فصلنامه زیست شناسی میکروارگانیسم ها
پیاپی 22 (تابستان 1396)

Cellulose is the most abundant biopolymer in the world. Cellulase, including endoglucanase, cellobiohydrolase and beta-glucosidase, catalyzes the hydrolysis of cellulose. Released glucose from enzymatic hydrolysis of cellulosic biomass is used in different biotechnology fields.
In this study, seven different Trichoderma species were obtained from Persian Type Culture Collection (PTCC) and in order to select the best ones, cellulase activity of native strains was determined. Sodium salt of carboxymethyl cellulose (CMC-Na), Avicel and cellobiose were used for endoglucanase, cellobiohydrolase (exoglucanase) and cellobiase (beta-glucosidase) assays, respectively. Kinetics of cellulose production was evaluated for the selected strain. Finally, random mutagenesis with 0.2 M sodium nitrate was done.
Among 7 different fungal species, Trichoderma parceramosum PTCC 5140 was selected as the best strain with the highest cellulase activity. This strain, by production of 0.182 U/ml of endoglucanase, 0.538 U/ml of exoglucanase and 0.109 U/ml of cellubiase, showed the highest amount of all three constituents of cellulolytic complex. Random mutagenesis and mutant selection of this strain caused to isolate 4 stable mutants that were able to produce 2 to 11 fold more enzymes compared with the parent strain.
Discussion and Evaluation of cellulase production in mutant strains of Trichoderma parceramosume PTCC 5140 showed that use of chemical mutagenesis with 2 to 11 fold increasing in enzyme activity is a potent method to improve cellulase complex activity. In the current study, obtained mutant strains could be introduced as a potent cellulase producer for further studies in bioconversion processes.

one dimensional nanocrystals especially nano-rods have attracted a great deal of attention due to their unique properties and wide applications in industry. Chemical and physical methods which are currently used to produce zinc oxide nano- rods, often leave toxic chemicals on surface of nanoparticles limiting their applications for health and medical purposes. Therefore, biological synthesis of zinc oxide nanoparticles has been considered as an environmentally friendly process and a potential alternative to chemical and physical methods.
Nano-rods of zinc oxide were produced by Xanthomonas campestris using zinc nitrate hexa hydrate as substrate, in a shaker incubator at 37 ° C and pH 7. The powder produced was then calcined at 600 ° C for 2 hours after drying. The synthesized ZnO nanoparticles were characterized using FTIR, XRD, SEM, EDX and UV–vis spectroscopy.
FTIR analysis was used to identify functional groups involved in the biosynthesis of ZnO NPs. The peak observed at 563 cm-1 corresponds to the stretching vibrations of ZnO NPs. XRD analysis revealed that the hexagonal ZnO nanoparticles synthesized were pure and crystalline in nature. The morphology and size of the powder were investigated using SEM analysis and the results showed that ZnO nano-rods have a diameter ranging from 122–200 nm with an average length about 300 nm. EDX analysis was performed for determination of the elemental composition and purity of samples. The recorded EDX spectrum revealed the high purity of the synthesized ZnO nano-rods without detection of any impurities. The absorption peak at 376 nm indicating the presence of zinc oxide nanoparticles was further confirmed by UV-Vis spectroscopy.
Discussion and The current research work describes a low-cost, unreported, nontoxic, simple, safe and eco-friendly method for the biosynthesis of zinc oxide nanoparticle using xanthomonas campestris as the reducing and capping agent.

Safflower straw is one of the agricultural residues that currently have no specific use. So, it can be used as a cheap source for biofuel production.
Alkaline pretreatment with NaOH was used to improve enzymatic hydrolysis, ethanol, and biogas production from safflower straw. The pretreatment was performed with 8% (w/v) NaOH solution at two different temperatures (0 and 100 °C) for 10-60 min. The liquid fraction of the pretreatment was subjected to anaerobic digestion for methane production. Solid fraction obtained from the pretreatments was used for bioethanol and biogas production in a parallel process. Therefore, two routes were followed for energy production from safflower straw: (1) ethanol production from solid fraction (2) biogas production from liquid fraction. In order to optimize the production of biofuels and compare different pretreatment conditions, the gasoline equivalents were calculated based on one tone of safflower straw after all pretreatment conditions. Statistical validation of the results was carried out using Minitab 16 software.
The highest glucose and ethanol production yields were obtained after the pretreatment at 100°C for 60 min. The glucose yield from untreated straw was only 20.6% and improved to 84.5%, whereas the corresponding ethanol yield was improved from 10.8% to 83.2%. The best improvement in methane production from solid fraction was achieved by pretreatment at 0 °C for 60 min, and resulted in 191.4 ml/g VS which was 99.6% higher than methane yield obtained from untreated straw. The highest gasoline equivalent was 124.9 L obtained after pretreatment at 0 oC for 60 min.
Discussion and Enzymatic hydrolysis and ethanol yields from the safflower straw were improved more by the pretreatment at high temperature. However, low temperature is desirable for methane production.

Halophiles and Halotolerant microorganisms are some of the extremophiles that are able to grow in medium containing sodium chloride and have dapted to life in salinity environments. Halophiles bacteria in saline soils by maintaining the food chain, decomposition of organic matter and improvement of soil structure and fertility improve soil conditions.
In order to isolate the halotoletant bacteria, from the Halophyte rhizosphere, four desert areas in Golestan province were sampled. To check the Extremophile of isolates, their resistance was tested for resistant to salinity, drought, temperature and PH. Also, plant growth promoting traits were measured.
Fromforty-five strains which were isolated, three strains (G3, G6 and G14) have demonstrated the ability of resistance to 35% salt. Isolates G6 and G3 phosphate solubiliziation power of 301 and 201 ppm, respectively. Isolated G6 micrograms produced auxin 20/7 Mg/ ml. G14 and G6 grow at 50 °C, pH = 10 and osmotic potential -0 /7MPa. While G3 strain grows at 50 °C, pH = 7/ 5 and osmotic potential -0 /49. The three strains of the bacterial genera Bacillus and Pseudomonas, respectively.
Discussion and In this study, isolates due to the growth in concentrations of salt and saturated salt tolerance of extreme environmental conditions and are likely halotolerant or halophile bacteria and its potential for use in various fields of biotechnology including biotech, industrial enzyme production and biological fertilizers for saline soil improvement.

Biodiesel as a biofuel, with renewable, biodegradable and free of polycyclic aromatic hydrocarbons properties, could be derived from fatty acids of the cells of the animals, plants, algae and bacteria. So, a wide range of approaches have been considered to increase the oil production in biodiesel resources. In this regard, biotechnology could be provided by promising vision based on application of corresponding molecular mechanisms.
Bearing in mind, a comprehensive survey on molecular mechanisms and key enzymes which are involved in the production and accumulation of the lipids in biodiesel resources have been considered based on literature study and in-silico investigation. Our survey led to the introduced diacyl glycerol acyl transferase (DGAT), waxester synthase/diacyl glycerol acyltransferase (WS/DGAT), oleosin, MLDP and TadA as the effective enzymes and proteins in lipid production and accumulation in selected biodiesel organisms. In-silico investigation has been performed via InterProScan 5¡ Motif scan¡ Conserved Domain¡ ProtParam¡ TMHMM¡ GC content calculator¡ NetNGlyc¡ NetPhos¡ Sulfinator¡ Protein Blastand MEGA6 programs for characterizing the structure, functions and homology survey of the selected sequences.
An investigation on the structure of the corresponding genes of the selected enzymes, led to reveal their various feature in the length, GC content as well as in intronic properties. On the other hand, this characterizing on the selected enzymes and proteins disclosed post-translational modifications in all of them, as well as their localization in the cells. Moreover MBOAT, DAGAT, UPF0089, Oleosin and apolipo protein have been revealed in their context as critical domains. On the other hand, homology survey of the selected enzymes and proteins led to introduce the Verniciafordii, Ricinuscommunis, Dunaliella parva , Thalassiosira pseudonana, Saitoella complicata, Rhodococcus imtechensis and Rhodococcus wratislaviensis species, as new sources of biodiesel with possible capability for lipid production.
Discussion and Overall, this survey provides a series of motifs and domains in biodiesel process, as well as introducing several organisms with potency in biodiesel production, which could be more examined in an experimental condition.

Elimination or reduction of petroleum hydrocarbons from natural resources such as water and soil is a serious problem of countries, particularly oil-rich countries of the world. Using white rotting fungi compost for bioremediation of soils contaminated by petroleum hydrocarbons is effective. The aim of this study is molecular identification and potential of anisolate of white rot fungi in bioremediation of petroleum contaminated soils.
Spent compost of white rotting fungi was inoculated with petroleum contaminated soil into 3%, 5% and 10% (w/w). Treatments were incubated at 25-23 °C for 3 months. Reduction of petroleum hydrocarbons in treated soil was determined by gas chromatography. Ecotoxicity of soil was evaluated by seed germination test.
Based on the genome sequence of 18s rRNA, it is revealed that this isolate is Ganoderma lucidum and this isolate is deposited as accession KX525204 in the Gene Bank database. Reduction of petroleum hydrocarbons in soil treated with compost (3, 5 and 10%) ranged from 42% to 71%. The germination index (%) in ecotoxicity tests ranged from 20.8% to 70.8%. Gas chromatography results also showed a decrease in soil Hydrocarbons compounds.
Discussion and The compost of Ganoderma lucidum, a white rot fungus, has a potential ability to remove petroleum hydrocarbons in contaminated soil. Removal of hydrocarbons was increased with increase in compost mixed with contaminated soil. Petroleum contaminated soil amended with spent compost of G.lucidum 10% during three months is appropriate to remove this pollutant.

Using rapid detection methods is important for detection of plant pathogens and also prevention through spreading pests in agriculture. Citrus brown spot disease caused by pathogenic isolates of Alternaria alternata is a common disease in Iran.
In this study, for the first time a PCR based molecular method was used for rapid diagnosis of brown spot disease. Nine isolates of A. Alternata were isolated in PDA medium from different citrus gardens. The plant pathogenic activity was examined in tangerine leaves for isolates. Results showed that these isolates are the agents of brown spot disease. PCR amplification of specific ACT-toxin gene was performed for DNA extracted from A. alternata isolates, with 11 different fungal isolates as negative controls and 5 DNA samples extracted from soil.
Results showed that A. alternata, the causal agent of brown spot disease, can be carefully distinguished from other pathogenic agents by performing PCR amplification with specific primers for ACT toxin gene. Also, the results from Nested-PCR method confirmed the primary reaction and the specificity of A. alternata for brown spot disease. PCR results to control samples of the other standard fungal isolates, showed no amplification band. In addition, PCR with the DNA extracted from contaminated soils confirmed the presence of ACT toxin gene.
Discussion and Molecular procedure presented here can be used in rapid identification and prevention of brown spot infection in citrus gardens all over the country.

Different studies undertaken in the animal modeling show that Interferon-gamma deficiency impairs healing process of Leishmania infection. It seems that the level of Interferon-gamma production could also affect the healing duration of Leishmania lesion in humans. The current study aims to investigate the possibility of Interferon-gamma application for recognition of cases afflicted by non-healing Leishmaniasis.
Peripheral blood mononuclear cells (PBMCs) of 32 patients, who were afflicted by healing or non-healing Leishmaniasis, were isolated and the levels of interferon-gamma were determined, using ELISA method. Afterwards, the cut-off point of interferon-gamma to identify patients afflicted by non-healing Leishmaniasis was calculated through ROC-Curve analysis. Furthermore, Leishmanin Skin Test (LST) was performed for every patient.
Levels of Interferon-gamma produced by PBMCs stimulated with Soluble Leishmania Antigen (SLA) or Phytohemaglotinine were significantly higher in healing patients, compared with non-healing individuals (p Discussion and Interferon-gamma deficiency could be considered as one of the causative factors in non-healing Leishmaniasis in humans. In fact, Interferon-gamma deficiency can identify some patients afflicted by non-healing Leishmaniasis.

Sulfur oxidizing bacteria are important in agriculture. The aim of this study was to identify the sulfur oxidizing bacteria in Sarcheshmeh (Kerman, Iran) agricultural soil.
Sampling was conducted from agricultural soil in Kerman province, Iran. After enrichment, samples were purified in the basic mineral culture containing sulfur as the only source of energy. Sulfur oxidation capability was evaluated based on the variation of pH and sulfate content and then strains of sulfur oxidizing were isolated and identified using morphological characteristics and phylogenic techniques. The best strains were selected and were evaluated for variation of pH and sulfate content by isolates in different pH and temperature.
Six sulfur-oxidizing strains were isolated and identified which had similarity to Thiobacillus and Starkeya genous. Isolates of 103, 129, 190 and 158 were mesophyll and neutrophil, but isolates of 192 and 156 were thermophile and alkalophill. Of these isolates, isolate 103 was 99.86% similar to Starkeya novella, identified as superior isolate, due to the most sulfur oxidation capability in the synthetic mineral medium containing sulfur.
Discussion and Isolate 103 was able to grow in neutral to slightly alkaline conditions with pH 7.5. Most of the growth of this isolate was at 30 ° C as aerobic. This isolate was facultative chemolithotroph and the results of this research showed capacity of the isolate for oxidizing sulfur, so that it could be used as a practical isolate in bio-fertilizers for neutral to alkaline and saline conditions in agriculture.

Polyhydroxyalkaniates (PHAS) are bacterial polymers that are formed as naturally occurring storage polyesters by a wide range of microorganisms usually under unbalanced growth conditions. Mechanical properties of PHAS make them suitable replacements for petrochemically produced bulk plastics (polyethylene, polypropylene etc.), but in contrast to these commodity plastics PHA are completely degradable to carbon dioxide and water through natural microbiological mineralization. PHAS can be produced by biotechnological processes under controlled conditions. Polyhydroxybotyric acid (PHB) was the first of thePHAS discovered and is the most abundant polyester found in bacteria.
First, a rich bacterial suspension prepared from poly(3-hydroxybutyrate), then the cells were broken down and ensure the absence of live cells.The cell free enzymes were added to M9 medium.Then the sample turbidity was measured at a wavelength of 600 nm and incubated at 30°C and centrifiuged at 120rpm. Next, the amount of PHB as well as solution turbidity were measured and compared with control. The physical and chemical analyzes of extracted PHB samples such as UV absorbtion, FTIR, HPLC and HNMR assesed and compared with standard sample.
In twelve days this experiment was conducted, from the third day, the amount of polyhydroxybutyrate was measurable and the highest polyhydroxybutyrate was gained on the tenth day. The level of PHB was constant. Moreover, the level of glucose was decreased gradually till the tenth day. By Physico-chemical analysis, production of polyhydroxybutyrate was approved.
Discussion and In this experiment, the reaction efficiency was 37.5 percent. Probably the rest of glucose could be changed to other intermediate compounds by other enzymes in the sample.Therefore, by purification of the enzyme, usage of specific substrate and optimization of conditions outside the cell could help the production of polyhydroxybutyrate continually. Production is very cost-effective, by this method.