فصلنامه زیست فناوری میکروبی
پیاپی 7 (زمستان 1389)

The properties of heavy crude oil present serious operational problems in heavy crude oil production and downstream processing. Heavy crude oil is extremely viscous and contains high concentration of asphaltene. In this research microorganisms capable of growing on asphaltene and using asphaltene as sole carbon and energy source were isolated and characterized. For this propose, oil contaminated soil from waste water treatment plant of Tehran Refinery as source of microorganisms was used. After adaptation of microorganisms with asphaltene, microorganisms were isolated and purified by cultivation and dilution. Subsequently microorganisms were characterized by PCR method. Isolated microbes are Bacillus Sp. PBCC13, Brevibacillus sp. PBCC14 and Serratia sp. PBCC12. They could be used for bioupgrading of crude oil and asphaltene.

Trichophyton rubrum is a wild spread fungi which cause the Dermatophytosis and approximately 80-93% of chronic or recurrent/recurdescent dermatophyte infections are created by trichophyton rubrum. Tricophyton proteins are members of different enzymatic families and dual role of these proteins in fungal pathogenesis as well as their allergenic behavior. Among trichophyton rubrum allergens, Tri r 2 and Tri r 4 can be mentioned. The present research aims to investigate the allergenic behavior of trichophyton rubrum and microsporum canis. Initially, the fungi trichophyton rubrum was cultured on PDA medium. Their conidia were subsequently gathered by sterilized PBS. DNA was extracted and PCR experiment was fulfilled using the specific primers Tri r4. Additionally, Guinea pig was used to study the allergenic behavior of the fungi trichophyton rubrum and microsporum canis. To confirm that the fungi were allergenic, total IgE antibody was assayed by ELISA method. Also, in CBC test performed on Guinea pig blood samples, percentage of blood cells especially eosinophils were evaluated. Conidia were confirmed by using PCR test for Tri r4. Results of allergenic behavior indicated that more IgE antibody, also neutrophil and eosinophil were produced in the Guinea pig in which trichophyton rubrum was inoculated and has a significant different compared to control group, whereas no significant different was observed in comparison with control group in the Guinea pig in which microsporum canis was inoculated. The present research indicates direct effects of allergens in enhancing titer of IgE antibody.

HBsu protein in Bacillus subtilis is a member of histone like protein family HU. The protein has an important role in structure and function of bacterial nucleoid. In sights of its importance and similarity between the members of HU family, in this study the purified expressed HBsu, as an immunogen, was injected in rabbit until the produced polyclonal antiserum was used in the next research about identification of the other HU family members. The recently expressed HBsu protein from hbs gene of Bacillus subtilis ATCC6633 was injected in two male rabbits according to a twenty-day schedule and immunoblotting was done for antiserum production checking. After twenty days, the antiserum against HBsu was produced and the presence of the antiserum was confirmed by immunoblotting. Also, extracted histone like protein from Halobacillus karajenesis was used for checking of its ability in identification of HBsu like protein that the answer was positive. The used method was rapid and with high ability for antiserum production against HBsu protein. The similarity between Halobacillus karajenesis HU and Bacillus subtilis HBsu is enough for the identification of this HU by antiserum against HBsu.

Heavy metals pollution has become one of the most serious environmental problems today. In the present study, different parts of bacteria (exopolymer, live and dead biomass) were used for adsorption of Zinc and Lead. The bacteria used in these experiments were Azotobacter and Rhizobium. Azotobacter and Rhizobium exopolymers were used for adsorption of heavy metals. Metals concentration was measured using a Varian AA-10 atomic absorption spectrophotometer. Azotobacter produced greater exopolymer in exponential growth phase, however Rhizobium in stationary phase had maximum exopolymer production. The zinc and lead adsorption in Azotobacter exopolymers was 10% greater than Rhizobium exopolymers. Biosorption capacity of live biomass of Azotobacter and Rhizobium for zinc and lead were 46% and 33.1% respectively, but about lead adsorption there was no significant difference between live biomass of bacteria. In zinc adsorption there was no difference between dead biomass of Rhizobium and Azotobacter. The amount of lead in 100 ml of urine, river water and well water was measured by using exopolymer biosensor. The Azotobacter exopolymer from 100 ml of urine, river water and well water adsorbed 0.1, 0.25 and 4 mg of Pb+2. g-1 exopolymer respectively, but Rhizobium exopolymer adsorbed only 0.8 mg of lead from 100 ml of well water. The result showed that exopolymer biosensor measures the small amount of lead in the environment.

The most important aspects of probiotic microorganisms are their useful effect on human health. The goal of this paper is to examine the Supernatant antimicrobial ability of separated probiotic bacteria from yogurt and probiotic tablet on the bacterial pathogen agents. In this study, it was isolated probiotic bacteria strains from yoghurt and probiotic tablet, was used and explored with the cooperation af biochimical method. Antimicrobial effects of these bacterial culture supernatants were evaluated on pathogen bacteria strains by using disc diffusion agar and well diffusion agar methods. To reduce the error test were repeated three times and the inhibitory zone diameter was measured, their antimicrobial ability was compared with each other. Six kinds of probiotic bacteria were determined from the sample of yogurt and probiotic tablet. These bacteria had shown a good antimicrobial ability against seven pathogen bacteria. The inhibitoriest effect was shown whit lactobacillus acidophilus of tablets against bacillus cereus according to well diffusion agar with average of inhibitory zone diameter 14mm. Also in the comparison between two method of well diffusion agar and disk diffusion agar, the well diffusion agar was the far more sensitive than disk diffusion agar method. According to this study the produced metabolite from probiotic bacteria can prevent the growth of pathogen bacteria which shows the positive role of this class of bacteia in human health and it would be suggested that is class of bacteria which are abundance in dairies, should be used more.

Antimicrobial properties of silver and copper nanoparticles were examined on B.cereus PTCC(1247), B.subtilis PTCC(6633). Active chlorine in sodium hypochlorite was causing the antimicrobial properties. The purpose of this study is the bactericidal effect of metal nanoparticles has been attributed to their small size and high surface to volume ratio, which allows them to interact closely with microbial membranes and is not merely due to the release of metal ions in solution. The antimicrobial effect of nanoparticles were quantified based on diameter of inhibition zone in the disk diffusion tests and well diffusion tests conducted in plates and by determining the minimum growth inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of nanoparticles in liquid batch cultures. Antimicrobial effects of nanoparticles and sodium hypochlorite were studied by counting colonies in different hours and dilution with cultured purplate and two layers methods. In this study was observed that B.subtilis depicted the highest sensitivity to nanoparticles compared to B.cereus and was more adversely affected by the silver nanoparticles. So that B.subtilis (MIC) of silver nanoparticles was measured 7 ppm and the nanoparticles of copper was measured 50 ppm, and sodium hypochlorite was measured 700 ppm. B.subtilis spores were killed in less dilution of antimicrobial substances and in zero time. B.subtilis spore and vegetative cell were more sensitive than the silver nanoparticles. The results indicated silver nanoparticles were more effectiveness compared to other antimicrobial substances that used in this stduy.

The presences of Cyanobacteria make numerous problems and its growths due to pollutes water with cyanotoxins that endangers human and aquatic health. Different species of cyanobacteria are known as producers of intracellular and extracellular metabolites with different biological activities like inhibiting and killing fungi and bacteria. The antibacterial effects of isolated cyanobacteria s strains from water sources were studied after preparing methanol extract with concertrations of 50 µl, 100 µl. ََAntibacterial properties of exteracts were determined by dilution in pipe method and its minimum inhibitory concentration was tested against some human pathogens like Staphylococcus aureus, Salmonella typhi, Bacillus cereus, E. coli and Proteus vulgaris with concentration of 0/0125g/ml and 0/025g/ml cause inhibition and death of Bacillus cereus, Proteus vulgaris and Salmonella typhi respectively and methanol extracts of Oscillatoria sp 2 with concentration of 0/0125g/m and 0/025g/ml can inhibited and killed S. typhi strains. This present survey shows methanol extracts of Oscillatoria has the more effects on pathogenic bacteria, but clinical usage needs more researches on mechanisem of function of cyanobacterial influential combinations on microbe factors.

Bioleaching is an economic process in which by using proper bacteria. One of the most important bacteria in this field is Acidithiobacillus ferrooxidans. This bacterium has an optimum growth in temperature interval 25-35 degrees centigrade and it has no desirable growth in lower than 20°C temperature. The main purpose of this research is to adapt these bacteria in lower temperatures, so that bioleaching process can be performed in all seasons of year including cold seasons. In this research, At.ferrooxidans is extracted from Pregnant leaching solution (PLS) of Sarcheshmeh copper mine and inoculated in TK and 9K media in 30°C. Then in order to adapt of At.ferrooxidans in low temperatures, this bacteria was enriched gradually at temperatures 25, 22, 18, 15, 12 and 4°C. In order to ensure the viability of bacteria at low temperatures and performing bioleaching process, the growth of At.ferrooxidans and ferric iron concentrations was measured by a spectrophotometer. The concentration of Cu (II) has been measured by Neocuprion at that temperature. In order to confirm At.ferrooxidans growth at 4°C extracellular proteins was measured by Bradford methods after six days. DNA of At.ferrooxidans strain has been extracted and region genes of 16SrDNA were amplified by PCR and gene sequencing was done. At.ferrooxidans has logarithmic growth at low temperatures such as 25, 22, 18, 15, 12 and 4 °C and is able to oxidize ferrous iron to ferric iron in bioleaching process and release Cu II in soluble form. It is shown that bacteria has the best growth at 25°C after 18 days and the rate of growth decrease with temperature reduction. Comparing the results of nucleotide sequence superior strains with the related strains showed that the bacteria is Acidithiobacillus ferrooxidans and was named as AL strain. At.ferrooxidans is able to adapt and perform bioleaching at low temperatures. Thus, according to the results, the process of iron oxidation and bioleaching at low temperatures is possible and precious metals such as gold and copper can be extracted at low temperatures.