فصلنامه زیست شناسی میکروارگانیسم ها
پیاپی 19 (پاییز 1395)

Viral hemorrhagic septicemia (VHS) is known as the most serious disease of rainbow trout in aquaculture industry. In this study, VHS virus glycoprotein genes were identified in farmed rainbow trout by RT-PCR test. Then nucleotide sequencing and phylogenetic analysis were performed on the viral isolates.
Suspected samples of VHS disease that collected from rainbow trout farms of Chaharmahal and Bakhtiari province were tested by RT-PCR method. Finally, the eight samples were identified as positive VHS strains and nucleotide sequencing was accomplished. Based on phylogenetic analysis, results of nucleotide sequences of glycoprotein gene were compared with other countries.
The results of present study showed that genetic variation in the glycoprotein gene was 0-7.4 percent. There was the highest similarity of glycoprotein gene of Iranian strains up to 100% with Germany strain (EU708818) and the lowest similarity was showed in Japan (AB672616) and Turkey (JF415091) strains with 60/92 percent in each.
Discussion and According to the genetic similarity of the VHS virus isolated from Iran with European strains, especially Germany and France, there is possibility of the relationship between the outbreak of VHS and eyed eggs imported from the mentioned countries.

Green microalga Chlamydomonas reinhardetii is concerned as a model system to study algal physiology, morphology, genetics and biofuel production. The aim of this study was optimization of the growth and starch production conditions. With this respect, the effect of pH and light intensities was investigated in alga Chlamydomonas reinhardetii.
The combination effect of these factors on growth and starch production was evaluated by design-Expert software and response surface method. The growth rate and starch content were evaluated using dry weight and spectrophotometric method respectively.
Regarding to the response surface curve, maximum growth rates and starch content were achieved at pH 7.4 and 8; and light intensity 7400 and 5300 lux respectively.
Discussion and Results showed that light intensity and pH play an important role in starch productivity of Chlamydomonas reinhardtii. Moreover, it was found that the response surface methodology was useful for enhancing the starch production in the algae.

drilling for exploration and production of petroleum requires the use of drilling fluid in order to lubricate and cool the drill bit. Oil based muds (OBMs) containing diesel, are the preferred drilling fluids because of their better lubricity. After mixing with drill cuttings, OBMs form complex industrial wastes containing hydrocarbons and brine. Bioremediation can be considered as a promising alternative to physicochemical methods for cleaning up drilling wastes because of its environmental friendly and cost effective properties. Due to high salinity and lack of active microflora in drill cuttings, developing halophilic consortia could be beneficial for enhancing bioremediation efficiency.
different oil-polluted and unpolluted saline soil and water samples have been used to enrich halophilic consortia able to degrade diesel hydrocarbons using a multistep enrichment technique. The growth capability of diesel-degrading consortia was evaluated by Bradford protein assay and standard plate count and growth curves were plotted. Furthermore, dominant strains of the consortia which were isolated from oil polluted saline soils were identified by molecular method.
Among 10 microbial consortia isolated from different environments, the best consortium was isolated from Qom oil-polluted soil sample which could grow very well on Bushnell Haas mineral medium containing diesel as the sole source of carbon. The bacterial count of the consortium was increased up to 1.88×1015 CFU/ml after 96h of incubation. Comparison of the growth curves indicated that in comparison to unpolluted samples, polluted ones reached the logarithmic phase more quickly and the final turbidity was more. 16S rRNA gene sequencing analysis revealed that the dominant strains of consortia isolated from oil polluted saline soil samples belonged to the genera of Dietzia, Microbacterium and Salinicola.
Discussion and Due to long term exposure of the polluted samples to oil pollution and high salinity, they have naturally been enriched by bacterial communities capable of degrading oil derivatives such as diesel in extreme conditions.

Rhizobia are bacteria that have a symbiosis with legumes. Rhizobia that have some capabilities like ACC deaminase and siderophore production in addition to nitrogen fixation are successful competitors in different environmental conditions for production of more nodules. The aim of this study was the screening of ACC deaminase and siderophore producing Rhizobia.
For screening of ACC deaminase producing rhizobia, all isolates were cultured on M9 medium with two different nitrogen sources, ACC and NH4Cl, and M9 medium without nitrogen source was considered as a control treatment. ACC deaminase producing Rhizobia were identified based on statistical comparison of growth rate on M9 medium with ACC against other mediums. Chrome Azurol S (CAS) liquid assay was used for siderophore producing Rhizobia. All treatments repeated two times and all data analyzed in the base of the completely randomized design. 16S rRNA gene sequencing was used for identifying the best ACC deaminase and siderophore producing strain.
14 isolates of Rhizobium were used for screening of ACC deaminase and siderophore producing isolates. Results showed that just R8 strain was isolated with the capability of ACC deaminase production. This isolate could have better growth on M9 with ACC than two other mediums. In the next experiment, it was determined that 8 isolates are siderophore producing rhizobia based on CAS liquid assay. The result showed that R12 isolate could produce more siderophore than others. According to 16S rRNA gene sequencing, R8 and R12 isolates identified as Sinorhizobium meliloti.
Discussion and The R8 strain that have the capability of ACC deaminase production probability can prevent adverse effects of ethylene during nodulation and abiotic stress. On the other side, R12 that are able to produce siderophore can prevent the growth of soil born pathogen in addition to facilitating nodulation and iron uptake by plants.

Shigella is a Gram-negative enteric bacilli that causes serious problems in advanced and developing countries. The purpose of this study was to evaluate colorimetric assay based on gold nanoparticles for rapid detection of S. flexneri using ipaH gene amplification products.
To identify the genus Shigella, the specific primers were used to amplify ipaH gene with polymerase chain reaction (PCR) method. Then, single-stranded probe was used to connect to the specific region of the PCR product. After addition of gold nanoparticles, the color changing was observed visually in the solution and imaged with scanning electron microscope (SEM). To examine the sensitivity of new method, a range of different concentrations of PCR products was used and the results were compared with gel electrophoresis method.
The hybridization of probes with amplified products of ipaH gene caused the accumulation of gold nanoparticles which resulted in color changing in the reaction solution from red to purple, but no color changing was observed in the negative control. Then, the accumulation of gold nanoparticles was approved with SEM.
Discussion and Because of the specificity of ipaH gene in S. flexneri, the colorimetric assay with high sensitivity can be used for specific detection of S. flexneri in the clinical specimen and food samples.

The accumulation of selenium oxyanions in the form of selenite (SeO3-2) in soil and water resources created increasingly worrying on human health and environment. The current project directed toward screening of selenite-resistant lactic acid bacteria for their potential use as safe biocatalysts in the selenite bio-remediation.
Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) values of selenite against isolated strains were evaluated using agar dilution method. Inhibitory effects of selenite against isolated strains were calculated employing well diffusion agar method. Turbidity testing was used to investigate bacterial growth. Identification of the potent bacterium strain was carried out based on bio-chemical tests and phylogenetic analysis. A colorimetric method using 3, 3-Diaminobenzidine hydrochloride has been developed for the microbial selenite removal.
Results indicated that Lactococcus raffinolactis strain seD2b exhibited the highest MIC (110 mM) and MBC (140 mM) values and also the lowest inhibitory effect with the average diameter of the inhibition zone (26.6 mm) in the presence of selenite. After 72 h incubation of the reaction medium containing 10mM of initial selenite concentration with resting cells of Lactococcus raffinolactis seD2b, the concentration of toxic selenite in the reaction medium decreased by 90.2% .
Discussion and Given the ability of Lactococcus raffinolactis strainseD2b in the remove and reduce selenite, screening of lactic acid bacteria as green bio-catalysts for their application in the bio-remediation of selenite and their economic importance on synthesis of selenium has been proposed.

Bacterial resistance to antibiotics increased the importance of developing novel antibacterial agents. Silver nanoparticles could be considered as one of the most important antibacterial agents. Regarding to the environmental concerns of widespread application of silver nanoparticles, the use of green methods for synthesis of the nanoparticles can lead to less concerns and to develop the nanoparticles with improved properties.
The ethanolic extract of Taxus baccata leaves was prepared and used in different concentrations for reduction of silver nitrate and synthesis of silver nanoparticles. The resulted nanoparticles were characterized by different methods including UV-Vis spectroscopy, Transmission electron microscopy, FT-IR, DLS, and Zetasizer. The antibacterial activity of silver nanoparticles was finally investigated using disk diffusion assay and minimum inhibitory concentration assay.
The ethanolic extract of T. baccata displayed high potential to synthesize high quality and stable silver nanoparticles with size below 50 nm. The nanoparticles displayed considerable antibacterial effect on all three tested bacteria. Minimum inhibitory concentration of 50 µg/ml obtained for S. pyogenes (ATCC 1447) and 25 µg/ml for E. coli (ATCC 25922) and S. aureus (ATCC 25923).
Discussion and The use of biological compounds especially plant extracts instead of dangerous and toxic chemicals for synthesis of silver nanoparticles could be considered as alternatives to reduce the environmental concerns of these nanoparticles. With respect to the considerable antibacterial activity of the nanoparticles synthesized with the present method, the nanoparticles can be used for different medical, food, and industrial applications.

Diesel oil is the major product of crude oil. This product is the main pollutant of environment and has hazardous effects on the environment. Bioremediation is the best method for remediation of this pollutant, because this method use nature potential for remediation and also this method has a low cost.
In this study, for isolation of diesel-oil degrading bacteria sampling performed from contaminated regions. The prevalent diesel-oil degrading bacteria were identified by biochemical and molecular methods. Some properties in these bacteria were analyzed such as: hydrophobicity and emulsification activity. The percentage of diesel-oil degradation was assayed for each strain by Gas Chromatography (GC) and qualitative test.
In this research 34 diesel-oil degrading bacteria were isolated. From these strains 10 strains were selected according to diesel-oil degradation. These strains belong to Achromobacter¡ Enterobacter and Klebsiella. The half percentages of diesel-oil were removed by these strains in 10 days of incubation. The best strain for degradation of diesel was Enterobacter cloacae strain L2 (72%). The highest hydrophobicity is related toKlebsiella oxytoca strain K2 (76%) and the best emulsification activity belongs to Achromobacter denitrificans strain B2 (58 %).
Discussion and The results of this research confirmed that diesel degrading bacteria have sufficient diversity in soil contaminated in Iran. These bacteria have good potential for remediation of diesel pollution from soil.

Polycyclic aromatic hydrocarbons (PAHs) are one of the most important pollutants that majorly originate from industrial production, transportation, refuse burning, gasification and plastic waste incineration and therefore must be degraded or removed. One of the safest and cheapest methods is the utilization of biologic processes such as bioremediation. In this method, microorganisms are employed to remove or reduce the pollutants toxicity. Bioremediation of anthracene as one of the tricyclic PAHs by a local bacterium that was isolated from wastewater of oil tankers station was investigated in this study.
Inorganic basal medium containing anthracene as the sole source of carbon and energy, biochemical tests and 16s rDNA gene analyses were applied respectively for bacterium isolation and identification. In wastewater analysis, the parameters such as total petroleum hydrocarbons (TPHs), aromatic hydrocarbons and biological oxygen demand (BOD) were measured. For anthracene degradation, different concentrations of anthracene (100, 150, 200, 250, 300 and 350 mg/l) were used. Gas chromatography (GC) and Gas chromatography-mass spectrometry (GC-Mass) were applied to determine anthracene residue and metabolites from anthracene degradation, respectively.
The current study showed the ability of Bacillus sp. Caspian1394to grow in wastewater and tolerate PAH pollutants with high concentration and also degrade anthracene. This bacterium degraded 100 and 350 mg/l concentrations of anthracene with 99.5% and 3.4% efficiency, respectively. Some of the metabolites derived from anthracene degradation were 1,4-methanoazulene, 2,5-cyclohexadiene, hexadecane, heptadecane, ocadecane and eicosane
Discussion and The studied bacterial strain can grow in wastewater with high contamination to hydrocarbons and tolerate high concentrations of different PAHs. It is a spore forming bacterium leading to its resistance to unfavorable conditions and also dispersion in different environments. Then, it can be proposed as a candidate in bioremediation of environments contaminated with anthracene.

Phenol and its derivatives are environmental pollutants commonly found in many industrial influents. Despite being toxic, phenol can be utilized by some microorganisms as sole carbon and energy sources.
In this research, halotolerant phenol biodegrading yeasts were isolated in GPY and MA media containing 250 ppm phenol and 5% NaCl. Among isolated strains, the best strain that was able to utilize phenol in presence of 5% NaCl was selected and identified by PCR of ITS gene. The resistance of the yeast isolate to different phenol concentrations and its toxicity besides phenol degradation at saline culture were investigated by spectroscopic measurement of residual phenol and cell dry weight.
In this research, 15 strains were isolated. Based on their ability for growth in different concentration of phenol and salt, ADH8 was selected as best strain. An inhibitory effect of phenol was observed at concentrations higher than 1250 ppm in presence of 5% NaCl, resulting in significant reduction of growth rates and phenol degradation. At concentration 750 ppm and 1000 ppm of phenol, all of the phenol was consumed after 24 h. Our results revealed that at the concentration of 1250 ppm of phenol, because of phenol toxicity on T. cutaneum, growth rate and phenol degradation was different. Higher lag phase and incubation time were observed in this concentration and total removal was detected after 48 h.
Discussion and The results indicate that the highly phenol-resistant halotolerant yeast represents a potentially promising means for biodegrading phenol pollution in wastewater treatment effluent and contaminated soil containing salt.

Nowadays, antimicrobial resistance among bacteria and other pathogenic microorganisms is a serious threat to management of infectious diseases. Therefore, investigation of new metabolite from new resources such as lichens are the subject of many research groups.
Acetone extract of two lichens of Glypholecia scabra and Rhizoplaca melanophthalm were prepared using a Soxhlet extraction. The antibacterial activities of the extracts were investigated on some Gram-positive and Gram-negative bacteria such as, Bacillus cereus, Bacillus subtilis, Staph aureus, Micrococcus luteus, Pseudomonas aeruginosa, Klebsiella pneumonia, Proteus vulgaris, Serratia marcessence, Escherichia coli using the Mueller Hinton Medium and Carbenicillin and Azithromycin standard antibiotics by Kirby’s method. Minimum inhibitory concentration and minimum bactericidal concentration of the extracts were determined.
Extract of Glypholecia scabra showed significant effects on Gram-positive bacteria such as, Bacillus cereus, Bacillus subtilis, Staph aureus, and Micrococcus luteus and also on Gram-negative Proteus vulgaris, respectively, with MIC values of 0.16, 0.26, 1.04, 0.65 and 0.11 mg/ml. Extract of Rhizoplaca melanophthalma showed inhibitory effects on Gram-positive bacteria Bacillus cereus, B. subtilis, Micrococcus luteus and Staphylococcus aureus , respectively, with values of 0.04, 0.08, 1.04 and 0.16 mg/ml but had no effect on Gram-negative bacteria.
Discussion and The antibacterial effect of Glypholecia scabra has not published before. The results of this research demonstrate the antibacterial effects of both Iranian native lichens. So acetone extract of the studied lichens maybe an alternative to chemical antibiotics that need to be studied more.

In recent years the use of rotifers as live food for rearing fish larvae and crustacean is very important. Rotifer has good characteristics such as size, nutritional value, slow swimming and these features are important in the development of early stage of aquatics. Growth and density are considered as important factors in the production of rotifers.
In this study, the effect of three different concentrations (1.5×106, 3×106, 4.5×106 cell/ml-1) of filamentous algae, Oscillatoria spp., on growth and reproduction of marine rotifer, Brachionus plicatilis was investigated. The rotifers were cultured in standard conditions in 500 ml plastic containers. The initial density of rotifers in different treatments was 30 ind/ml.
At the end of the fifth day, the highest density (453.66 ± 6.33) (M±SE) was seen in high concentration (4.5×106 cell.ml-1), which were significantly (PDiscussion and The present study showed that filamentus algae, Oscillatoria spp. can be used in culturing of marine rotifer and changing different concentrations can produce different density of rotifers.

Any point at which over time, the earth continuously exudes a mud-like substance, may sometimes be referred to as a "mud volcano". The mud produced by mud volcanoes is often a slurry of fine solids suspended in liquids which may include water, which is frequently acidic or salty, and hydrocarbon fluids. Mud volcanoes in the southeastern bank of the Caspian Sea and the North Eastern of Golestan province of the country are among the most important of mud volcanoes in Iran and therefore, were microbiologically examined. In this study biodiversity of aerobic and anaerobic bacteria, dominant species, was investigated.
Material and In order to investigate the microbial diversity of the mud volcanoes the samples were cultured in aerobic and anaerobic conditions. The common culture media were used for cultivating aerobic bacteria whereas the thioglycollate broth were used to culture anaerobic bacteria. In addition to study the halophilic trait, hydrolase enzymes and physiological characteristics of the isolates were evaluated.
Among all of the isolates twenty strains were selected for further study from which 15 strains were aerobes and 5 strains were facultative anaerobes. Most isolates were gram positive bacilli that are capable of producing hydrolases. These enzymes have potential applications in biotechnology. All isolates were able to grow in medium containing 0-15% NaCl and considered as the salt tolerant. In order to identify the facultative anaerobes, following polymerase chain reactions the sequencing and blast steps were done.
Discussion and A BLAST search enabled us to compare the 16S rRNA sequences of the strains with a database of sequences the homology study showed that the strains Akh.1, Akh.2, Akh.3, Akh.4 and Akh.5 share homology with Bacillus niacin (99.61%) and Clostridium bifermentas (99.74%), Proteiniborus ethanoligenes (94.9%), Clostridium ultunense (95.7%) and Clostridium methylpentosum (91.14%), respectively.

Common scab which caused by Streptomyces scabies and other closely related species is a serious disease of potato and other tuber crops such as beet, radish, and turnip and etc. Although, the potato is one of the most important food crop in Northwest of Iran and common scab disease is prevalent in these areas, there is no accurate information about the exact causal agent of potato common scab and also their strains distribution in the area.
Material and During 2010-2012 harvesting seasons, potato tubers with common scab symptoms were collected from different fields in East Azerbaijan and Kurdistan provinces. Collected isolates were characterized based on morphological and physiological and biochemical assays. Also PCR assay was performed using universal primer for Streptomyces genus and specific primer for S. scabies species. Pathogenicity assay was performed on potato slices and radish seedlings too.
Based on morphological and physiological characteristics and biochemical test, 32 bacterial strains were identified as Streptomyces. Pathogenicity assay were positive in 26 isolates. All selected 32 isolates were amplified a 1027-bp fragment with using universal primer for Streptomyces genus and 26 isolates were amplified a 475-bp fragment using a pair primers which specific for S. scabies and its closely related plant pathogenic species. The PCR assay for detection of taxtomine (txtA) showed that all selected 26 pathogenic isolates could produce expected 522-bp band which indicating a significant correlation between pathogenicity capability of isolates on potato slices and radish seedlings and existing of txtA gene.
Discussion and This is the first report of molecular identification of S. scabies in Northwest of Iran. The results demonstrated that there is significant distribution of this pathogen in the potato farms of Azerbaijan and Kurdistan provinces.

The endophytic bacteria are non-pathogenic organisms residing plant tissues which produced growth factors to increase metabolism, growth and resistance of plants to the pathogens and environmental tensions. The present study was aimed (i) to isolate and molecular characterization of endophytic bacteria from peppermint, chamomile, and asparagus (ii) to investigate antagonistic effect of isolates on fungal pathogens including Aspergillus niger, Alternaria alternata, Fusarium oxysporum and Mucor hiemalis (iii) to survey of isolates effect on plant growth characters.
The endophytic bacteria were isolated from root tissue of peppermint, chamomile, and asparagus. The antagonistic effect of endophytic isolates were investigated on fungal pathogens including Alternaria alternata, Fusarium oxysporum and Mucor hiemalis using agar well diffusion assay testing on Mueller-Hinton agar. The seeds of plants inoculated to the bacterial suspension and then were cultured in vases containing 300 g soil. The isolates effect was studied on plant growth parameters and analyzed by statistical software SAS 9.2.
A total of 6 endophytic bacteria (E1-E6) belonging Bacillus sp. were isolated from the plant root. The statistical and variance analysis indicated significance effect of isolates on fungal pathogens inhibition and plant growth improvement. The growth of peppermint, chamomile, and asparagus were showed high length of the plants which had been treated with the endophytic bacteria rather than the non-treatment of plants.
Discussion and In this study, the endophytic bacteria were isolated from peppermint, chamomile, and asparagus, then verified antagonistic effect of isolates on current fungal pathogens, also on plant growth improvement. Regarding the potential of endophytic bacteria, the organisms can be used as biocontrols in agriculture, increasing resistance of plants against fungal pathogens, environmental stresses and improving growth of plants.