فصلنامه زیست شناسی میکروارگانیسم ها
پیاپی 7 (پاییز 1392)

Halophiles، especially haloarchaea are one of the most important groups of extremophiles. Halophilic hydrolases have been studied worldwide and have been considered for biotechnology and industrial technologies. This study is the first report in amylopullulanase production in halophilic microorganisms. Materials and method s: A halophilic archaeon، Halorubrum sp. strain Ha25، produced extracellular halophilic organic solvent-tolerant amylopullulanase. The enzyme was purified using ethanol precipitation and anion exchange chromatography method. Molecular mass of purified enzyme was determined by SDS–PAGE method. After purification، the enzyme was characterized. To study the effects of organic solvents in the stability of the enzyme، the enzyme solution was incubated in the presence of various organic compounds and then، residual enzyme activity was measured. Mode of action of the enzyme was determined by thin-layer chromatography. Molecular weight of the purified enzyme was estimated to be 140 kDa by SDS–PAGE method. Optimum temperature for amylolitic and pullulytic activities was 50 °C. Optimum pH for amylolitic activity was 7. 0 and for pullulytic activity was 7. 5. This enzyme was active over a wide range of concentrations (0-4. 5 M) of NaCl. The effect of organic solvents on the amylolitic and pullulytic activities showed that this enzyme was more stable in the presence of non-polar organic solvents than polar solvents. The enzyme solely hydrolyzed pullulan and soluble starch to glucose. Discussion and Halorubrum sp. strain Ha25 produces thermophilic and extremely halophilic amylopullulanase. The catalytic function under multi extreme condition of high temperature، high salinity، and low water activity might possess biotechnological and commercial values such as treatment waste solutions with starch residues، high salt content and solvents.

Due to the vast growth of world population، consumption of a lot of energy، limited energy supply and rising prices of fuel oil in the future، other alternative energy source is essential. Ethanol is renewable and a safe fuel and it is mainly based on microbial fermentation. The purpose of this study was i solation of high ethanol producing bacteria from the fermentation process of alcohol industries and optimization of growth conditions to be introduced to the industries. The samples that were collected from fermentation tanks of alcohol industries were enriched in ZSM medium. To isolate the ethanol producing bacteria، the enriched culture was transferred on RMA agar. Bacterial growth conditions and their effects on ethanol production were optimized based on pH، growth temperature، agitation، fermentation time، initial substrate concentration and carbon and nitrogen sources. In addition، the morphological، physiological and molecular characterizations were investigated for identification of the isolates. Three bacterial isolates ZYM7، ZYM8 and ZYM9 were isolated from fermentation tank. All isolates were able to produce ethanol 5. 00، 7. 60 and 4. 00 gL-1 after 48 hours، respectively. The results demonstrated that all isolates were able to consume most sugars sources specially pentose carbon xylose. The isolate ZYM7 produced 13. 00 gL-1 ethanol by consumption of xylose. The results of morphological and physiological characteristics showed that ZYM7 belonged to Lactobacillus sp. and ZYM8 and ZYM9 belonged to Acetobacter sp. Moreover، 16S rRNA sequencing and phylogenetic analyses exhibited that ZYM7 was similar to Lactobacillus rhamnosus with 99% homology and ZYM8 and ZYM9 were similar to Acetobacter pasteurianus with 99 and 98% homology، respectively. Discussion and The results showed that that the isolated bacteria were suitable candidates to produce ethanol from raw material enriched with xylose. These bacteria could be introduced to fermentation industries.

Zinc oxide nanoparticles have quite a few applications in the fields of biology، optics، mechanics، magnetism، energy، hygiene and medicine. Due to serious problems associated with physiochemical synthesis of ZnO nanoparticles، including environmental pollution، complicated and costly processes، there is a growing need to develop a simple biological procedure for synthesis of nanoparticles to achieve the monodisperse-sized particles with a higher purity، low energy consumption and a cleaner environment. We conducted this investigation to screen and isolate native fungi strains capable of high zinc metal tolerance ability and a potential for extracellular synthesis of ZnO nanoparticles using fungal secretions as biological catalysts. 15 different strains of fungi were isolated from soil samples collected from lead and zinc mines of Angoran-Zanjan using conventional enrichment process and characterized initially based on macroscopic and microscopic characteristics and colony morphology. The intrinsic tolerance of the isolated strains to zinc toxic metal was measured in the synthetic and complex media using the agar dilution method. The supernatants of isolated fungi were incubated with zinc acetate solution in a shaker incubator for 72h; then، the strain that was able to synthesis ZnO nanoparticle was identified. The ZnO nanoparticles formation was investigated by using spectroscopic techniques and microscopic observations. Among the 15 isolated strains، the strain ZRS14 had highest zinc metal tolerance ability and was selected and identified as Aspergillus niger strain ZRS14 (GenBank accession number KF414527) based on morphological and molecular phylogenetic analysis. For synthesis of ZnO nanoparticles by isolated A. niger ZRS14، fungal cell-free filtrate of the strain was collected and incubated in the presence of zinc acetate solution at a final concentration of 250 mg/l zinc metal ion at 28º C for 72 h on a rotary shaker (150 rpm) under dark conditions. Results obtained by visual observations، spectral data achieved from UV–vis، XRD spectrum and SEM micrographs revealed the extracellular formation of ZnO nanoparticles with narrow size distribution and average particle size of 32 nm with the collected cell-free filtrate of A. niger isolate ZRS14. Discussion and Owing to the extracellular synthesis of ZnO nanoparticles، the obtained results in the current study suggest that the A. niger strain ZRS14 has a considerable potentiality that can be efficiently used as an eco-friendly biocatalyst for the preparative synthesis of ZnO nanoparticles. The present investigation is the first report on the biological synthesis of ZnO nanoparticles using newly isolated strain of Aspergillus niger ZRS14.

Evolution and dissemination of multidrug-resistant Salmonella enterica isolates have been reported. Resistance to antibiotic within Salmonella serotypes is a serious global concern. Integrons are genetic units that participate in capturing and dissemination of mobile gene cassettes among Gram-negative microorganisms. Integron mobility is facilitated by transposons and plasmids. Antibiotic resistance phenotype of 32 Salmonella enterica isolates of animal origin which were collected during 2011–2012 were investigated. All samples were assessed by culture method and standard biochemical tests for identification of Salmonella strains. After DNA extraction، the presence of class I integron was examined by PCR. The most common resistant phenotypes were to cefalothin (100%)، chloramphenicol (68/7%)، ampicillin (62/5%)، tetracycline (56/2%)، amoxicillin/ clavulanate (50%)، sulfamethoxazole (43/7%). Class 1 integrons were found in (55/5%) and (64. 2 %) of Salmonella isolates from livestock and poultry respectively. Discussion and Integron positive isolates had higher resistance to tetracycline، chloramphenicol، ampicillin sulfamethoxazole and amoxicillin / clavulanate compared with integron negative isolates. The ability of integrons to integrate resistance gene to antimicrobial agents makes them the main sources in the diffusion of antibiotic resistance.

Biodegradation is one of the main degrading factors of leather artifacts. Determination of fungi and other microorganisms has vital importance for conservation of valuable objects made from organic materials. Usual classic techniques for identification of pathogens and fungi have some difficulties، but FTIR spectroscopy has been introduced as useful technique for characterization and determination of microorganisms. In this study، historic leathers were related to the Seljuk period. The leathers were sampled in laboratory and fungi were cultivated in SDA medium. Genuses of fungi were identified according to macroscopic، microscopic and morphologic characteristics. Microorganisms were analyzed by ATR-FTIR spectrometer and differences of spectral properties were assessed. Spectral properties were assessed according to form and absorption region of peaks. Cluster analysis was used for evaluation of different regions and areas on the measured spectra. Results showed the presence of Penicillium sp. (9 samples، 33. 3%)، Aspergillus sp. (5 samples، 18. 5%)، Cladosporium sp. (4 samples، 14. 8%)، Rhizoctonia sp. (2 samples، 7. 4%)، Trichophyton sp. (1 sample، 3. 7%) and also one sample of yeast (3. 7%). Two species of fungi remained unidentified (7. 4%) and there was not any growth in 3 samples (11. 1%). Spectral properties indicated to structural differences between microorganisms (fungi، yeast، bacteria). It showed characteristics of fungi genus and species in some cases. Discussion and Active fungi induce decomposition، hydrolysis and decay of leather and its vegetable tannins. The decay leads to aesthetic change and health problems. Therefore، it is necessary to remove of fungi and appropriate treatment of leather artifacts. Application of ATR-FTIR spectroscopy had good results for identification of fungi، yeast and bacteria. Assessments showed the potential of ATR-FTIR spectroscopy for easy and rapid discrimination between microorganisms and specially fungi genus and species in some cases. This study indicated the possibility of developing FTIR–ATR spectroscopy as a reliable method for rapid identification of fungal pathogens. Advantages of the method showed that it was an appropriate technique for fungi evaluation in large scale. IR spectroscopy could help in identification and classification of fungi. Combining the advantages of FTIR spectroscopy with other used routine techniques، might improve the efficiency of fungal classification and identification. Chemical composition of the samples can be simultaneously analyzed. Chemical analysis by FTIR spectroscopy results to better understanding of fungi growth and complex chemical processes during fungal development and substrate degradation.

Polyphosphates، also called volutin granules، are linear polymers from orthophosphates linked by energy-rich phosphoanhydride bands that have been seen in bacteria، yeasts، fungi، plants and animals. These polymers are completely safe and nontoxic، and have numerous applications in food and drug industries. Due to the great importance and wide range of the utilization of these polymers in various industries، several factors such as various carbon sources، carbon source concentration and phosphorus concentration were studied and optimized. In order to increase polyphosphate production in Bacillus megaterium strain G11. The optimization process was carried out with determination of the amount of polyphosphate accumulated in cell and phosphorus removed from the medium. One-way ANOVA and Tukey tests were used in order to determine whether there was a significant difference between data obtained in this research. Growth of B. megaterium in the presence of sucrose (OD=3. 026) was better than glucose (OD=2. 616) whereas polyphosphate production and phosphorus removal from medium were higher in the presence of glucose (0. 033 g g-1 dry cell weight and 1. 61 g l-1، respectively). On the other hand، polyphosphate production and phosphorus removal from medium coordinately were decreased with increasing glucose concentration. Furthermore، in studying the effects of phosphorus، we faced two phases of rising and falling. Actually، the increase of phosphorus concentration (0. 25-1 g l-1) in medium caused an increase in polyphosphate production and phosphorus removal from medium whereas both of them were decreased with a more increase in amount of phosphorus (1-4 g l-1). One-way ANOVA and Tukey tests showed that there was a significant difference (P<0. 01) between data obtained at each optimization step and the best glucose and di potassium phosphate concentrations for polyphosphate production were 5 and 0. 5 g l-1 respectively. Discussion and According to data obtained in this study، the best carbon source for cell growth and proliferation was not the best source for polyphosphate production and phosphorus removal from medium. Since there was no statically significant difference between 1، 2، and 0. 5 g l-1 concentrations for polyphosphate production، therefore، for industrialization and economization in the level of phosphorus consumption، 0. 5 g l-1 was the best phosphorus concentration.