The effects of nutrients and physical conditions on phytase production were investigated with a recently isolated strain of Aspergillus tubingensis SKA under solid state fermentation on wheat bran. The nutrient factors investigated included carbon source, nitrogen source, phosphate source and concentration, metal ions (salts) and the physical parameters investigated included inoculum size, pH, temperature and fermentation duration. Our investigations revealed that optimal productivity of phytase was achieved using wheat bran supplemented with: 1.5% glucose. 0.5% (NH 4 ) 2 SO 4 , 0.1% sodium phytate. Additionally, optimal physical conditions were 1 × 10 5 spore/g substrate, initial pH of 5.0, temperature of fermentation 30˚C and fermentation duration of 96 h. Overall, a 34% improvement in phytase activity was achieved by using the optimal conditions.
In order to evaluate the efficiency of using Polymerase Chain Reaction (PCR) in the identifications of microorganisms causing microbial keratitis, 20 corneal scraping samples were collected from patients who attended the Eye Casualty Unit at the Southampton General Hospital in the United Kingdom. Samples cultured on blood agar and chocolate agar incubated at 37 ͦ C for 24hrs and on sabrouad agar at 28 ͦ C for one week. PCR procedure was performed with the primer paired that targeted to the 16S rRNA for bacterial species and 18S rRNA gene for fungal species, in addition to the species specific primer for the most common microbial keratitis causatives microorganisms. Results in the regards showed that out of the 20 presumed cases of keratitis, PCR showed positivity in 75% of them, from these 55% were due to the fungal infection and 20% of the cases indicated that the keratitis belonged to bacterial infections: In comparison, only 25% of positivity was obtained by the cultural method. The species specific primer showed that half of the 20% bacterial infection cases were caused by S. aureus and the other 10% referred to S.epidermidis infection. While the candida albicans primer gave a positive result only in 72% of the original percentage (55%), the rest 28% may belong to the other fungal infection. Depending on the above results, it can be concluded that PCR not only proved to be an effective rapid method for the diagnosis of bacterial and fungal keratitis, but was also more accurate and sensitive method than the culture methods.
Lettuce is one of the most important edible plant worldwide. At the time that lettuce is the candidate plant to carry the foreign vaccine gene for human. The B subunits of toxin of Vibrio cholerae (CTB) are candidate vaccine antigens. This research was conduct to express CTB gene in lettuce chloroplast. Genes required in this study were obtained by polymerase chain reaction (PCR) technique using specific forward and reverse primers, and these genes were CTB, BADH, prrn promoter and many other regulatory genes. Some of these genes were isolated from their hosts and some were obtained from previous work available at Daniell laboratory. All these genes beside many techniques for ligation, extension, sequencing, orientation confirmation were used to construct the cassette vector pLS-BADH-LS-CTB which carries the gene of interest. In this work the CTB gene with BADH gene were transferred to the chloroplast of lettuce plant and selection of transgenic plant was performed on the MS medium containing BA and NaCl without any antibiotic selectable marker. Integration of an unmodified CTB-coding sequence into chloroplast genomes (up to 1000 copies per cell) resulted in the accumulation of up to 6.2% of total soluble lettuce leaves protein as functional oligomers (620-fold higher expression levels than that of the unmodified CTB gene expressed via the nuclear genome). PCR and Southern blot analyses confirmed stable integration of the CTB gene and BADH gene into the chloroplast genome in addition to the integration in the right orientation and in specific region between trnaI\trnA.Western blot analysis showed that the chloroplast synthesized CTB assembled into oligomers and were antigenically identical with purified native CTB.
This study was conducted to determine the role of lipopolysaccahride-liposome conjugate (LPS-LIP) as a potential vaccine against shigellosis in mice by determining the IgG titer. One hundred stool samples were collected from patients with diarrhea from different hospitals in Baghdad city during the period December 2011-May 2012. Thirty isolates were suspected to be Shigella. Four isolates of Shigella flexneri were obtained after performing some biochemical tests and Api System. Antibiotic sensitivity test was carried out and results revealed that all isolates were resist to some antibiotics used in this study. One isolate was selected for LPS extraction by phenol hot water extraction method. Chemical characterization of the extracted LPS revealed that the carbohydrate content was 2.34 mg/ml, while the protein concentration was 0.52µg/ml. Partial purification of the extracted LPS was carried out by using gel-filtration chromatography on Sephacryl S-200 and results showed that three peaks were obtained and protein, carbohydrate concentration were estimated for each peak. The second peak observed to have the highest carbohydrate content was (25%). and the lowest contaminated protein was (0.001%). The partial purified LPS was analyzed by Sodium Dodecyl Sulphate Gel Electrophoresis (SDS-PAGE) and results reveled that two bands with molecular weights 100 and 150 KDa were present. Band with MW 100 kDa represented LPS. IgG titer was estimated by Elisa technique. A significant increase in IgG titer was recorded in mice treated with conjugate after infection with S. flexneri.
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