Search results for: Pseudomonas putida

Authors: O. Antypenko, I. Vasilieva, S. Kovalenko

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Investigations for new effective and less toxic antimicrobials agents are always up-to-date. The tetrazole derivatives are quite interesting objects as for synthesis as well as for pharmacological screening. Thus, some derivatives of tetrazole demonstrated antimicrobial activity, namely 5-phenyl-tetrazolo[1,5-c]quinazoline was effective one against Staphylococcus aureus and Esherichia faecalis (MIC = 250 mg/L). Besides, investigation of the 9-bromo(chloro)-5-morpholin(piperidine)-4-yl-tetrazolo[1,5-c]quinazoline’s antimicrobial activity against Esherichia coli and Enterococcus faecalis, Pseudomonas aeruginosa and Staphylococcus aureus revealed that sensitivity of Gram-positive bacteria to the compounds was higher than that of Gram-negative bacteria. So, our previously synthesized, 31 derivatives of 1-(2-(1H-tetrazol-5-yl)-R1-phenyl)-3-R2-phenyl(ethyl)ureas were decided to test for their in vitro antibacterial activity against Gram-positive bacteria (Staphylococcus aureus ATCC 25923, Enterobacter aerogenes, Enterococcus faecalis ATCC 29212), Gram-negative bacteria (Pseudomonas aeruginosa ATCC 9027, Escherichia coli ATCC 25922, Klebsiella pneumoniae 68) and antifungal properties against Candida albicans ATCC 885653. Agar-diffusion method was used for determination of the preliminary activity compared to well-known reference antimicrobials. All the compounds were dissolved in DMSO at a concentration of 100 μg/disk, using inhibition zone diameter (IZD, mm) as a measure for the antimicrobial activity. The most active turned to be 3 structures, that inhibited several bacterial strains: 1-ethyl-3-(5-fluoro-2-(1H-tetrazol-5-yl)phenyl)urea (1), 1-(4-bromo-2-(1H-tetrazol-5-yl)-phenyl)-3-(4-(trifluoromethyl)phenyl)urea (2) and 1-(4-chloro-2-(1H-tetrazol-5-yl)phenyl)-3-(3-(trifluoromethyl)phenyl)urea (3). IZM (mm) was 40 (Escherichia coli), 25 (Klebsiella pneumonia) for compound 1; 12 (Pseudomonas aeruginosa), 15 (Staphylococcus aureus), 10 (Enterococcus faecalis) for compound 2; 25 (Staphylococcus aureus), 15 (Enterococcus faecalis) for compound 3. The most sensitive to the activity of the substances were Gram-negative bacteria Pseudomonas aeruginosa. While none of compound effected on Candida albicans. Speaking about, reference drugs: Amikacin (30 µg/disk) showed 27 and Ceftazide (30 µg/disk) 25 against Pseudomonas aeruginosa. That is, unfortunately, higher than studied 1-(2-(1H-tetrazol-5-yl)-R1-phenyl)-3-R2-phenyl(ethyl)ureas. Obtained results will be used for further purposeful optimization of the leading compounds in the more effective antimicrobials because of the ever-mounting problem of microorganism’s resistance.

Keywords: antimicrobial, antifungal, compounds, 1-(2-(1H-tetrazol-5-yl)-R1-phenyl)-3-R2-phenyl(ethyl)ureas

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Authors: Harsha Thaira, Ritu Raval, Keyur Raval

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Melanin has immense applications in the field of agriculture, cosmetics and pharmaceutical industries due to its photo-protective, UV protective and anti- oxidant activities. However, its production is limited to costly chemical methods or harsh extractive methods from hair which ultimately gives poor yields. This makes the cost of melanin very high, to the extent of US Dollar 300 per gram. Some microorganisms are reported to produce melanin under stress conditions. Out of all melanin producing organisms, Pseudomonas stutzeri can grow in sea water and produce melanin under saline stress. The objective of this study was to develop a sea water based bioprocess. Effects of different growth media and process parameters on melanin production using sea water were investigated. The marine bacterial strain Pseudomonas stutzeri HMGM-7(MTCC 11712) was selected and the effect of different media such as Nutrient Broth (NB), Luria Bertini (LB) broth, Bushnell- Haas broth (BHB) and Trypticase Soy broth (TSB) and various medium components were investigated with one factor at a time approach. Parameters like shaking frequency, inoculum age, inoculum size, pH and temperature were also investigated in order to obtain the optimum conditions for maximum melanin production. The highest yield of melanin concentration, 0.306 g/L, was obtained in Trypticase Soy broth at 36 hours. The yield was 1.88 times higher than the melanin obtained before optimization, 0.163 g/L at 36 hours. Studies are underway to optimize medium constituents to further enhance melanin production.

Keywords: melanin, marine, bioprocess, pseudomonas

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Authors: Alisha Akya, Afsaneh salami

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Background and Objective: Pseudomonas aeruginosa is an opportunistic pathogen associated with nosocomial infections. One of the major concerns for the treatment of P. aeruginosa infections is its resistant to a variety of antibiotics. The purpose of this study was to assess the dissemination of p. aeruginosa isolates obtained from major hospitals in Kermanshah, west of Iran. Materials and Methods: Antibiotic susceptibility testing was performed using the minimal inhibitory concentrations. Mettalo-beta-lactamase was investigated using the double disk diffusion (DDST) test and PCR. Molecular typing was performed by pulsed-field gel electrophoresis (PFGE). Results: The 60 P. aeruginosa isolates, 30 (50%) were resistant to gentamicin, 38 (63/3%) to piperacilin, 42 (70%) to ceftazidime, and 45 (75%) to cefepime. Twenty-nine (48/3%) isolates were MBLs producer based on the DDST test. Five (8/3%) isolates were positive for VIM gene and 4 of them were from burn specimens. PFGE analysis among MBLs producers revealed 12 distinct genotype patterns. A pattern covering the highest number of strains was determined as the dominant clone. Conclusions: Our study showed that P. aeruginosa strains can be spread between patients in hospitals or acquired from different environmental sources. P. aeruginosa isolates were highly resistant to antibiotics and, therefore, the susceptibility of isolates to antibiotics should be tested before treatment. Given the clinical significance of MBLs producing isolates, identification of these organisms is essential in the hospitals in order to get a better therapeutic response and control of bacterial dissemination.

Keywords: clonal dissemination, mettalo-beta-lactamase, Pseudomonas aeruginosa, PFGE

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Authors: Solomon Gebremariam, Mulugeta Naizigi, Aregawi Haileselassie

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Background: Knowledge of the pattern of bacterial isolates and their antimicrobial susceptibility is crucial for guiding empirical treatment and infection prevention and control measures. Objective: The aim of this study was to analyze the pattern of bacterial isolates and their susceptibility patterns from various specimens. Methods: Retrospectively, a total of 1067 microbiological culture results that were isolated, characterized, and identified by standard microbiological methods and whose antibiotic susceptibility was determined using CLSI guidelines between 2017 and 2019 were retrieved and analyzed. Data were entered and analyzed using the Stata release 10.1 statistical package. Result: The positivity rate of culture was 26.04% (419/1609). The most common bacteria isolated were S. aureus 23.8% (94), E. coli 15.1% (60), Klebsiella pneumonia 14.1% (56), Pseudomonas aeruginosa 8.5% (34), and CONS 7.3% (29). S. aureus and CONS showed a high (58.1% - 96.2%) rate of resistance to most antibiotics tested. They were less resistant to Vancomycin which is 18.6% (13/70) and 11.8% (2/17), respectively. Similarly, the resistance of E. coli, Klebsella pneumonia, and Pseudomonas aeruginosa was high (69.4% - 100%) to most antibiotics. They were less resistant to Ciprofloxacilin, which is 41.1% (23/56), 19.2% (10/52), and 16.1% (5/31), respectively. Conclusion: This study has shown that there is a high rate of antibiotic resistance among bacterial isolates in this hospital. A combination of Vancomycin and Ciprofloxacin should be considered in the choice of antibiotics for empirical treatment of suspected infections due to S. aureus, CONS, E. coli, Klebsiella pneumonia, Pseudomonas such as in infections within hospital setup.

Keywords: antimicrobial, resistance, bacteria, hospital

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Authors: Mostafa M. Abo Elsoud, Heba I. Elkhouly, Nagwa M. Sidkey

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Production of rhamnolipids by Pseudomonas aeruginosa has attracted a growing interest during the last few decades due to its high productivity compared with other microorganisms. In the current work, rhamnolipids production by P. aeruginosa PAO1 was statistically modeled using Taguchi orthogonal array, numerically optimized and validated. Prickly Pear Peel (Opuntia ficus-indica) has been used as a carbon source for production of rhamnolipid. Finally, the optimum conditions for rhamnolipid production were applied in 5L working volume bioreactors at different aerations, agitation and controlled pH for maximum rhamnolipid production. In addition, kinetic studies of rhamnolipids production have been reported. At the end of the batch bioreactor optimization process, rhamnolipids production by P. aeruginosa PAO1 has reached the worldwide levels and can be applied for its industrial production.

Keywords: rhamnolipids, pseudomonas aeruginosa, statistical optimization, tagushi, opuntia ficus-indica

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Authors: Zonglin Yang, Tatsuya Akiyama, Kerry S. Williamson, Michael J. Franklin, Thiruvarangan Ramaraj

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Pseudomonas aeruginosa is an opportunistic pathogen that forms surface-associated microbial communities (biofilms) on artificial implant devices and on human tissue. Biofilm infections are difficult to treat with antibiotics, in part, because the bacteria in biofilms are physiologically heterogeneous. One measure of biological heterogeneity in a population of cells is to quantify the cellular concentrations of ribosomes, which can be probed with fluorescently labeled nucleic acids. The fluorescent signal intensity following fluorescence in situ hybridization (FISH) analysis correlates to the cellular level of ribosomes. The goals here are to provide computationally and statistically robust approaches to automatically quantify cellular heterogeneity in biofilms from a large library of epifluorescent microscopy FISH images. In this work, the initial steps were developed toward these goals by developing an automated biofilm detection approach for use with FISH images. The approach allows rapid identification of biofilm regions from FISH images that are counterstained with fluorescent dyes. This methodology provides advances over other computational methods, allowing subtraction of spurious signals and non-biological fluorescent substrata. This method will be a robust and user-friendly approach which will enable users to semi-automatically detect biofilm boundaries and extract intensity values from fluorescent images for quantitative analysis of biofilm heterogeneity.

Keywords: image informatics, Pseudomonas aeruginosa, biofilm, FISH, computer vision, data visualization

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Authors: Omar Alhamd, Peter A. Thomas, Trevor J. Greenhough, Annette K. Shrive

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The Pseudomonas syringae species complex includes many plant pathogenic strains with highly specific interactions with varied host species and cultivars. The rapid spread of these bacteria over the last ten years has become a cause for concern. Nanoparticles have previously shown promise in microbiological action. We have therefore investigated in vitro and in vivo the effects of different types and sizes of nanoparticles in order to provide quantitative information about their effect on the bacteria. The effects of several different nanoparticles against several bacteria strains were investigated. The effect of NP on bacterial growth was studied by measuring the optical density, biochemical and nutritional tests, and transmission electron microscopy (TEM) to determine the shape and size of NP. Our results indicate that their effects varied, with either a negative or a positive impact on both bacterial and plant growth. Additionally, the methods of exposure to nanoparticles have a crucial role in accumulation, translocation, growth response and bacterial growth. The results of our studies on the behaviour and effects of nanoparticles in model plants showed. Cerium oxide (CeO₂) and silver (Ag) NP showed significant antibacterial activity against several pathogenic bacteria. It was found that titanium nanoparticles (TiO₂) can have either a negative or a positive impact, according to concentration and size. It is also thought that environmental conditions can have a major influence on bacterial growth. Studies were therefore also carried out under some environmental stress conditions to test bacterial survival and to assess bacterial virulence. All results will be presented including information about the effects of different nanoparticles on Pseudomonas syringae bacteria.

Keywords: plant microbiome, nanoparticles, 16S rRNA gene sequencing, bacterial survival

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Authors: Togzhan D. Mukasheva, Anel A. Omirbekova, Raikhan S. Sydykbekova, Ramza Zh. Berzhanova, Lyudmila V. Ignatova

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Following plants-barley (Hordeum sativum), alfalfa (Medicago sativa), grass mixture (red fescue-75%, long-term ryegrass - 20% Kentucky bluegrass - 10%), oilseed rape (Brassica napus biennis), resistant to growth in the contaminated soil with oil content of 15.8 g / kg 25.9 g / kg soil were used. Analysis of the population showed that the oil pollution reduces the number of bacteria in the rhizosphere and rhizoplane of plants and enhances the amount of spore-forming bacteria and saprotrophic micromycetes. It was shown that regardless of the plant, dominance of Pseudomonas and Bacillus genera bacteria was typical for the rhizosphere and rhizoplane of plants. The frequency of bacteria of these genera was more than 60%. Oil pollution changes the ratio of occurrence of various types of bacteria in the rhizosphere and rhizoplane of plants. Besides the Pseudomonas and Bacillus genera, in the presence of hydrocarbons in the root zone of plants dominant and most typical were the representatives of the Mycobacterium and Rhodococcus genera. Together the number was between 62% to 72%.

Keywords: pollution, root system, micromycetes, identification

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

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Pseudomonas species possess a variety of promising properties of antifungal and growth promoting activities in the wheat plant. In the present study, Pseudomonas fluorescens MTCC-9768 is tested against plant pathogenic fungus Tilletia indica, causing Karnal bunt, a quarantine disease of wheat (Triticum aestivum) affecting kernels of wheat. It is one of the 1/A1 harmful diseases of wheat worldwide under EU legislation. This disease develops in the growth phase by the spreading of microscopically small spores of the fungus (teliospores) being dispersed by the wind. The present chemical fungicidal treatments were reported to reduce teliospores germination, but its effect is questionable since T. indica can survive up to four years in the soil. The fungal growth inhibition tests were performed using Dual Culture Technique, and the results showed inhibition by 82.5%. The interaction of antagonist bacteria-fungus causes changes in the morphology of hyphae, which was observed using Lactophenol cotton blue staining and Scanning Electron Microscopy (SEM). The rounded and swollen ends, called ‘theca’ were observed in interacted fungus as compared to control fungus (without bacterial interaction). This bacterium was tested for its antagonistic activity like protease, cellulose, HCN production, Chitinase, etc. The growth promoting activities showed increase production of IAA in bacteria. The bacterial secondary metabolites were extracted in different solvents for testing its growth inhibiting properties. The characterization and purification of the antifungal compound were done by Thin Layer Chromatography, and Rf value was calculated (Rf value = 0.54) and compared to the standard antifungal compound, 2, 4 DAPG (Rf value = 0.54). Further, the in vivo experiments showed a significant decrease in the severity of disease in the wheat plant due to direct injection method and seed treatment. Our results indicate that the extracted and purified compound from the antagonist bacteria, P. fluorescens MTCC-9768 may be used as a potential biocontrol agent against T. indica. This also concludes that the PGPR properties of the bacteria may be utilized by incorporating it into bio-fertilizers.

Keywords: antagonism, Karnal bunt, PGPR, Pseudomonas fluorescens

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Authors: Nastaran Soltani

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Heavy metals such as Mercury are toxic elements that enter the environment through different ways and endanger the environment, plants, animals, and humans’ health. Microbial activities reduce the amount of heavy metals. Therefore, an effective mechanism to eliminate heavy metals in the nature and factory slops, is using bacteria living in polluted areas. Karun River in Khuzestan Province in Iran has been always polluted by heavy metals as it is located among different industries in the region. This study was performed based on the data from sampling water and sediments of four stations across the river during the four seasons of a year. The isolation of resistant bacteria was performed through enrichment and direct cultivation in a solid medium containing mercury. Various bacteria such as Pseudomonas sp., Serratia Marcescens, and E.coli were identified as mercury-resistant bacteria. The power of these bacteria to remove mercury varied from 28% to 86%, with strongest power belonging to Pseudomonas sp. isolated in spring making a good candidate to be used for mercury biological removal from factory slops.

Keywords: bacteria, Karun River, mercury, biological removal, mercury-resistant

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Authors: V. Hakimzadeh, M. Noori, M. maleki

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The aim of this study was to determine the composition and antimicrobial effect of Mentha piperita essential oil in "in-vitro" condition. The chemical composition of the essential oil obtained by hydro-distillation was examined by GC/MS and the antimicrobial effect was studied on the growth of seven microbial species including Bacillus cereus, Pseudomonas aeruginosa and Proteus vulgaris using micro-dilution method. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined. Chemical composition analysis identified a total of 28 compounds in which the main components were menthol (32%), mentone (13.4), menthyl acetate (12%), 1,8-cineole (8.2%) and neomenthol (4%) representing 69.6 % of the total oil. Other separated components accounted for less than 30.4% of the oil. Results of antimicrobial analysis showed that the MIC values for Bacillus cereus, Pseudomonas aeruginosa and Proteus vulgaris was respectively 50, 200 and 100 µg/ml and the MBC was determined at 200, 400 and 200 µg/ml respectively. The results of the present study indicated that Mentha piperita essential oil had significant antimicrobial activity.

Keywords: antimicrobial activity, essential oil composition, Mentha piperita

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Authors: C. S. Jeba Samuel

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Soil, especially from oil field has posed a great hazard for terrestrial and marine ecosystems. The traditional treatment of oil contaminated soil cannot degrade the crude oil completely. So far, biodegradation proves to be an efficient method. During biodegradation, crude oil is used as the carbon source and addition of nitrogenous compounds increases the microbial growth, resulting in the effective breakdown of crude oil components to low molecular weight components. The present study was carried out to evaluate the biodegradation of crude oil by hydrocarbon-degrading microorganism Pseudomonas aeruginosa isolated from natural environment like oil contaminated soil. Pseudomonas aeruginosa, an oil degrading microorganism also called as hydrocarbon utilizing microorganism (or “HUM” bug) can utilize crude oil as sole carbon source. In this study, the biodegradation of crude oil was conducted with modified mineral basal salt medium and nitrogen sources so as to increase the degradation. The efficacy of the plasmid from the isolated strain was incorporated into E.coli DH5 α host to speed up the degradation of oil. The usage of molecular techniques has increased oil degradation which was confirmed by the degradation of aromatic and aliphatic rings of hydrocarbons and was inferred by the lesser number of peaks in Fourier Transform Infrared Spectroscopy (FTIR). The gas chromatogram again confirms better degradation by transformed cells by the lesser number of components obtained in the oil treated with transformed cells. This study demonstrated the technical feasibility of using direct inoculation of transformed cells onto the oil contaminated region thereby leading to the achievement of better oil degradation in a shorter time than the degradation caused by the wild strain.

Keywords: biodegradation, aromatic rings, plasmid, hydrocarbon, Fourier Transform Infrared Spectroscopy (FTIR)

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Authors: Mohammad Aladwan, Adelia Skripova

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Bioremediation aspects of crude oil-polluted fields can be achieved by isolation and identification of bacterial species from oil-contaminated soil in order to choose the most active isolates and increase the strength of others. In this study, oil-degrading bacteria were isolated and identified from oil-contaminated soil samples in northeastern Jordan. The bacterial growth count (CFU/g) was between 1.06×10⁵ and 0.75×10⁹. Eighty-two bacterial isolates were characterized by their morphology and biochemical tests. The identified bacterial genera included: Klebsiella, Staphylococcus, Citrobacter, Lactobacillus, Alcaligenes, Pseudomonas, Hafnia, Micrococcus, Rhodococcus, Serratia, Enterobacter, Bacillus, Salmonella, Mycobacterium, Corynebacterium, and Acetobacter. Molecular identification of a universal primer 16S rDNA gene was used to identify four bacterial isolates: Microbacterium esteraromaticum strain L20, Pseudomonas stutzeri strain 13636M, Klebsilla pneumoniae, and uncultured Klebsilla sp., known as new strains. Our results indicate that their specific oil-degrading bacteria isolates might have a high strength of oil degradation from oil-contaminated sites. Staphylococcus intermedius (75%), Corynebacterium xerosis (75%), and Pseudomonas fluorescens (50%) showed a high growth rate on different types of hydrocarbons, such as crude oil, toluene, naphthalene, and hexane. In addition, monooxygenase and catechol 2,3-dioxygenase were detected in 17 bacterial isolates, indicating their superior hydrocarbon degradation potential. Total petroleum hydrocarbons were analyzed using gas chromatography for soil samples. Soil samples M5, M7, and M8 showed the highest levels (43,645, 47,805, and 45,991 ppm, respectively), and M4 had the lowest level (7,514 ppm). All soil samples were analyzed for heavy metal contamination (Cu, Cd, Mn, Zn, and Pb). Site M7 contains the highest levels of Cu, Mn, and Pb, while Site M8 contains the highest levels of Mn and Zn. In the future, these isolates of bacteria can be used for the cleanup of oil-contaminated soil.

Keywords: bioremediation, 16S rDNA gene, oil-degrading bacteria, hydrocarbons

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Authors: Omar Pillaca-Pullo, Arturo Alejandro-Paredes, Carol Flores-Fernandez, Marijuly Sayuri Kina, Amparo Iris Zavaleta

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Aqueous two-phase system (ATPS) is an interesting alternative for separating industrial enzymes due to it is easy to scale-up and low cost. Polyethylene glycol (PEG) mixed with potassium phosphate or magnesium sulfate is one of the most frequently polymer/salt ATPS used, but the consequences of its use is a high concentration of phosphates and sulfates in wastewater causing environmental issues. Citrate could replace these inorganic salts due to it is biodegradable and does not produce toxic compounds. On the other hand, statistical design of experiments is widely used for ATPS optimization and it allows to study the effects of the involved variables in the purification, and to estimate their significant effects on selected responses and interactions. The 24 factorial design with four central points (20 experiments) was employed to study the partition and purification of proteases produced by Pseudomonas sp. in PEG/citrate ATPS system. ATPS was prepared with different sodium citrate concentrations [14, 16 and 18% (w/w)], pH values (7, 8 and 9), PEG molecular weight (2,000; 4,000 and 6,000 g/mol) and PEG concentrations [18, 20 and 22 % (w/w)]. All system components were mixed with 15% (w/w) of the fermented broth and deionized water was added to a final weight of 12.5 g. Then, the systems were mixed and kept at room temperature until to reach two-phases separation. Volumes of the top and bottom phases were measured, and aliquots from both phases were collected for subsequent proteolytic activity and total protein determination. Influence of variables such as PEG molar mass (MPEG), PEG concentration (CPEG), citrate concentration (CSal) and pH were evaluated on the following responses: purification factor (PF), activity yield (Y), partition coefficient (K) and selectivity (S). STATISTICA program version 10 was used for the analysis. According to the obtained results, higher levels of CPEG and MPEG had a positive effect on extraction, while pH did not influence on the process. On the other hand, the CSal could be related with low values of Y because of the citrate ions have a negative effect on solubility and enzymatic structure. The optimum values of Y (66.4 %), PF (1.8), K (5.5) and S (4.3) were obtained at CSal (18%), MPEG (6,000 g/mol), CPEG (22%) and pH 9. These results indicated that the PEG/citrate system is accurate to purify these Pseudomonas sp. proteases from fermented broth as a first purification step.

Keywords: citrate, polyethylene glycol, protease, Pseudomonas sp

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Authors: Fatma M. Benrabha

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The aim of the study was to determine the type and pattern of antibiotic susceptibility of the pathogenic microorganisms causing chronic suppurative otitis media (CSOM), which could lead to better therapeutic decisions and consequently avoidance of appearance of resistance to specific antibiotics. Most frequently isolated agents were Pseudomonas aeruginosa 28.5%; followed by Staphylococcus aureus 18.2%; proteus mirabilis 13.9%; Providencia stuartti 6.7%; Bacteroides melaninogenicus, Aspergillus sp., candida sp., 4.2% each; and other microorganisms were represented in 3-0.2%. Drug sensitivities pattern of Pseudomonas aeruginosa showed that ciprofloxacin was active against the majority of isolates (93.9%) followed by ceftazidime 86.2%, amikacin 76.2% and gentamicin 40.8%. However, Staphylococcus aureus isolates were resistant to penicillin 72.7%, erythromycin 28.6%, cephalothin 18.2%, cloxacillin 8.3% and ciprofloxacin was active against 96.2% of isolates. The resistance pattern of proteus mirabilis were 55.6% to ampicillin, 47.1% to carbencillin, 29.4% to cephalothin, 14.3% to gentamicin and 4.8% to amikacin while 100% were sensitive to ciprofloxacin. We conclude that ciprofloxacin is the best drug of choice in treatment of CSOM caused by the common microorganisms.

Keywords: otitis media, chronic suppurative otitis media (CSOM), microorganism, drug sensitivity

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Authors: H. Kadi, A. Moussaoui, A. Medah, N. Benayahia, Nahal Bouderba

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For thousands of years, Punica granatum L. has been used in traditional medicine all over the world and predate the introduction of antibacterial drugs. The aim of the present study was to investigate the antibacterial activity of aqueous and ethanolic extracts of Punica granatum L. bark obtained by decoction and maceration. The different extracts of Punica granatum L. (Lythraceae) bark have been tested for antibacterial activity against Gram-positive bacteria (Staphylococcus aureus, Bacillus stearothermophilus) and Gram-negative bacteria (Escherichia coli, Pseudomonas aeruginosa) by disc diffusion method. The ethanolic macerate extract showed the strong in vitro antibacterial activity against Pseudomonas aeruginosa with zone inhibition of 24.4 mm. However, the results tests by disc diffusion method revealed the effectiveness of ethanolic decoctate against Gram-positive bacteria (Staphylococcus aureus and Bacillus stearothermophilus) with diameter zone of inhibition varying with 21.1mm and 23.75 mm respectively.

Keywords: Punica granatum L. bark, antibacterial activity, maceration, decoction

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Authors: András Fülöp, Lejla Heilmann, Zsolt Szabó, Ákos Koós

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Microbial fuel cells (MFCs) present a sustainable biotechnological solution to future energy demands. The aim of this study was to construct soil based, single cell, membrane-less MFC systems, operated without treatment to continuously power on-site monitoring and control systems during the soil bioremediation processes. Our Pseudomonas aeruginosa 541 isolate is an ideal choice for MFCs, because it is able to produce pyocyanin which behaves as electron-shuttle molecule, furthermore, it also has a significant antimicrobial effect. We tested several materials and structural configurations to obtain long term high power output. Comparing different configurations, a proton exchange membrane-less, 0.6 m long with 0.05 m diameter MFC tubes offered the best long-term performances. The long-term electricity production were tested from starch, yeast extract (YE), carboxymethyl cellulose (CMC) with humic acid (HA) as a mediator. In all cases, 3 kΩ external load have been used. The two best-operated systems were the Pseudomonas aeruginosa 541 containing MFCs with 1 % carboxymethyl cellulose and the MFCs with 1% yeast extract in the anode area and 35% hydrogel in the cathode chamber. The first had 3.3 ± 0.033 mW/m2 and the second had 4.1 ± 0.065 mW/m2 power density values. These systems have operated for 230 days without any treatment. The addition of 0.2 % HA and 1 % YE referred to the volume of the anode area resulted in 1.4 ± 0.035 mW/m2 power densities. The mixture of 1% starch with 0.2 % HA gave 1.82 ± 0.031 mW/m2. Using CMC as retard carbon source takes effect in the long-term bacterial survivor, thus enable the expression of the long term power output. The application of hydrogels in the cathode chamber significantly increased the performance of the MFC units due to their good water retention capacity.

Keywords: microbial fuel cell, bioremediation, Pseudomonas aeruginosa, biotechnological solution

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Authors: Nagat M. Saeed, Mabruka S. Elashheb, Fatma M. Ben Rabaha, Aisha M Edrah

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The aim of the study was to determine the type and pattern of antibiotic susceptibility of the pathogenic microorganisms causing chronic suppurative otitis media (CSOM), which could lead to better therapeutic decisions and consequently avoidance of appearance of resistance to specific antibiotics. Most frequently isolated agents were Pseudomonas aeruginosa 28.5%; followed by Staphylococcus aureus 18.2%; proteus mirabilis 13.9%; Providencia stuartti 6.7%; Bacteroides melaninogenicus, Aspergillus sp., candida sp., 4.2% each; and other microorganisms were represented in 3-0.2%. Drug sensitivities pattern of Pseudomonas aeruginosa showed that ciprofloxacin was active against the majority of isolates (93.9%) followed by ceftazidime 86.2%, amikacin 76.2% and gentamicin 40.8%. However, Staphylococcus aureus isolates were resistant to penicillin 72.7%, erythromycin 28.6%, cephalothin 18.2%, cloxacillin 8.3% and ciprofloxacin was active against 96.2% of isolates. The resistance pattern of proteus mirabilis was 55.6% to ampicillin, 47.1% to carbencillin, 29.4% to cephalothin, 14.3% to gentamicin and 4.8% to amikacin while 100% were sensitive to ciprofloxacin. We conclude that ciprofloxacin is the best drug of choice in the treatment of CSOM caused by the common microorganisms.

Keywords: otitis media, chronic suppurative otitis media (CSOM), microorganisms, drug sensitivity

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Authors: Memoona Ramzan, Bushra Tabassum, Anwar Khan, Muhammad Tariq, Mudassar Fareed Awan, Idrees Ahmad Nasir, Zahida Qamar, Naila Shahid, Tayyab Husnain

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Biological control is a novel approach being used in crop protection nowadays. Bacteria like Bacillus and Pseudomonas are reported for this purpose and few of their products are commercially available too. Rhizosphere and phyllosphere of healthy cotton plants were used as a source to isolate bacteria capable of exhibiting properties worthy for selection as biocontrol agent. For this purpose all isolated strains were screened for the activities like phosphate solubilization, Indole acetic acid (IAA) production and biocontrol against fungi. Two strains S1HL3 and S1HL4 showed phosphate solubilization and IAA production simultaneously while two other JS2HR4 and JS3HR2 were good inhibitors of fungal pathogens. Through biochemical and molecular characterization these bacteria were identified as P. aeruginosa, Burkholderia and Bacillus respectively. In green house trials of these isolates against Cotton leaf curl virus (CLCuV), seven treatments including individual bacterial isolate and consortia were included. Treated plants were healthy as compared to control plants in which upto 74% CLCuV symptomatic plants exist. Maximum inhibition of CLCuV was observed in T7 treated plants where viral load was only 0.4% as compared to control where viral load was upto 74%. This treatment consortium included Bacillus and Pseudomonas isolates; S1HL3, S1HL4, JS2HR4 and JS3HR2. Principal Component Biplot depicted highly significant correlation between percentage viral load and the disease incidence.

Keywords: cotton leaf curl virus, biological control, bacillus, pseudomonas

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Authors: M. L. Mangena, N. Mokoena, K. Rashamuse, M. G. Tlou

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Tertiary alcohol ester (TAE) hydrolases are a group of esterases (EC 3.1.1.-) that catalyze the kinetic resolution of TAEs and as a result, they are sought-after for the production of optically pure tertiary alcohols (TAs) which are useful as building blocks for number biologically active compounds. What sets these enzymes apart is, the presence of a GGG(A)X-motif in the active site which appears to be the main reason behind their activity towards the sterically demanding TAEs. The genome of Pseudomonas syringae pv. maculicola (Psm) comprises a multitude of genes that encode esterases. We therefore, hypothesize that some of these genes encode TAE hydrolases. In this study, Psm was screened for TAE hydrolase activity using the linalyl acetate (LA) plate assay and a positive reaction was observed. As a result, the genome of Psm was screened for esterases with a GGG(A)X-motif using the motif search tool and two potential TAE hydrolase genes (PsmEST1 and 2, 1100 and 1000bp, respectively) were identified, PsmEST1 was amplified by PCR and the gene sequenced for confirmation. Analysis of the sequence data with the SingnalP 4.1 server revealed that the protein comprises a signal peptide (22 amino acid residues) on the N-terminus. Primers specific for the gene encoding the mature protein (without the signal peptide) were designed such that they contain NdeI and XhoI restriction sites for directional cloning of the PCR products into pET28a. The gene was expressed in E. coli JM109 (DE3) and the clones screened for TAE hydrolase activity using the LA plate assay. A positive clone was selected, overexpressed and the protein purified using nickel affinity chromatography. The activity of the esterase towards LA was confirmed using thin layer chromatography.

Keywords: hydrolases, tertiary alcohol esters, tertiary alcohols, screening, Pseudomonas syringae pv., maculicola genome, esterase activity, linalyl acetate

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Authors: Shikha Masand, Sudesh Kumar Yadav

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Certain protein kinases have been shown to be crucial for plant cell signaling pathways associated with plant immune responses. Here we identified a horsegram [Macrotyloma uniflorum (Lam.) Verdc.] malectin-like leucine rich receptor-like protein kinase (RLK) gene MuRLK. The functional MuRLK protein preferentially binds to mannose and N-acetyl glucosamine residues. MuRLK exists in the cytoplasm and also localizes to the plasma membrane of plant cells via its N-terminus. Over-expression of MuRLK in Arabidopsis enhances the basal resistance to infection with Pseudomonas syringae pv. tomato, Alternaria brassicicola and Hyaloperonospora arabidopsidis, are associated with elevated ROS bursts, MAPK activation, thus ultimately leading to hypersensitive cell death. Moreover, salicylic acid-dependent and jasmonic acid-dependent defense responses are also enhanced in the MuRLK-overexpressed plants that lead to HR-induced cell death. Together, these results suggest that MuRLK plays a key role in the regulation of plant cell death, early and late defense responses after the recognition of microbial pathogens.

Keywords: horsegram, Pseudomonas syringae pv. tomato, MuRLK, ROS burst, cell death, plant defense

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Authors: Sujata Sharma, Avinash Singh, Lovely Gautam, Pradeep Sharma, Mau Sinha, Asha Bhushan, Punit Kaur, Tej P. Singh

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Peptidyl-tRNA hydrolase (Pth) Pth is an essential bacterial enzyme that catalyzes the release of free tRNA and peptide moeities from peptidyl tRNAs during stalling of protein synthesis. In order to design inhibitors of Pth from Pseudomonas aeruginosa (PaPth), we have determined the structures of PaPth in its native state and in the bound states with two compounds, amino acylate-tRNA analogue (AAtA) and 5-azacytidine (AZAC). The peptidyl-tRNA hydrolase gene from Pseudomonas aeruginosa was amplified by Phusion High-Fidelity DNA Polymerase using forward and reverse primers, respectively. The E. coliBL21 (λDE3) strain was used for expression of the recombinant peptidyl-tRNA hydrolase from Pseudomonas aeruginosa. The protein was purified using a Ni-NTA superflow column. The crystallization experiments were carried out using hanging drop vapour diffusion method. The crystals diffracted to 1.50 Å resolution. The data were processed using HKL-2000. The polypeptide chain of PaPth consists of 194 amino acid residues from Met1 to Ala194. The centrally located β-structure is surrounded by α-helices from all sides except the side that has entrance to the substrate binding site. The structures of the complexes of PaPth with AAtA and AZAC showed the ligands bound to PaPth in the substrate binding cleft and interacted with protein atoms extensively. The residues that formed intermolecular hydrogen bonds with the atoms of AAtA included Asn12, His22, Asn70, Gly113, Asn116, Ser148, and Glu161 of the symmetry related molecule. The amino acids that were involved in hydrogen bonded interactions in case of AZAC included, His22, Gly113, Asn116, and Ser148. As indicated by fittings of two ligands and the number of interactions made by them with protein atoms, AAtA appears to be a more compatible with the structure of the substrate binding cleft. However, there is a further scope to achieve a better stacking than that of O-tyrosyl moiety because it is not still ideally stacked. These observations about the interactions between the protein and ligands have provided the information about the mode of binding of ligands, nature and number of interactions. This information may be useful for the design of tight inhibitors of Pth enzymes.

Keywords: peptidyl tRNA hydrolase, Acinetobacter baumannii, Pth enzymes, O-tyrosyl

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Authors: Mohsina Akhter

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Cancer has become a wide spread disease around the globe and takes many lives every year. Different treatments are being practiced but all have potential side effects with somewhat less specificity towards target sites. Pseudomonas aeruginosa is known to secrete a protein azurin with special anti-cancer function. It has unique cell penetrating peptide comprising of 18 amino acids that have ability to enter cancer cells specifically. Reported function of Azurin is to stabilize p53 inside the tumor cells and induces apoptosis through Bax mediated cytochrome c release from mitochondria. At laboratory scale, we have made recombinant azurin through cloning rpTZ57R/T-azu vector into E.coli strain DH-5α and subcloning rpET28-azu vector into E.coli BL21-CodonPlus (DE3). High expression was ensured with IPTG induction at different concentrations then optimized high expression level at 1mM concentration of IPTG for 5 hours. Purification has been done by using Ni+2 affinity chromatography. We have concluded that azurin can be a remarkable improvement in cancer therapeutics if it produces on a large scale. Azurin does not enter into the normal cells so it will prove a safe and secure treatment for patients and prevent them from hazardous anomalies.

Keywords: azurin, pseudomonas aeruginosa, cancer, therapeutics

Procedia PDF Downloads 275

Authors: Felora Rafiei, Shahram Shoaei

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The aim of this studies was to evaluate effect of nitroxin, super nitro plus and biophosphorus on yield and yield components of maize (Zea mays) under different levels of chemical fertilizers in the condition of normal and difficiet irrigation. Experiment laid out as split plot factorial based on randomized complete block design with three replications. Main plots includes two irrigation treatments of 70 (I1), 120(I2) mm evaporation from class A pan. Sub plots were biological fertilizer and chemical fertilizer as factorial biological fertilizer consisting of nitroxin: Azospirillium lipoferum, Azospirillium brasilens, Azotobacter chroococcum Azotobacter agilis (108 CFU ml-1) (B1), super nitro plus (Azospirillium spp, + Pseudomonas fluorescence + Bacillus subtilis (108 CFU ml-1) + biological fungicide) (B2), biophosphorus (Pseudomonas spp + Bacillus spp (107 CFU ml-1) (B3), and chemical fertilizer consisting of NPK (C1), N5oP5oK5o (C2) and NoPoKo (C3).The results showed that usage of biological fertilizer have positive effects on chemical fertilizers use efficiency and tolerance to drought stress in maize. Also with use of biological fertilizer can decrease usage of chemical fertilizers.

Keywords: biological fertilizer, chemical fertilizer, yield component, yield, corn

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Authors: Ganeshkumar, Ashika, Valavan, Ramesh, Thangam, Chirayu

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MSCs are found in much higher concentration in the Wharton’s jelly compared to the umbilical cord blood, which is a rich source of hematopoietic stem cells. Umbilical cord tissue is collected at the time of birth; it is processed and stored in liquid nitrogen for future therapeutical purpose. The source of contamination might be either from vaginal tract of mother or from hospital environment or from personal handling during cord tissue sample collection. If the sample were contaminated, decontamination procedure will be done with 70% ethanol (1 minute) in order to avoid sample rejection. Ethanol is effective against a wide range of bacteria, protozoa and fungi and has low toxicity to humans. Among the 1954 samples taken for the study, 24 samples were found to be contaminated with microorganism. The organisms isolated from the positive samples were found to be E. coli, Stenotrophomonas maltophilia, Pseudomonas aueroginosa, Enterococcus fecalis, Acinetobacter bowmani, Staphylococcus epidermidis, Enterobacter cloacae, and Proteus mirabilis. Among these organisms 70% ethanol successfully eliminated E. coli, Enterococcus fecalis, Acinetobacter bowmani, Staphylococcus epidermidis, and Proteus mirabilis. 70% ethanol was unsuccessful in eliminating Stenotrophomonas maltophilia, Pseudomonas aueroginosa, and Enterobacter cloacae. Stenotrophomonas maltophilia and Pseudomonas aueroginosa have the ability to form biofilm that make them resistant to alcohol. Biofilm act as protective layer for bacteria and which protects them from host defense and antibiotic wash. Finally it was found 70% ethanol wash saved 58.3% cord tissue samples from rejection and it is ineffective against 41% of the samples. The contamination rate can be reduced by maintaining proper aseptic techniques during sample collection and processing.

Keywords: umblical cord tissue, decontamination, 70% ethanol effectiveness, contamination

Procedia PDF Downloads 317

Authors: Clara Franch de la Cal, Christopher J Morris, Michael McArthur

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Cystic fibrosis (CF) is an autosomal recessive genetic disorder characterized by abnormal transport of chloride and sodium across the lung epithelium, leading to thick and viscous secretions. Within which CF patients suffer from repeated bacterial pulmonary infections, with Pseudomonas aeru-ginosa (PA) eliciting the greatest inflammatory response, causing an irreversible loss of lung func-tion that determines morbidity and mortality. The cell wall of PA is a permeability barrier to many antibacterials and the rise of Mutli-Drug Resistant strains (MDR) is eroding the efficacy of the few remaining clinical options. In addition when PA infection becomes established it forms an antibi-otic-resistant biofilm, embedded in which are slow growing cells that are refractive to drug treat-ment. Making the development of new antibacterials a major challenge. This work describes the development of new type of nanoparticulate oligonucleotide antibacterial capable of tackling PA infections, including MDR strains. It is being developed to both block growth and prevent biofilm formation. These oligonucleotide therapeutics, Transcription Factor Decoys (TFD), act on novel genomic targets by capturing key regulatory proteins to block essential bacterial genes and defeat infection. They have been successfully transfected into a wide range of pathogenic bacteria, both in vitro and in vivo, using a proprietary delivery technology. The surfactant used self-assembles with TFD to form a nanoparticle stable in biological fluids, which protects the TFD from degradation and preferentially transfects prokaryotic membranes. Key challenges are to adapt the nanoparticle so it is active against PA in the context of biofilms and to formulate it for administration by inhalation. This would allow the drug to be delivered to the respiratory tract, thereby achieving drug concentrations sufficient to eradicate the pathogenic organisms at the site of infection.

Keywords: antibacterials, transcriptional factor decoys (TFDs), pseudomonas aeruginosa

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Authors: Daria Olkiewicz, Maciej Walczak

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In this paper, the biocidal effects of trans-cinnamaldehyde (a main component of cinnamon oil) and geraniol (a constituent of Pelargonium graveolens essential oil) are presented. The activity of the combination of trans-cinnamaldehyde and geraniol was tested against 3 bacterial strains: Staphylococcus aureus ATCC6538 (Gramm+), Escherichia coli ATCC8739 (Gramm-, Lac+) and Pseudomonas aeruginosa KKP 991(Gramm-, Lac-). The biocidal activity of trans-cinnamaldehyde-geraniol mixture against bacteria mentioned above was evaluated by disk-diffusion method. The model strains were exposed on 1, 2.5, 5 and 10 mg of trans-cinnamaldehyde-geraniol mixture per disk, and all strains were susceptible to this combination of plant compounds. For all microorganisms, also Minimal Inhibitory Concentration (MIC) and Minimal Bactericidal Concentration (MBC) were estimated. For Staphylococcus aureus MIC was 0.0625 mg/ml of the trans-cinnamaldehyde and geraniol mixture, and MBC was 1.25 mg/ml; For Escherichia coli MIC=0.5 mg/ml, MBC=1 mg/ml, and finally Pseudomonas aeruginosa was inhibited in 0.5 mg/ml, and minimal biocidal concentration of tested mixture for it was 1.25 mg/ml. There are also reports about the synergistic working of trans-cinnamaldehyde and geraniol against microorganisms and the antimicrobial activity of polymers enriched with trans-cinnamaldehyde or geraniol, therefore the successful development and introduction to the today life of biocidal polymer enriched with trans-cinnamaldehyde and geraniol are possible.

Keywords: antibacterial activity, biocidal polymers, geraniol, trans-cinnamaldehyde

Procedia PDF Downloads 142

Authors: M. Mohammadi

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In order to evaluate the effect of phosphate and Zn bio-fertilizers on the yield, phytic acid (PA), Zn concentration and PA/Zn molar ratio in bean, a field experiment was carried out for two years. The treatments included two cultivars of bean (Talash and Sadri), four levels of P (P₀, P₁: 100 kg ha^-1 triple super phosphate (TSP), P2: 50 kg ha^-1 TSP + phosphate bio-fertilizer, P₃: phosphate bio-fertilizer), three levels of Zn (Zn₀, Zn₁: 50 kg ha^-1 ZnSO4, Zn₂: Zn bio-fertilizer). Phosphate bio-fertilizer consisted of inoculum of mycorrhizal fungus and Azotobacter and Zn bio-fertilizer consisted of Pseudomonas bacteria. The results revealed that there was significant difference between yield and Zn concentration between years. The effect of cultivar was significant on studied parameters. The lowest content of PA and PA/Zn were obtained from Talash. P treatment caused to significant difference on parameters in which P₂ caused to increase yield, P and Zn concentration, and decrease PA and PA/Zn by 21.8%, 38.2%, 33.4%, 17.4% and 38.6% respectively. Zn treatment caused to significant difference on studied parameters. The maximum number of parameters were obtained from Zn₁ and Zn₂. The higher Zn concentration led to lower content of PA and PA/Zn. Using of P and Zn bio–fertilizers were caused to increasing nutrient uptake, improving growth condition and reducing PA and PA/Zn molar ratio.

Keywords: mycorrhizae, phosphorus, pseudomonas, zinc

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Authors: Muhammad Naeem Chaudhry, Fahim Nawaz, Rana Nauman Shabbir

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New strategies aimed at increasing the resilience of crop plants to the negative effects of climate change represent important research priorities of plant scientists. The use of soil microorganisms to alleviate abiotic stresses like drought has gained particular importance in recent past. A field experiment was planned to investigate the effect of phosphorous solubilizing bacteria on yield and seed quality of Camelina (Camelina sativa L.) under water deficit conditions. The study was conducted at Agronomic Research Farm, University College of Agriculture and Environmental Sciences, The Islamia University Bahawalpur, during 4th week of November, 2013. The available seeds of Camelina sativa were inoculated with two bacterial strains (pseudomonas and Bacillus spp.) and grown under various water stress levels i.e. D0, (four irrigations), D3 (three irrigation), D2 (two irrigations), and D1 (one irrigation). The results revealed that drought stress significantly reduced the plant growth and yield, consequently reducing protein contents and oil concentration in camelina. The exposure to drought stress decreased plant height (16%), plant population (27%), number of fertile branches (41-59%), number of pods per plant (35%) and seed per pod (33%). Drought stress also exerted a negative impact on yield characteristics by reducing the 1000-seed weight (65%), final seed yield (52%), biological yield (22%) and harvest index (39%) of camelina. However, the inoculation of seeds with Pseudomonas and Bacillus spp. promoted the plant growth characterized by increased plant height and enhanced plant population. It was noted that inoculation of seeds with Pseudomonas resulted in the maximum plant population (113.4 cm), primary branches (19 plant-1), and number of pods (664 plant-1), whereas Bacillus inoculation resulted in maximum plant height (113.4 cm), seeds per pod (15.9), 1000-seed weight (1.85 g), and seed yield (3378.8 kg ha-1). Moreover, the inoculation with Bacillus also significantly improved the quality attributes of camelina and gave 3.5% and 2.1% higher oil contents than Pseudomonas and control (no-inoculation), respectively. Similarly, the same strain also resulted in maximum protein contents (33.3%). Our results confirmed the hypothesis that inoculation of seeds with phosphorous solubilizing bacterial strains is an effective, viable and environment-friendly approach to improve yield and quality of camelina under water deficit conditions. However, further studies are suggested to investigate the physiological and molecular processes, stimulated by bacterial strains, for increasing drought tolerance in food crops.

Keywords: Camelina, drought stress, phosphate solubilizing bacteria, seed quality

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Authors: N. Meziou-Chebouti, A. Merabet, Y. Chebouti N. Behidj

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This work is a contribution to the knowledge of physicochemical characteristics of mature carob followed by evaluation of the activity, antimicrobial phenolics leaves and green pods of Ceratonia siliqua L. physicochemical study shows that mature carob it has a considerable content of sugar (50.90%), but poor in proteins (7%), fat (8%) and also has a high mineral content. The results obtained from phenolic extracts of leaves and green pods of Ceratonia siliqua L. show a wealth leaf phenolic extract especially flavonoids (0,545 mg EqQ/g) relative to the extract of green pods (0,226 mgEqQ/g). Polyphenols leaves have a slightly inhibitory effect on the growth of strains: Staphylococcus aureus, Escherichia coli, Klebsiella pneumoiae, Streptococcus sp and Sanmonella enteritidis, a strong inhibitory effect on the growth of Pseudomonas strain aerogenosa. Moreover, polyphenols pod have a slightly inhibitory effect on the growth of Streptococcus sp strains, Pseudomonas and aerogenosa Sanmonella enteritidis, a slightly inhibitory effect on the growth of Klebsiella pneumoniae strains, E. coli and Staphylococcus aureus.

Keywords: antimicrobial activity, bacteria, clove, Ceratonia siliqua, polyphenols

Procedia PDF Downloads 321