الفهرس 
1. INTRODUCTIONOnly 14 pages are availabe for public view
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Abstract
Nowadays, there is an intense need for discovery of new bioactive agents for the treatment of human, animal, and plant pathogens. Marine bacteria have been recognized as an important resource for novel bioactive compounds. The secondary metabolites from marine bacteria, especially those with unique color pigments have diverse biological properties such as antibiotic activity. The aim of this work is to isolate local marine bacteria which are able to produce bioactive pigments and evaluate the antimicrobial activity of this pigment against different pathogenic microbes. As well as, study the morphological, biochemical and molecular characterization of the most active producer. characterization of the extracted pigment using chemical tools and maintaining of Optimized conditions for maximum production of this pigment. In addition to, molecular studies on selected strain including protein profile and detection of pigment biosynthetic gene. In the present study, pigmented marine bacteria from different marine environments were isolated. Out of twenty one water samples only fourteen pigmented bacteria were isolated. The antimicrobial activity of pigments produced by these isolates against some pathogenic microbes was estimated using a disc diffusion method `according to Kirby-Bauer bioassay technique. The pigment produced by isolate (M1) had the highest average of inhibition zone against the tested pathogenic microbes. The most potent strain (M1) was isolated from El-Max bay, Alexandria government, Egypt. This isolate was purified, maintained and stored for further studies. The isolate M1 subjected to colony characterization and biochemical identification. Based on these results, this isolate showed uniform biochemical characteristics of Pseudomonas aeruginosa. The identification was confirmed using 16S rRNA gene analysis. The amplification of 16S rRNA gene of this strain was performed using PCR technique. The PCR product of 16S rRNA gene was detected using agarose gel electropherises and gave 370 bp. The new sequence of this PCR product was compared using the BLAST algorithm with available 16S rRNA gene sequences from organisms in the GenBank databases. The nucleotide alignment and distance matrix showed high similarity value (99 %) with species: P. aeruginosa. The sequence was submitted to gene bank and the strain was identified as P. aeruginosa strain OSh1 and had NCBI accession number: KT032066. In this study, Pyocyanin pigment was extracted from culture supernatants of P. aeruginosa strain OSh1 by standard chemical methodes and characterized using Ultraviolet-Visible scanning spectrophotometer and GC-MS analysis. Many trails to enhance the production of bioactive pyocyanin pigment by P. aeruginosa strain OSh1 using different growth condition as; incubation period, different media, pH, temperatures, inoculums sizes and static-shaking condition during the growth were undertaking. In this study the antimicrobial activity of purified pyocyanin (Needle-like crystals) at different concentrations, was monitored against some pathogenic microbes.The results indicated that pyocyanin has a high antimicrobial activity against most of tested microbes. Molecular studies on pyocyanin producing marine P. aeruginosa strain OSh1including detection of the phenazine –modifying gene (PhzS) involved in biosynthesis of pyocyanin and protein profile also undertaken. PCR assay was developed to detect the phenazine–modifying gene (PhzS) in P. aeruginosa strain using PHZS-F and PHZS-R primers. The amplified fragment was detected and separated on agarose gel electrophoresis (1%), and the 448 bp band of the phenazine biosynthetic gene was detected and not found in E- Coli strain, which used as negative control.To confirm the PCR results, cellular proteins of P. aeruginosa strain OSh1were separated by SDS-PAGE. Protein profile of P. aeruginosa strain OSh1 that growed on King’s A medium gave eight protein bands among of which 36 KDa and 43 that was corresponding to two enzymes encoded by two phenazine-modifying genes PhzM and PhzS, respectively. These bands were not found when P. aeruginosa strain OSh1growed on LB medium which used as negative contro.