful. Considering the harmful effects of chemical fungicides on man as well as environment, many countries are developing biological control as the superior substitute to chemical control. Repeated use of fungicides can be responsible for the development of resistance in fungal pathogens against certain chemical compounds. The present investigation is based on the isolation and evaluation of antifungal properties of some isolated (from natural manure) and commercial bacterial strains against Tilletia indica. Total 23 bacterial isolates were obtained and antagonistic activity of all isolates and commercial bacterial strains (Bacillus subtilis MTCC8601, Bacillus pumilus MTCC 8743, Pseudomonas aeruginosa) were tested against T. indica by dual culture plate assay (pour plate and streak plate). Test for the production of antifungal volatile organic compounds (VOCs) by antagonistic bacteria was done by sealed plate method. Amongst all s1, s3, s5, and B. subtilis showed more than 80% inhibition. Production of extracellular hydrolytic enzymes such as protease, beta 1, 4 glucanase, HCN and ammonia was studied for confirmation of antifungal activity. s1, s3, s5 and B. subtilis were found to be the best for protease activity and s5 and B. subtilis for beta 1, 4 glucanase activity. Bacillus subtilis was significantly effective for HCN whereas s3, s5 and Bacillus subtilis for ammonia production. Isolates were identified as Pseudomonas aeruginosa (s1) and B. licheniformis (s3, s5) by various biochemical assays and confirmed by16s rRNA sequencing. Use of microorganisms or their secretions as biocontrol agents to avoid plant diseases is ecologically safe and may offer long term of protection to crop. The above study reports the promising effects of these strains in better pathogen free crop production and quality maintenance as well as prevention of the excessive use of synthetic fungicides.

Keywords: antagonistic, antifungal, biocontrol, Karnal bunt

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Authors: Bassam O AlJohny

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The growth of Bacillus aquimaris was inhibited from 6 - 20 % of NaCl but it showed some tolerance when Diatomaceous earth (DE) added from 2 - 12% NaCl. Concerning the effect of NaCl on polyol production, we can conclude that, the test bacterium showed some tolerance to NaCl by producing glycerol up to 8 % of NaCl. Then decreased sharply. The addition of DE decrease the amount of polyol and glycerol remarkably and this due to the productive effect of DE to the bacterial cells. The SEM figures represented the presence of electron dense bodies due to the accumulation of small particles of DE as protective molecules.

Keywords: Bacillus aquimaris, Diatomaceous earth (DE), osmoticstress, sodium chloride

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Authors: Narasimhulu Korrapati

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The objective of this present study is to optimize the process parameters for batch biosorption of Cr(VI) and Cu(II) ions by Bacillus subtilis using Response Surface Methodology (RSM). Batch biosorption studies were conducted under optimum pH, temperature, biomass concentration and contact time for the removal of Cr(VI) and Cu(II) ions using Bacillus subtilis. From the studies it is noticed that the maximum biosorption of Cr(VI) and Cu(II) was by Bacillus subtilis at optimum conditions of contact time of 30 minutes, pH of 4.0, biomass concentration of 2.0 mg/mL, the temperature of 32°C in batch biosorption studies. Predicted percent biosorption of the selected heavy metal ions by the design expert software is in agreement with experimental results of percent biosorption. The percent biosorption of Cr(VI) and Cu(II) in batch studies is 80% and 78.4%, respectively.

Keywords: heavy metal ions, response surface methodology, biosorption, wastewater

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Authors: Samira Ferhat, Redha Alouaoui, Leila Trifi, Abdelmalek Badis

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The objective of this work is the use of natural surfactant (biosurfactant) and synthetic (sodium dodecyl sulfate and tween 80) for environmental application. In fact the solubility of the polycyclic hydrocarbon (naphthalene) and the desorption of the heavy metals in the presence of surfactants. The microorganisms selected in this work are bacterial strain (Bacillus licheniformis) for the production of biosurfactant for use in this study. In the first part of this study, we evaluated the effectiveness of surfactants solubilization certain hydrocarbons few soluble in water such as polyaromatic (case naphthalene). Tests have shown that from the critical micelle concentration, decontamination is performed. The second part presents the results on the desorption of heavy metals (for copper) by the three surfactants, using concentrations above the critical micelle concentration. The comparison between the desorption of copper by the three surfactants, it is shown that the biosurfactant is more effective than tween 80 and sodium dodecyl sulfate.

Keywords: surfactants, biosurfactant, naphthalene, copper, critical micelle concentration, solubilization, desorption

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Authors: Sarisha Devnath, Oluwatosin A. Ijabadeniyi

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In recent years milk contamination has become a major problem in households; due to the likely occurrence of bacteria, even after the milk has been processed. One such genus of bacteria causing unwanted growth is Bacillus. This research project looks at the presence of spore formers in processed milk from household refrigerators and the effect of pasteurization and high temperature on Bacillus spores activation. 24 samples each of UHT milk and pasteurised milk from 24 households were sampled for the presence of spore formers. While anaerobic spore formers were not found in any of the samples, the average aerobic spore formers in UHT milk and pasteurized milk however were 5.77 cfu/ml and 5.88 cfu/ml respectively. After sequencing, it was detected that the mixed culture contained Bacillus cereus, for both pasteurised and UHT milk samples. For the activation study, raw milk samples were collected and subjected to four different temperatures; 65˚C, 72˚C, 80˚C, 100˚C respectively. Samples were stored for 7 days at 5˚C and 10˚C and analysed daily. The average aerobic spore formers in raw milk for samples stored at 5˚C range between 4.67-6.00 cfu/ml while it ranges between 4.84-6.00 cfu/ml at 10˚C, signifying that the high temperatures could have resulted in germination of dominant spores. Statistical analysis conducted on these results indicated a significant difference between the numbers of colonies present at the different treatment temperatures the bacterium was exposed to. This work showed that household milk may constitute public health risk furthermore; pasteurization and higher temperatures may not be effective to remove aerobic spore formers because of Bacillus spores activation.

Keywords: sporeformers, bacillus, spores, activation, milk

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Authors: Stephy Saavedra, Annsy C. Arredondo, Gisele Monteiro, Adalberto Pessoa Jr, Carol N. Flores-Fernandez, Amparo I. Zavaleta

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L-asparaginase from bacterial sources is used in leukemic treatment and food industry. This enzyme is classified based on its affinity towards L-asparagine and L-glutamine. Likewise, ansZ genes express L-asparaginase with higher affinity to L-asparagine. The aim of this work was to clone and express of ansZ gene from Bacillus sp. CH11 isolated from Chilca salterns in Peru. The gene encoding L-asparaginase was cloned into pET15b vector and transformed in Escherichia coli BL21 (DE3) pLysS. The expression was carried out in a batch culture using LB broth and 0.5 mM IPTG. The recombinant L-asparaginase showed a molecular weight of ~ 39 kDa by SDS PAGE and a specific activity of 3.19 IU/mg of protein. The cloning and expression of ansZ gene from this halotolerant Bacillus sp. CH11 allowed having a biological input to improve a future scaling-up.

Keywords: ansZ gene, Bacillus sp, Chilca salterns, recombinant L-asparaginase

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Authors: Saleh H. Salmen, Sulaiman Ali Alharbi, Hany M. Yehia, Mohammad A. Khiyami, Milton Wainwright, Naiyf S. Alharbi, Arunachalam Chinnathambi

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An antifungal protein synthesized by Bacillus cereus has been partially purified by the use of ammonium sulfate precipitation and Sephadex-G-200 column chromatography. The protein was produced from Bacillus cereus grown in potato Dextrose Broth Medium (PDB) at 30 ºC for 3 days at 100 rpm. The protein showed antagonistic effect against some fungi and yeasts. Crude extract from medium and semi-purified protein were tested in vitro against both fungi and yeasts using the disc diffusion method in order to detect the inhibitory effect of the protein. Zones of inhibition of the following diameter were found (mm) were Alternaria alternate (28), Rhodotorula glutinis (20), Fusarium sp. (16), Rhizopus sp. (15), Penicillium digitatum (13), Mucor sp. (13) and Aspergillus niger (10). The isolated protein was found to have a molecular weight of ~35kDa by sodium deodecyl sulfate-poly acrylamide gel electrophoresis. The data showed that the protein of Bacillus cereus has antifungal activity, a fact which points to the possibility of using it as a bio-control agent against some fungi, findings which emphasize the potential role of B. cereus as an important bio-control agent.

Keywords: bacillus cereus, ~35kDa protein, molds, yeasts

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Authors: Roni Rachel Mendelson, Caileigh Pudela

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Bacillus cereus is a toxin-producing, facultatively anaerobic gram-positive bacterium that is widely distributed environmentally. It can quickly multiply at room temperature with an abundantly present preformed toxin. When ingested, this toxin can cause gastrointestinal illness, which is the commonly known manifestation of the disease. Bacillus cereus sepsis is a disease that is mostly concerning in the population of the immunocompromised patients. One of them is acute lymphoblastic leukemia’s patients during induction. Pediatric acute lymphoblastic leukemia is a common pediatric hematologic malignancy. It is characterized by the rapid proliferation of poorly differentiated lymphoid progenitor cells inside the bone marrow. We present here a 21-month-old boy undergoing induction chemotherapy for acute lymphoblastic leukemia who developed bacillus sepsis bacteremia and, as a result, multi organ failure leading to seizures and multiple strokes. Our case report highlights the extensive overall and neurological damage that can be caused because of bacillus cereus bacteremia, which can lead to higher mortality rate and decreased in survivorship in a highly curable disease. It is very subtle and difficult to recognize and appears to be deteriorating extremely fast. There should be a low threshold for work up and empiric coverage for neutropenic patients during acute lymphoblastic leukemia induction therapy.

Keywords: acute lymphoblastic leukemia, bacillus cereus, immunocompromised, sepsis

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