Search results for: Fusarium strains

Authors: Lali Kutateladze, Tamar Urushadze, Tamar Dudauri, Besarion Metreveli, Nino Zakariashvili, Izolda Khokhashvili, Maya Jobava

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Lignocellulosic waste streams from agriculture, paper and wood industry are renewable, plentiful and low-cost raw materials that can be used for large-scale production of liquid and gaseous biofuels. As opposed to prevailing multi-stage biotechnological processes developed for bioconversion of cellulosic substrates to ethanol where high-cost cellulase preparations are used, Consolidated Bioprocessing (CBP) offers to accomplish cellulose and xylan hydrolysis followed by fermentation of both C6 and C5 sugars to ethanol in a single-stage process. Syntrophic microbial consortium comprising of anaerobic, thermophilic, cellulolytic, and saccharolytic bacteria in the genus Clostridia with improved ethanol productivity and high tolerance to fermentation end-products had been proposed for achieving CBP. 65 new strains of anaerobic thermophilic cellulolytic and saccharolytic Clostridia were isolated from different wetlands and hot springs in Georgia. Using new isolates, fermentation of mechanically pretreated wheat straw and corn stalks was done under oxygen-free nitrogen environment in thermophilic conditions (T=550C) and pH 7.1. Process duration was 120 hours. Liquid and gaseous products of fermentation were analyzed on a daily basis using Perkin-Elmer gas chromatographs with flame ionization and thermal detectors. Residual cellulose, xylan, xylose, and glucose were determined using standard methods. Cellulolytic and saccharolytic bacteria strains degraded mechanically pretreated herbaceous cellulosic wastes and fermented glucose and xylose to ethanol, acetic acid and gaseous products like hydrogen and CO2. Specifically, maximum yield of ethanol was reached at 96 h of fermentation and varied between 2.9 – 3.2 g/ 10 g of substrate. The content of acetic acid didn’t exceed 0.35 g/l. Other volatile fatty acids were detected in trace quantities.

Keywords: anaerobic bacteria, cellulosic wastes, Clostridia sp, ethanol

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Authors: Fatma Koksal Cakirlar, Murat Gunaydin, Nevri̇ye Gonullu, Nuri Kiraz

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During the last few decades, the increasing prevalence of methicillin resistant-CoNS isolates has become a common problem worldwide. Macrolide-lincosamide-streptogramin B (MLSB) antibiotics are effectively used for the treatment of CoNS infections. However, resistance to MLSB antibiotics is prevalent among staphylococci. The aim of this study is to determine species distribution and the incidence of inducible clindamycin resistance in CoNS isolates caused nosocomial bacteremia in our hospital. Between January 2014 and October 2015, a total of 484 coagulase-negative CoNS isolates were isolated from blood samples of patients with true bacteremia who were hospitalized in intensive care units and in other departments of Istanbul University Cerrahpasa Medical Hospital. Blood cultures were analyzed with the BACTEC 9120 system (Becton Dickinson, USA). The identification and antimicrobial resistance of isolates were determined by Phoenix automated system (BD Diagnostic Systems, Sparks, MD). Inducible clindamycin resistance was detected using D-test. The species distribution was as follows: Staphylococcus epidermidis 211 (43%), S. hominis 154 (32%), S. haemolyticus 69 (14%), S. capitis 28 (6%), S. saprophyticus 11 (2%), S. warnerii 7 (1%), S. schleiferi 5 (1%) and S. lugdunensis 1 (0.2%). Resistance to methicillin was detected in 74.6% of CoNS isolates. Methicillin resistance was highest in S.hemoliticus isolates (89%). Resistance rates of CoNS strains to the antibacterial agents, respectively, were as follows: ampicillin 77%, gentamicin 20%, erythromycin 71%, clindamycin 22%, trimethoprim-sulfamethoxazole 45%, ciprofloxacin 52%, tetracycline 34%, rifampicin 20%, daptomycin 0.2% and linezolid 0.2%. None of the strains were resistant to vancomycin and teicoplanin. Fifteen (3%) CoNS isolates were D-test positive, inducible MLSB resistance type (iMLSB-phenotype), 94 (19%) were constitutively resistant (cMLSB -phenotype), and 237 (46,76%) isolates were found D-test negative, indicating truly clindamycin-susceptible MS phenotype (M-phenotype resistance). The incidence of iMLSB-phenotypes was higher in S. epidermidis isolates (4,7%) compared to other CoNS isolates.

Keywords: bacteremia, inducible MLSB resistance phenotype, methicillin-resistant, staphylococci

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Authors: Andrea Parisi, Samantha Vilkins, Luis Furuya-Kanamori, John A. Crump, Benjamin P. Howden, Darren Gray, Kathryn Glass, Martyn Kirk

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Objectives: Salmonella is a leading cause of foodborne enterocolitis worldwide. Nontyphoidal Salmonella (NTS) infections that are Multi-Drug Resistant (MDR) (non-susceptible to ≥1 agent in ≥3 antimicrobial categories) may result in more severe outcomes, although these effects have not been systematically examined. We conducted a systematic review and meta-analysis to examine impacts of MDR NTS on health in high-income settings. Methods: We systematically reviewed the literature from scientific databases, including PubMed, Scopus and grey literature sources, using PRISMA guidelines. We searched for data from case-control studies, cohorts, outbreaks, reports and theses, imposing no language restriction. We included only publications from January 1990 to September 2016 from high income countries as classified by World Bank. We extracted data from papers on duration of illness, hospitalisation rates, morbidity and mortality for MDR and non-MDR NTS strains. Results: After removing duplicates, the initial search revealed 4258 articles. After further screening, we identified 16 eligible studies for the systematic review, and 9 of these were included in meta-analysis. NTS serotypes differed among the reported studies but serotype Typhimurium, Enteritidis, Newport and Heidelberg were among the most often reported as MDR pathogens. Salmonella infections that were MDR were associated with excess bloodstream infections (OR 1.63; 95%CI 1.18-2.26), excess hospitalisations (OR 2.77; 95%CI 1.47-5.21) and higher mortality (OR 3.54; 95%CI 1.10-11.40). Conclusions: MDR NTS infections are a serious public health concern. With the emergence of MDR Salmonella strains in the high-income countries, it is crucial to restrict the use of antimicrobials both in animals and humans, and intervene to prevent foodborne infections.

Keywords: Antimicrobial Resistance, Bloodstream Infection, Health Outcomes, Hospitalisation, Invasive Disease, Multi-Drug Resistance (MDR), Mortality, Nontyphoidal Salmonella

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Authors: P. M. G. Pathiraja, P. Egodawatta, A. Goonetilleke, V. S. J. Te'o

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The production and use of Polychlorinated biphenyls (PCBs), a group of synthetic halogenated hydrocarbons have been restricted worldwide due to its toxicity and categorized as one of the twelve priority persistent organic pollutants (POP) by the Stockholm Convention. Low reactivity and high chemical stability of PCBs have made them highly persistent in the environment and bio-concentration and bio-magnification along the food chain contribute to multiple health impacts in humans and animals. Remediating environments contaminated with PCBs is a challenging task for decades. Use of microorganisms for remediation of PCB contaminated soils and sediments have been widely investigated due to the potential of breakdown these complex contaminants with minimum environmental impacts. To achieve an effective bioremediation of polychlorinated biphenyls (PCBs) contaminated environments, microbes were sourced from environmental samples and tested for their ability to hydrolyze PCBs under different conditions. Comparison of PCB degradation efficiencies of four naturally occurring facultative bacterial cultures isolated through selective enrichment under aerobic and anaerobic conditions were simultaneously investigated in minimal salt medium using 50 mg/L Aroclor 1260, a commonly used commercial PCB mixture as the sole source of carbon. The results of a six-week study demonstrated that all the tested facultative Achromobacter, Ochrobactrum, Lysinibacillus and Pseudomonas strains are capable of degrading PCBs under both anaerobic and aerobic conditions while assisting hydrophobic PCBs to make solubilize in the aqueous minimal medium. Overall, the results suggest that some facultative bacteria are capable of effective in degrading PCBs under anaerobic conditions through reductive dechlorination and under aerobic conditions through oxidation. Therefore, use of suitable facultative microorganisms under combined anaerobic-aerobic conditions and combination of such strains capable of solubilization and breakdown of PCBs has high potential in achieving higher PCB removal rates.

Keywords: bioremediation, combined anaerobic-aerobic degradation, facultative microorganisms, polychlorinated biphenyls

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Authors: Mahmoudi Noureddine

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In this study the use of two variable refined plate theories of laminated composite plates to static response of laminated plates. The plate theory accounts for parabolic distribution of the transverse shear strains, and satisfies the zero traction boundary conditions on the surfaces of the plate without using shear correction factor. The validity of the present theory is demonstrated by comparison with solutions available in the literature and finite element method. The result is presented for the static response of simply supported rectangular plates under uniform sinusoidal mechanical loadings.

Keywords: bending, composite, laminate, plates, fem

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Authors: H. Alijani, M. Jabari, S. Matroodi, H. Zolqarnein, A. Sharafi, I. Zamani

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Actinomycetes, Gram-positive bacteria, are interesting as a main producer of secondary metabolites and are important industrially and pharmaceutically. The marine environment is a potential source for new actinomycetes, which can provide novel bioactive compounds and industrially important enzymes. The aims of this study were to isolate and identify novel actinomycetes from Persian Gulf sediments and screen these isolates for the production of secondary metabolites, especially antibiotics, Using phylogenetic (16S rRNA gene sequence), morphological and biochemical analyses. 15 different actinomycete strains from Persian Gulf sediments at a depth of 5-10 m were identified. DNA extraction was done using Cinnapure DNA Kit. PCR amplification of 16S rDNA gene was performed using F27 and R1492 primers. Phylogenetic tree analysis was performed using the MEGA 6 software. Most of the isolated strains belong to the genus namely Streptomyces (14), followed by Nocardiopsis (1). Antibacterial assay of the isolates supernatant was performed using a standard disc diffusion assay with replication (n=3). The results of disk diffusion assay showed that most active strain against Proteus volgaris and Bacillus cereus was AMJ1 (16.46±0.2mm and 13.78±0.2mm, respectively), against Salmonella sp. AMJ7 was the most effective strain (10.13±0.2mm), and AMJ1 and AHA5 showed more inhibitory activity against Escherichia coli (8.04±0.02 mm and 8.2±0.03 ). The AMJ6 strain showed best antibacterial activity against Klebsiella sp. (8.03±0.02mm). Antifungal activity of AMJ2 showed that it was most active strain against complex (16.05±0.02mm) and against Aspergillus flavus strain AMJ1 was most active strain (16.4±0.2mm) and highest antifungal activity against Trichophyton mentagrophytes, Microsporum gyp serum and Candida albicans, were shown by AHA1 (21.03±0.02mm), AHA3 and AHA7 (18±0.03mm), AMJ6 (21.03±0.2mm) respectively. Our results revealed that the marine actinomycetes of Persian Gulf sediments were potent source of novel antibiotics and bioactive compounds and indicated that the antimicrobial metabolites were extracellular. Most of the secondary metabolites and antibiotics are extracellular in nature and extracellular products of actinomycetes show potent antimicrobial activities.

Keywords: antibacterial activity, antifungal activity, marine actinomycetes, Persian Gulf

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Authors: Sohan Sengupta, Arnab Pramanik, Abhrajyoti Ghosh, Maitree Bhattacharyya

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Newly emerged phyto-pathogens and multi drug resistance have been threating the world for last few decades. Actinomycetes, the most endowed group of microorganisms isolated from unexplored regions of the world may be the ultimate solution to these problems. Thus the aim of this study was to isolate several bioactive actinomycetes strains capable of producing antimicrobial secondary metabolite from Sundarbans, the only mangrove tiger land of the world. Fifty four actinomycetes were isolated and analyzed for antimicrobial activity against fifteen test organisms including three phytopathogens. Nine morphologically distinct and biologically active isolates were subjected to polyphasic identification study. 16s rDNA sequencing indicated eight isolates to reveal maximum similarity to the genus streptomyces, whereas one isolate presented only 93.57% similarity with Streptomyces albogriseolus NRRL B-1305T. Seventy-one carbon sources and twenty-three chemical sources utilization assay revealed their metabolic relatedness. Among these nine isolates three specific strains were found to have notably higher degree of antimicrobial potential effective in a broader range including phyto-pathogenic fungus. PCR base whole genome screen for PKS and NRPS genes, confirmed the occurrence of bio-synthetic gene cluster in some of the isolates for novel antibiotic production. Finally the strain SMS_SU21, which showed antimicrobial activity with MIC value of 0.05 mg ml-1and antioxidant activity with IC50 value of 0.242±0.33 mg ml-1 was detected to be the most potential one. True prospective of this strain was evaluated utilizing GC-MS and the bioactive compound responsible for antimicrobial activity was purified and characterized. Rare bioactive actinomycetes were isolated from unexplored heritage site. Diversity of the biosynthetic gene cluster for antimicrobial compound production has also been evaluated. Antimicrobial compound SU21-C has been identified and purified which is active against a broad range of pathogens.

Keywords: actinomycetes, sundarbans, antimicrobial, pks nrps, phyto-pathogens, GC-MS

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Authors: Marina Arino Martin, John McEvoy, Eakalak Khan

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Endoxifen is an active metabolite responsible for the effectiveness of tamoxifen, a chemotherapeutic drug widely used for endocrine responsive breast cancer and chemo-preventive long-term treatment. Tamoxifen and endoxifen are not completely metabolized in human body and are actively excreted. As a result, they are released to the water environment via wastewater treatment plants (WWTPs). The presence of tamoxifen in the environment produces negative effects on aquatic lives due to its antiestrogenic activity. Because endoxifen is 30-100 times more potent than tamoxifen itself and also presents antiestrogenic activity, its presence in the water environment could result in even more toxic effects on aquatic lives compared to tamoxifen. Data on actual concentrations of endoxifen in the environment is limited due to recent discovery of endoxifen pharmaceutical activity. However, endoxifen has been detected in hospital and municipal wastewater effluents. The detection of endoxifen in wastewater effluents questions the treatment efficiency of WWTPs. Studies reporting information about endoxifen removal in WWTPs are also scarce. There was a study that used chlorination to eliminate endoxifen in wastewater. However, an inefficient degradation of endoxifen by chlorination and the production of hazardous disinfection by-products were observed. Therefore, there is a need to remove endoxifen from wastewater prior to chlorination in order to reduce the potential release of endoxifen into the environment and its possible effects. The aim of this research is to isolate and identify bacteria strain(s) capable of degrading endoxifen into less hazardous compound(s). For this purpose, bacteria strains from WWTPs were exposed to endoxifen as a sole carbon and nitrogen source for 40 days. Bacteria presenting positive growth were isolated and tested for endoxifen biodegradation. Endoxifen concentration and by-product formation were monitored. The Monod kinetic model was used to determine endoxifen biodegradation rate. Preliminary results of the study suggest that isolated bacteria from WWTPs are able to growth in presence of endoxifen as a sole carbon and nitrogen source. Ongoing work includes identification of these bacteria strains and by-product(s) of endoxifen biodegradation.

Keywords: biodegradation, bacterial degraders, endoxifen, wastewater

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Authors: Ayman K. El Essawy, Amal M. Hosny, Hala M. Abu Shady

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The rapid detection of TB and drug resistance, both optimizes treatment and improves outcomes. In the current study, respiratory specimens were collected from 155 patients. Conventional susceptibility testing and MIC determination were performed for rifampicin (RIF) and isoniazid (INH). Genotype MTBDRplus assay, which is a molecular genetic assay based on the DNA-STRIP technology and specific gene sequencing with primers for rpoB, KatG, and mab-inhA genes were used to detect mutations associated with resistance to rifampicin and isoniazid. In comparison to other categories, most of rifampicin resistant (61.5%) and isoniazid resistant isolates (47.1%) were from patients relapsed in treatment. The genotypic profile (using Genotype MTBDRplus assay) of multi-drug resistant (MDR) isolates showed missing of katG wild type 1 (WT1) band and appearance of mutation band katG MUT2. For isoniazid mono-resistant isolates, 80% showed katG MUT1, 20% showed katG MUT1, and inhA MUT1, 20% showed only inhA MUT1. Accordingly, 100% of isoniazid resistant strains were detected by this assay. Out of 17 resistant strains, 16 had mutation bands for katG distinguished high resistance to isoniazid. The assay could clearly detect rifampicin resistance among 66.7% of MDR isolates that showed mutation band rpoB MUT3 while 33.3% of them were considered as unknown. One mono-resistant rifampicin isolate did not show rifampicin mutation bands by Genotype MTBDRplus assay, but it showed an unexpected mutation in Codon 531 of rpoB by DNA sequence analysis. Rifampicin resistance in this strain could be associated with a mutation in codon 531 of rpoB (based on molecular sequencing), and Genotype MTBDRplus assay could not detect the associated mutation. If the results of Genotype MTBDRplus assay and sequencing were combined, this strain shows hetero-resistance pattern. Gene sequencing of eight selected isolates, previously tested by Genotype MTBDRplus assay, could detect resistance mutations mainly in codon 315 (katG gene), position -15 in inhA promotes gene for isoniazid resistance and codon 531 (rpoB gene) for rifampicin resistance. Genotyping techniques allow distinguishing between recurrent cases of reinfection or reactivation and supports epidemiological studies.

Keywords: M. tuberculosis, rpoB, KatG, inhA, genotype MTBDRplus

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Authors: A. Berraf-Tebbal, Z. Bouznad, , A.J.L. Phillips

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Phaeomoniella is a fungus genus in the mitosporic ascomycota which includes Phaeomoniella chlamydospora specie associated with two declining diseases on grapevine (Vitis vinifera) namely Petri disease and esca. Recent studies have shown that several Phaeomoniella species also cause disease on many other woody crops, such as forest trees and woody ornamentals. Two new species, Phaeomoniella zymoides and Phaeomoniella pinifoliorum H.B. Lee, J.Y. Park, R.C. Summerbell et H.S. Jung, were isolated from the needle surface of Pinus densiflora Sieb. et Zucc. in Korea. The identification of species in Phaeomoniella genus can be a difficult task if based solely on morphological and cultural characters. In this respect, the application of molecular methods, particularly PCR-based techniques, may provide an important contribution. MSP-PCR (microsatellite primed-PCR) fingerprinting has proven useful in the molecular typing of fungal strains. The high discriminatory potential of this method is particularly useful when dealing with closely related or cryptic species. In the present study, the application of PCR fingerprinting was performed using the micro satellite primer M13 for the purpose of species identification and strain typing of 84 Phaeomoniella -like isolates collected from grapevines with typical symptoms of dieback. The bands produced by MSP-PCR profiles divided the strains into 3 clusters and 5 singletons with a reproducibility level of 80%. Representative isolates from each group and, when possible, isolates from Eutypa dieback and esca symptoms were selected for sequencing of the ITS region. The ITS sequences for the 16 isolates selected from the MSP-PCR profiles were combined and aligned with sequences of 18 isolates retrieved from GenBank, representing a selection of all known Phaeomoniella species. DNA sequences were compared with those available in GenBank using Neighbor-joining (NJ) and Maximum-parsimony (MP) analyses. The phylogenetic trees of the ITS region revealed that the Phaeomoniella isolates clustered with Phaeomoniella chlamydospora reference sequences with a bootstrap support of 100 %. The complexity of the pathosystems vine-trunk diseases shows clearly the need to identify unambiguously the fungal component in order to allow a better understanding of the etiology of these diseases and justify the establishment of control strategies against these fungal agents.

Keywords: Genotyping, MSP-PCR, ITS, phylogeny, trunk diseases

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Authors: Ahilya Singh, Brad Carte, Ramesh Subramani, William Aalbersberg

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A total of 37 actinomycete strains were purified from 25 Solomon Islands marine sediments using four different types of isolation media. Among them, 54% of the strains had obligate requirement of seawater for growth. The ethyl acetate extract of 100 ml fermentation product of each strain was screened for antimicrobial activity against multidrug resistant human pathogens and cytotoxic activity against brine shrimps. A total of 67% of the ethyl acetate extracts showed antimicrobial and/or cytotoxic activities. A strain F-1915 was selected for isolation and evaluation of bioactive compound(s) based on its bioactive properties and chemical profile analysis using the LC-MS. The strain F-1915 was identified to have 96% sequence similarity to Streptomyces violaceusniger on the basis of 16S rDNA sequences using BLAST analysis. The 16S rDNA revealed that the strain F-1915 is a new member of MAR4 clade of actinomycetes. The MAR4 clade is an interesting clade of actinomycetes known for the production of pharmaceutically important hybrid isoprenoid compounds. The ethyl acetate extract of the fermentation product of this strain was purified by silica gel column chromatography and afforded the isolation of one bioactive pure compound. Based on the 1D and 2D NMR spectral data of compound 1 it was identified as a new mono-brominated phenazinone, Marinophenazimycin A, a structure which has already been studied by external collaborators at Scripps Institution of Oceanography but is yet to be published. Compound 1 displayed significant antimicrobial activity against drug resistant human pathogens. The minimum inhibitory concentration (MIC) of compound 1 was against Methicillin Resistant Staphylococcus aureus (MRSA) was about 1.9 μg/ml and MIC recorded against Amphotericin Resistant Candida albicans (ARCA) was about 0.24 μg/ml. The bioactivity of compound 1 against ARCA was found to be better than the standard antifungal agent amphotericin B. Compound 1 however did not show any cytotoxic activity against brine shrimps.

Keywords: actinomycetes, antimicrobial activity, brominated phenazine, MAR4 clade, marine natural products, multidrug resistent, 1D and 2D NMR

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Authors: Yassir Adam Shuaib, Stefan Niemann, Eltahir Awad Khalil, Ulrich Schaible, Lothar Heinz Wieler, Mohammed Ahmed Bakhiet, Abbashar Osman Mohammed, Mohamed Abdelsalam Abdalla, Elvira Richter

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Tuberculosis (TB) is a chronic bacterial disease of humans and animals and it is characterized by the progressive development of specific granulomatous tubercle lesions in affected tissues. In a six-month study, from June to November 2014, a total of 2,304 carcasses of cattle, camel, sheep, and goats slaughtered at East and West Gaash slaughterhouses, Kassala, were investigated during postmortem, in parallel, 101 sputum samples from TB suspected patients at Kassala and El-Gadarif Teaching Hospitals were collected in order to investigate tuberculosis in animals and humans. Only 0.1% carcasses were found with suspected TB lesions in the liver and lung and peritoneal cavity of two sheep and no tuberculous lesions were found in the carcasses of cattle, goats or camels. All samples, tissue lesions and sputum, were decontaminated by the NALC-NaOH method and cultured for mycobacterial growth at the NRZ for Mycobacteria, Research Center Borstel, Germany. Genotyping and molecular characterization of the grown strains were done by line probe assay (GenoType CM and MTBC) and 16S rDNA, rpoB gene, and ITS sequencing, spoligotyping, MIRU-VNTR typing and next generation sequencing (NGS). Culture of the specimens revealed growth of organisms from 81.6% of all samples. Mycobacterium tuberculosis (76.2%), M. intracellulare (14.2%), mixed infection with M. tuberculosis and M. intracellulare (6.0%) and mixed infection with M. tuberculosis and M. fortuitum and with M. intracellulare and unknown species (1.2%) were detected in the sputum samples and unknown species (1.2%) were detected in the samples of one of the animals tissues. From the 69 M. tuberculosis strains, 25 (36.2%) were showing either mono-drug-resistant or multi-drug-resistant or poly-drug-resistant but none was extensively drug-resistant. In conclusion, the prevalence of TB in animals was very low while in humans M. tuberculosis-Delhi/CAS lineage was responsible for most cases and there was an evidence of MDR transmission and acquisition.

Keywords: animal, human, slaughterhouse, Sudan, tuberculosis

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Authors: Lachin Mikjtarnejad, Mohsen Farzaneh

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Various microbes, such as fungi and bacteria species, are naturally found in the fruit microbiota, and some of them act as a pathogen and result in fruit rot. Among non-pathogenic microbes, yeasts (single-celled microorganisms belonging to the fungi kingdom) can colonize fruit tissues and interact with them without causing any damage to them. Although yeasts are part of the plant microbiota, there is little information about their interactions with plants in comparison with bacteria and filamentous fungi. According to several existing studies, some yeasts can colonize different plant species and have the biological control ability to suppress some of the plant pathogens. It means those specific yeast-colonized plants are more resistant to some plant pathogens. The major objective of the present investigation is to isolate yeast strains from apple fruit and screen their ability to control Penicillium expansum, the causal agent of blue mold of fruits. In the present study, psychrotrophic and epiphytic yeasts were isolated from apple fruits that were stored at low temperatures (0–1°C). Totally, 42 yeast isolates were obtained and identified by molecular analysis based on genomic sequences of the D1/D2 and ITS1/ITS4 regions of their rDNA. All isolated yeasts were primarily screened by' in vitro dual culture assay against P. expansum by measuring the fungus' relative growth inhibition after 10 days of incubation. The results showed that the mycelial growth of P. expansum was reduced between 41–53% when challenged by promising yeast strains. The isolates with the strongest antagonistic activity belonged to Metschnikowia pulcherrima A13, Rhodotorula mucilaginosa A41, Leucosporidium Scottii A26, Aureobasidium pullulans A19, Pichia guilliermondii A32, Cryptococcus flavescents A25, and Pichia kluyveri A40. The results of seven superior isolates to inhibit blue mold decay on fruit showed that isolates A. pullulans A19, L. scottii A26, and Pi. guilliermondii A32 could significantly reduce the fruit rot and decay with 26 mm, 22 mm and 20 mm zone diameter, respectively, compared to the control sample with 43 mm. Our results show Pi. guilliermondii strain A13 was the most effective yeast isolates in inhibiting P. expansum on apple fruits. In addition, various biological control mechanisms of promising biological isolates against blue mold have been evaluated to date, including competition for nutrients and space, production of volatile metabolites, reduction of spore germination, production of siderophores and production of extracellular lytic enzymes such as chitinase and β-1,3-glucanase. However, the competition for nutrients and the ability to inhibit P. expansum spore growth have been introduced as the prevailing mechanisms among them. Accordingly, in our study, isolates A13, A41, A40, A25, A32, A19 and A26 inhibited the germination of P. expansum, whereas isolates A13 and A19 were the strongest inhibitors of P. expansum mycelia growth, causing 89.13% and 81.75 % reduction in the mycelial surface, respectively. All the promising isolates produced chitinase and β-1,3-glucanase after 3, 5 and 7 days of cultivation. Finally, based on our findings, we are proposing that, Pi. guilliermondiias as an effective biocontrol agent and alternative to chemical fungicides to control the blue mold of apple fruit.

Keywords: yeast, yeast enzymes, biocontrol, post harvest diseases

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Authors: Evy Latifah, Eli Korlina, Hanik Anggraeni, Kuntoro Boga, Joko Mariyono

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During the rainy season, the tomato plants are vulnerable to various diseases. A disease that attacks the leaves of tomato plants (foliar diseases) such as late blight (Phytophtora infestans) and spotting bacteria (bacterial spot / Xanthomonas sp.) In addition, there is a disease that attacks the roots such as fusarium and bacterial wilt. If not immediately anticipated, it will decrease the quality and quantity of crop yields. In fact, it can lead to crop failure. The aim of this research is to know the production of tomato grafting by using Timoty and CLN 3024 tomatoes at rain shelter during rainy season in lowland. Data were analyzed using analysis of variance and tested further by Least Significant Difference (LSD) level of 5 %. The parameters measured were plant height (cm), stem diameter (cm), number of fruit space, canopy extended, number of branches, number of productive branches, and the number of stem segments. The results show at the beginning of growth until the end of the treatment without grafting with relative rain shelter displays the highest plant height. This was followed by extensive crop canopy. For tomato grafting and non-grafting using rain shelter able to produce the number of branches and number of productive branches at most. While at the end of the growth in the number of productive branches generated as much. Highest production of tomatoes produced by tomato dig rafting to use the shelter.

Keywords: field trail, wet and dry season, production, diseases, rain shelter

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Authors: Afrah E. Mohammed, Mudawi M. Nour

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Environmentally friendly green synthesis of nanomaterial has a very significant part in nanotechnology. In the present research, the synthesis of silver nanoparticles (AgNPs) was established by treating silver ions with the aqueous extract of Calligonum comosum green shoots at room temperature. AgNPs formation was firstly detected by the colour change of mixed extract (plant extract and AgNO3). Further characterization was done by ultraviolet (UV)-Vis spectrophotometer, transmission electron microscopy (TEM), scanning electron microscopy (SEM), zeta potential and fourier transform infrared spectroscopy (FTIR). The peak values for UV-VIS- spectroscopy were in the range of 440 nm, TEM micrograph showed a spherical shape for the particles and zeta potential showed the formation of negative charged nanoparticles with an average size of about 105.8 nm. 1635.41 and 3249.83 cm−1 are the peaks detected from the FTIR analysis. In this study, biosynthesized silver nanoparticles mediated by C. comosum were tested for their antimycotic activity using a well diffusion method against fungal species; Aspergillus flavus, Penicillium sp, Fusarium oxysporum. Our findings indicated that biosynthesized AgNPs showed an efficient antimycotic activity against tested species. The antimycotic action of AgNPs varied according to different fungal species. Results confirmed the ability of C. comosum green shoot extract to act as an reducing and stabilizing agent during the synthesis of AgNPs.

Keywords: AGNPS, zeta potential, TEM, SEM

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Authors: Marie Bajnarová

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The core part of the research project is represented by taking a perspective on the role of an educator in Juvenile Institutional Centres. In accordance with the research questions, the research explores impact of the environment, situations, practices, attitudes, values and also experience of the respondents. Art activities minimize risky behaviours and contribute to a healthy lifestyle. They also help children and adolescents with conduct disorders develop positive social behaviour, psychosocial skills and cope with difficult life situations.

Keywords: Juvenile Detention Centres, drawing, conduct disorders, art therapy

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Authors: Evy Latifah, Eko Widaryanto, M. Dawam Maghfoer, Arifin

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Tomato (Lycopersicon esculentum Mill.) is potential vegetables to develop, because it has high economic value and has the potential to be exported. There is a decrease in tomato productivity due to unfavorable growth conditions such as bacterial wilt, fusarium wilt, high humidity, high temperature and inappropriate production technology. Grafting technology is one alternative technology. In addition to being able to control the disease in the soil, grafting is also able to increase the growth and yield of production. Besides, it is also necessary to know the economic benefits if using grafting technology. A promising eggplant rootstock for tomato grafting is Solanum torvum. S. torvum is selected as a rootstock with high compatibility. The purpose of this research is to know the effect of grafting several varieties of tomatoes with Solanum torvum as a rootstock. The experiment was conducted in Agricultural Extension Center Pare. Experimental Garden of Pare Kediri sub-district from July to early December 2016. The materials used were tomato Cervo varieties, Karina, Timoty, and Solanum torvum. Economic analysis, growth, and yield including plant height, number of leaves, percentage of disease and tomato production were used as performance measures. The study showed that grafting tomato Timoty scion with Solanum torvum as rootstock had higher production. Financially, grafting tomato Timoty and Cervo scion had higher profit about. 28,6% and 16,3% compared to Timoty and Cervo variety treatment without grafting.

Keywords: grafting technology, economic analysis, growth, yield of tomato, Solanum torvum

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Authors: Messaouda Belaid, Salima Kebbouche-Gana, Djamila Benaziza

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Our study focuses on determining the antibacterial activity of some honeys from northern Algeria. To test this activity, the agar well diffusion methods was employed. The bacterial strains tested were Staphylococcus aureus, Bacillus subtilis, Streptococcus faecalis, Klebsiella pneumoniae, Escherichia coli, and Pseudomonas aeroginosae. The results showed that all the microbes tested were inhibited by all honey used in this study but Those bacteria that appear to be more sensitive to all honey tested are Staphylococcus aureus and Pseudomonas aeroginosae.

Keywords: honey, antibacterial activity, Northern Algeria, Staphylococcus aureus

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Authors: Abu S. Mustafa, Mohammad W. Khan, Faraz Shaheed Khan, Nazima Habibi

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Brucellosis is a zoonotic disease of worldwide prevalence. There are at least four species and several strains of Brucella that cause human disease. Brucella genomes have very limited variation across strains, which hinder strain identification using classical molecular techniques, including PCR and 16 S rDNA sequencing. The aim of this study was to perform whole genome sequencing of clinical isolates of Brucella and perform bioinformatics and comparative genomics analyses to determine the existence of genetic differences across the isolates of a single Brucella species and strain. The draft sequence data were generated from 15 clinical isolates of Brucella melitensis (biovar 2 strain 63/9) using MiSeq next generation sequencing platform. The generated reads were used for further assembly and analysis. All the analysis was performed using Bioinformatics work station (8 core i7 processor, 8GB RAM with Bio-Linux operating system). FastQC was used to determine the quality of reads and low quality reads were trimmed or eliminated using Fastx_trimmer. Assembly was done by using Velvet and ABySS softwares. The ordering of assembled contigs was performed by Mauve. An online server RAST was employed to annotate the contigs assembly. Annotated genomes were compared using Mauve and ACT tools. The QC score for DNA sequence data, generated by MiSeq, was higher than 30 for 80% of reads with more than 100x coverage, which suggested that data could be utilized for further analysis. However when analyzed by FastQC, quality of four reads was not good enough for creating a complete genome draft so remaining 11 samples were used for further analysis. The comparative genome analyses showed that despite sharing same gene sets, single nucleotide polymorphisms and insertions/deletions existed across different genomes, which provided a variable extent of diversity to these bacteria. In conclusion, the next generation sequencing, bioinformatics, and comparative genome analysis can be utilized to find variations (point mutations, insertions and deletions) across different genomes of Brucella within a single strain. This information could be useful in surveillance and epidemiological studies supported by Kuwait University Research Sector grants MI04/15 and SRUL02/13.

Keywords: brucella, bioinformatics, comparative genomics, whole genome sequencing

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Authors: O. P. Yadav

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Ag-N co-doped ZnS and ZnS/Fe2O3 composite nanoparticles have been synthesized by chemical and sol-gel methods. As-synthesized nanomaterial have been characterized by XRD and TEM techniques and their antimicrobial effects were studied using paper disc diffusion technique against gram positive (Staphylococcus aureus) and gram negative (Escherichia coli) bacteria. As-synthesized nanomaterial showed potent antimicrobial activity against studied bacterial strains. Antimicrobial activity of synthesized nanomaterial has also been compared with some commonly used antibiotics.

Keywords: antibiotic, Escherichia coli, nanomaterial, TEM, Staphylococcus aureus

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Authors: Sze Wing Ho, Nagendra P. Shah

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Lactic acid bacteria (LAB) have shown the beneficial effects on human gastrointestinal tract, such as protects diarrhea induced by lactose intolerance or enteric pathogens. Citrulline is a non-protein amino acid and also the precursors of arginine and nitric oxide, it has shown to enhance intestinal barrier function. Citrulline has shown to improve the growth of some strains of LAB, it is important for LAB to have a sufficient cell concentration to contribute the effects. Therefore, the aims of this study were to investigate the effect of combining citrulline with LAB on the anti-adhesion effect against pathogens and the effect on the cell integrity. The effect of citrulline on selected LAB was determined by incubating in 0%, 0.1% or 0.2% citrulline enriched MRS broth for 18 h. The adhesion ability of LAB and the anti-adhesion effect of LAB and citrulline against pathogens were performed on IPEC-J2 cell line. Transepithelial electrical resistance (TEER) assay was used to measure the tight junction (TJ) integrity. TJ proteins (claudin-1, occludin and zonula occluden-1 (ZO-1)) were determined by western blot analysis. It found that the growth of Lactobacillus helveticus ASCC 511 was significantly stimulated by 0.2% citrulline compared with control during 18 h fermentation. The adhesion of L. helveticus ASCC 511 and Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) ASCC 756 was increased when supplemented with citrulline. Citrulline has shown significant inhibitory effect on the adhesion of Escherichia coli PELI0480 (O157:H7), Shigella sonnei ATCC 25931, Staphyloccocus aureus CMCC26003 and Cronobacter sakazakii ATCC 29544. The anti-adhesion effect of L. helveticus ASCC 511, L. bulgaricus ASCC 756 and Lactobacillus paracasei ASCC 276 against Cronobacter sakazakii ATCC 29544 was significantly enhanced with citrulline supplementation. Treatments with citrulline and LAB were able to maintain the TEER of IPEC-J2 cell and shown the positive effect on the TJ proteins. In conclusion, citrulline had stimulating effect on some strains of LAB and determined to improve the adhesion of LAB on intestinal epithelial cell, to enhance the inhibitory effect on enteric pathogens adhesion as well as had beneficial effects on maintaining cell integrity. It implied LAB supplemented with citrulline might have advantageous effects on gastrointestinal tracts.

Keywords: citrulline, lactic acid bacteria, amino acid, anti-adhesion effect, cell integrity

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Authors: Elizabeth Starzycka-Korbas, Michal Starzycki, Wojciech Rybinski, Mirosława Dabert

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For a few years, the populations of pathogenic fungi occurring in winter oilseed rape in Malyszyn were analyzed. Brassica napus L. in Poland and in the world is a source of energy for both the men (oil), and animals, as post-extraction middling, as well as a motor fuel (oil, biofuel) therefore studies of this type are very important. The species composition of pathogenic fungi can be an indicator of seed yield. The occurrence of oilseed rape pathogens during several years were analyzed using the sequencing method DNA ITS. The results were compared in the gene bank using the program NCBI / BLAST. In field conditions before harvest of oilseed rape presence of pathogens infesting B. napus has been assessed. For example, in 2015, 150 samples have been isolated and applied to PDA medium for the identification of belonging species. From all population has been selected mycelium of 83 isolates which were sequenced. Others (67 isolates) were pathogenic fungi of the genus Alternaria which are easily to recognize. The population of pathogenic species on oilseed rape have been identified after analyzing the DNA ITS and include: Leptosphaeria sp. 38 (L. maculans 25, L. biglobosa 13), Alternaria sp. 29, Fusarium sp. 3, Sclerotinia sclerotiorum 7, heterogeneous 6, total of 83 isolates. The genus Alternaria sp. fungi wear the largest share of B. napus pathogens in particular years. Another dangerous species for oilseed rape was Leptosphaeria sp. Populations of pathogens in each year were different. The number of pathogens occurring in the field and their composition is very important for breeders and farmers because of the possible selection of the most resistant genotypes for sowing in the next growing season.

Keywords: B. napus, DNA ITS Sequencing, pathogenic fungi, population

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Authors: Ellana Boada, Eric Santos-Clotas, Alba Cabrera-Codony, Maria Martin, Lluis Baneras, Frederic Gich

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Volatile methylsiloxanes (VMS) are a group of manmade silicone compounds widely used in household and industrial applications that end up on the biogas produced through the anaerobic digestion of organic matter in landfills and wastewater treatment plants. The presence of VMS during the biogas energy conversion can cause damage on the engines, reducing the efficiency of this renewable energy source. Non regenerative adsorption onto activated carbon is the most widely used technology to remove siloxanes from biogas, while new trends point out that biotechnology offers a low-cost and environmentally friendly alternative to conventional technologies. The first objective of this research was to enrich, isolate and identify bacterial species able to grow using siloxane molecules as a sole carbon source: anoxic wastewater sludge was used as initial inoculum in liquid anoxic enrichments, adding D4 (as representative siloxane compound) previously adsorbed on activated carbon. After several months of acclimatization, liquid enrichments were plated onto solid media containing D4 and thirty-four bacterial isolates were obtained. 16S rRNA gene sequencing allowed the identification of strains belonging to the following species: Ciceribacter lividus, Alicycliphilus denitrificans, Pseudomonas aeruginosa and Pseudomonas citronellolis which are described to be capable to degrade toxic volatile organic compounds. Kinetic assays with 8 representative strains revealed higher cell growth in the presence of D4 compared to the control. Our second objective was to characterize the community composition and diversity of the microbial community present in the enrichments and to elucidate whether the isolated strains were representative members of the community or not. DNA samples were extracted, the 16S rRNA gene was amplified (515F & 806R primer pair), and the microbiome analyzed from sequences obtained with a MiSeq PE250 platform. Results showed that the retrieved isolates only represented a minor fraction of the microorganisms present in the enrichment samples, which were represented by Alpha, Beta, and Gamma proteobacteria as dominant groups in the category class thus suggesting that other microbial species and/or consortia may be important for D4 biodegradation. These results highlight the need of additional protocols for the isolation of relevant D4 degraders. Currently, we are developing molecular tools targeting key genes involved in siloxane biodegradation to identify and quantify the capacity of the isolates to metabolize D4 in batch cultures supplied with a synthetic gas stream of air containing 60 mg m⁻³ of D4 together with other volatile organic compounds found in the biogas mixture (i.e. toluene, hexane and limonene). The isolates were used as inoculum in a biotrickling filter containing lava rocks and activated carbon to assess their capacity for siloxane removal. Preliminary results of biotrickling filter performance showed 35% of siloxane biodegradation in a contact time of 14 minutes, denoting that biological siloxane removal is a promising technology for biogas upgrading.

Keywords: bacterial cultivation, biogas upgrading, microbiome, siloxanes

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Authors: Zafar Iqbal, Sana Irshad Khan

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Organic extract from newly isolated plant growth promoting fungus (PGPF) Penicillium sp EU0013 was subjected to bioassays including anti fungal (disc diffusion) cytotoxicity (brine shrimp lethality), herbicidal (Lemna minor) and nematicidal activities. Metabolite profile of the extract was also assessed using HPLC analysis with the aim to identify bioactive natural products in the extract as new drug candidate(s). The extract showed anti fungal potential against tested fungal pathogens. Growth of the Wilt pathogen Fusarium oxyosproum was inhibited up to 63% when compared to negative reference. Activity against brine shrimps was weak and mortality up to 10% was observed at concentration of 200 µg. mL-1. The extract exhibited no toxicity against Lemna minor frond at 200 µg. mL-1. Nematicidal activity was observed very potent against root knot nematode and LC50 value was calculated as 52.5 ug. mL-1 using probit analysis. Methodically assessment of metabolites profile by HPLC showed the presence of kojic acid (Rt 1.4 min) and aflatoxin B1 (Rt 5.9 min) in the mycellial extract as compared with standards. The major unidentified metabolite was eluted at Rt 8.6 along with other minor peaks. The observed high toxicity against root knot nematode was attributed to the unidentified compounds that make fungal extract worthy of further exploration for isolation and structural characterization studies for development of future commercial nematicidal compound(s).

Keywords: penicillium, nematicidal activity, metabolites, HPLC

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Authors: Waheed Ahmad Safi, Shunichi Nakamura

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Concrete filled steel I-girder (CFIG) bridge was proposed and the bending and shear strength was confirmed by experiments. The area surrounded by the upper and lower flanges and the web is filled with concrete in CFIG, which is used to the intermediate support of a continuous girder. Three-dimensional finite element models were established to simulate the bending and shear behaviors of CFIG and to clarify the load transfer mechanism. Steel plates and filled concrete were modeled as a three-dimensional 8-node solid element and steel reinforcement bars as a three-dimensional 2-node truss element. The elements were mostly divided into the 50 x 50 mm mesh size. The non-linear stress-strain relation is assumed for concrete in compression including the softening effect after the peak, and the stress increases linearly for concrete in tension until concrete cracking but then decreases due to tension stiffening effect. The stress-strain relation for steel plates was tri-linear and that for reinforcements was bi-linear. The concrete and the steel plates were rigidly connected. The developed FEM model was applied to simulate and analysis the bending behaviors of the CFIG specimens. The vertical displacements and the strains of steel plates and the filled concrete obtained by FEM agreed very well with the test results until the yield load. The specimens collapsed when the upper flange buckled or the concrete spalled off. These phenomena cannot be properly analyzed by FEM, which produces a small discrepancy at the ultimate states. The FEM model was also applied to simulate and analysis the shear tests of the CFIG specimens. The vertical displacements and strains of steel and concrete calculated by FEM model agreed well with the test results. A truss action was confirmed by the FEM and the experiment, clarifying that shear forces were mainly resisted by the tension strut of the steel plate and the compression strut of the filled concrete acting in the diagonal direction. A trail design with the CFIG was carried out for a four-span continuous highway bridge and the design method was established. Construction cost was estimated about 12% lower than that of a conventional steel I-section girder.

Keywords: concrete filled steel I-girder, bending strength, FEM, limit states design, steel I-girder, shear strength

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Authors: Omaima A. Sharaf, Tarek A. Moussa, Said M. Badr El-Din, H. Moawad

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Background: Polycyclic aromatic hydrocarbons (PAHs) pose great environmental and human health concerns for their widespread occurrence, persistence, and carcinogenic properties. PAHs releases due to anthropogenic activities to the wider environment have led to higher concentrations of these contaminants than would be expected from natural processes alone. This may result in a wide range of environmental problems that can accumulate in agricultural ecosystems, which threatened to become a negative impact on sustainable agricultural development. Thus, this study aimed to evaluate the physico-chemical, and microbial properties of the compost leachate (CL) to assess its role as nutrient and microbial source (biostimulation/bioaugmentation) for developing a cost-effective bioremediation technology for PAHs contaminated sites. Material and Methods: PAHs-degrading bacteria were isolated from CL that was collected from a composting site located in central Scotland, UK. Isolation was carried out by enrichment using phenanthrene (PHR), pyrene (PYR) and benzo(a)pyrene (BaP) as the sole source of carbon and energy. The isolates were characterized using a variety of phenotypic and molecular properties. Six different isolates were identified based on the difference in morphological and biochemical tests. The efficiency of these isolates in PAHs utilization was assessed. Further analysis was performed to define taxonomical status and phylogenic relation between the most potent PAHs-utilizing bacterial strains and other standard strains, using molecular approach by partial 16S rDNA gene sequence analysis. Results indicated that the 16S rDNA sequence analysis confirmed the results of biochemical identification, as both of biochemical and molecular identification of the isolates assigned them to Bacillus licheniformis, Pseudomonas aeruginosa, Alcaligenes faecalis, Serratia marcescens, Enterobacter cloacae and Providenicia which were identified as the prominent PAHs-utilizers isolated from CL. Conclusion: This study indicates that the CL samples contain a diverse population of PAHs-degrading bacteria and the use of CL may have a potential for bioremediation of PAHs contaminated sites.

Keywords: polycyclic aromatic hydrocarbons, physico-chemical analyses, compost leachate, microbial and biochemical analyses, phylogenic relations, 16S rDNA sequence analysis

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Authors: Victor Iorungwa Gwa, Ebenezer Jonathan Ekefan

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Potential of Trichoderma harzianum for biological control of postharvest fungal rot of white yam (Dioscorea rotundata Poir) tubers in storage was studied. Pathogenicity test revealed the susceptibility of healthy looking yam tubers to Aspergillus niger, Botryodiplodia theobromae, and Fusarium oxysporum f. sp. melonganae after fourteen days of inoculation. Treatments comprising A. niger, B. theobromae, and F. oxysporum each paired with T. harzianum and were arranged in completely randomized design and stored for five months. Experiments were conducted between December 2015 and April 2016 and December 2016 and April 2017. Results showed that tubers treated with the pathogenic fungi alone caused mean percentage rot of between 6.67 % (F. oxysporum) and 22.22 % (A. niger) while the paired treatments produced only between 2.22 % (T. harzianum by F. oxysporum) and 6.67 % (T. harzianum by A. niger). In the second year of storage, mean percentage rot was found to be between 13.33 % (F. oxysporum) and 28.89 % (A. niger) while in the paired treatment rot was only between 6.67 % (F. oxysporum) and 8.89% (A. niger). Tubers treated with antagonist alone produced 0.00 % and 2.22 % in the first and second year, respectively. Result revealed that there was a significant difference (P ≤ 0.05) in mean percentage rot between the first year and the second year except where B. theobromae was inoculated alone, A. niger and T. harzianum paired and B. theobromae and T. harzianum paired. The most antagonised fungus in paired treatment for both years was F. oxysporum f. sp. melonganae, while the least antagonised, was A. niger and B. theobromae. It is, therefore, concluded that T. harzianum has potentials to control rot causing pathogens of yam tubers in storage. This can compliment or provide better alternative ways of reducing rot in yam tubers than by the use of chemical fungicides which are not environmentally friendly.

Keywords: biological control, fungal rot, postharvest, Trichoderma harzianum, white yam

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Authors: Omaima A. Sharaf, Wafaa M. Abd El-Rahim, Hassan Moawad, Michael J. Sadowsky

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Textile industry is one of the important industries in the worldwide. It is known that the eco-friendly industrial and agricultural activities are significant for socio-economic stability of all countries. The absence of appropriate industrial waste water treatments is essential barrier for sustainable development in food and agricultural sectors especially in developing country like Egypt. Thus, the development of enzymatic bioremediation technology for textile dye removal will enhance the collaboration between scientists who develop the technology and industry where this technology will be implemented towards the safe disposal of the textile dye wastes. Highly efficient microorganisms are of most importance in developing and using highly effective biological treatment processes. Bacterial degradation of azo dyes is generally initiated by an enzymatic step that involves cleavage of azo linkages, usually with the aid of an azoreductase as electron donor. Thus, expanding the spectrum of microorganisms with high enzymatic activities as azoreductases and discovering novel azo-dye degrading enzymes, with enhanced stability and superior catalytic properties, are necessary for many environmental and industrial applications. Consequently, the use of molecular tools has become increasingly integrated into the understanding of enzyme properties and characterization. Researchers have utilized a gene cloning and expression methods as a tool to produce recombinant protein for decolorizing dyes more efficiently. Thus, presumptive evidence for the presence of genes encoding azoreductases in the genomes of selected local, and most potent azo-degrading strains were obtained by using specific oligonucleotides primers. These potent strains have been isolated from textile industrial wastewater in Egypt and identified using 16S rRNA sequence analysis as 'Lysinibacillus macrolidesB8, Brevibacillus parabrevisB11, Bacillus coagulansB7, and B. cereusB5'. PCR products of two full length genes designated as (AZO1;621bp and AZO2;534bp) were detected. BLASTx results indicated that AZO1 gene was corresponding to predicted azoreductase from of Bacillus sp. ABP14, complete genome, multispecies azoreductase [Bacillus], It was submitted to the gene bank by an accession no., BankIt2085371 AZO1 MG923210 (621bp; 207 amino acids). AZO1 was generated from the DNA of our identified strains Lysinibacillus macrolidesB8. On the other hand, AZO2 gene was corresponding to a predicted azoreductase from Bacillus cereus strain S2-8. Gene bank accession no. was BankIt2085839 AZO2 MG932081 (534bp;178 amino acids) and it was amplified from our Bacillus coagulansB7. Both genes were successfully cloned into pCR2.1TOPO (Invitrogen) and in pET28b+ vectors, then they transformed into E. coli DH5α and BL21(DE3) cells for heterologous expression studies. Our recombinant azoreductases (AZO1&AZO2) exhibited potential enzyme activity and efficiently decolorized an azo dye (Direct violet). They exhibited pH stability between 6 and 8 with optimum temperature up to 60°C and 37 °C after induction by 1mM and 1.5mM IPTG, for both AZO1 &AZO2, respectively. These results suggested that further optimization and purification of these recombinant proteins by using different heterologous expression systems will give great potential for the sustainable utilization of these recombinant enzymes in several industrial applications especially in wastewater treatments.

Keywords: azoreductases, decolorization, enzyme activity, gene cloning and expression

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Authors: Fouzia Qamar, Shahida Hasnain

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The present study is primarily concerned with the alteration in haemocyte profile of adult male Periplaneta americana with emphasis on the effect of bacterial inoculations on the haemogram i.e., total haemocyte count (THC) and differential haemocyte count (DHC) of different haemocyte types of the target insect. Haemolymph cellular profile showed considerable alterations under the effect of nine strains of Agrobacterium tumefaciens after 8, 16 and 24 hrs of treatment thereby signifying the potential role of Agrobacterium tumefaciens as a possible biocontrol agent against the house hold pests.

Keywords: Agrobacterium tumefaciens, Periplaneta americana, Haemolymph, cellular parametes

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Authors: Satya Eswari Jujjavarapu, Swast Dhagat

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Contagious diseases enact severe public health problems and have upsetting consequences. The cyclic lipopeptides explained by bacteria Bacillus, Paenibacillus, Pseudomonas, Streptomyces, Serratia, Propionibacterium and fungus Fusarium are very critical in confining the pathogens. As the degree of drug resistance upsurges in unparalleled manner, the perseverance of searching novel cyclic lipopeptides is being professed. The intense study has shown the implication of these bioactive compounds extending beyond antibacterial and antifungal. Lipopeptides, composed of single units of peptide and fatty acyl moiety, show broad spectrum antimicrobial effects. Among the surplus of cyclic lipopeptides, only few have materialized as strong antibiotics. For their functional vigor, polymyxin, daptomycin, surfactin, iturin and bacillomycin have been integrated in mainstream healthcare. In our work daptomycin has been a major part of antimicrobial resource since the past decade. Daptomycin, a cyclic lipopeptide consists of 13-member amino acid with a decanoyl side-chain. This structure of daptomycin confers it the mechanism of action through which it forms pore in the bacterial cell membrane resulting in the death of cell. Daptomycin is produced by Streptococccus roseoporus and acts against Streptococcus pneumonia (PSRP), methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE). The PDB structure and ligands of daptomycin are available online. The molecular docking studies of these ligands with the lipopeptides were performed and their docking score and glide energy were recorded.

Keywords: daptomycin, molecular docking, structure-based drug design, lipopeptide

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