Pseudomonas aeruginosa is one of the Gram-negative bacteria widespread everywhere in the Pseudomonadaceae family and is able to survive in a wide range of environments. The aim of this study was to determine of virulent exoU genotype and phylogenetic tree of P. aeruginosa isolates using real time PCRand Sanager sequencing. 206 clinical samples were collected from wounds and burns from both sexes. Samples were cultured on MacConkey agar, Blood agar and Cetrimide agar in order to obtain the bacterial isolates depending on their phenotypic characteristics; biochemical tests as diagnosed with Api 20E; Microgen GnA + B-ID and Molecular detection of exoU mutation of Pseudomonas aeruginosa isolates by using RT-PCR and Sanger method. The results showed that 12(24%) of (50) isolates were positive for ExoU gene.The ExoU gene were used to analyze the DNA sequence of local isolates and compare them with some standard global isolates according to the ExoU gene available in the GenBank database of the NCBI database. The results showed that the isolates were (98-100%) matched between local isolates and the international standard isolates. The results of the RT-PCR of the ExoU gene were used to detect mutations related to Pseudomonas aeruginosa, and the transitions showed the frequency of 5 mutations of the ExoU gene in all isolates. In addition, there were three types of mutations such as the predicted frame mutation effect was accompanied by only a transition 3, while 4 mutations were shown silent in all isolates. In addition, 2 mutations were shown to be mismatched. Neighbor joining phylogenetic tree for ExoU sequences which indicate one cluster divided into two subgroups. Group 1 showed sister group with P. aeruginosa strain BA7823 (India) reference strain from GenBank database which was used for determining the mutations and polymorphisms in the local isolates of this study.

• Collee J G, Fraser A G, Marmion B P, and Simmons A (1996). Practical medical microbiology. 14th ed. Longman. Singapore publishers (pet) ltd. Singapore.363-373.
• Forbes, B. A., Sahm, D. F., & Weissfeld, A. S. (2007). Diagnostic microbiology. St Louis: Mosby.‏
• Futuyma D J (2013). Evolution (3rd ed.). Sinauer. ISBN 978-1605351155.
• http://www.ncbi.nlm.nih.gov, National Center for Biotechnology Information, U.S. National Library of Medicine, 8600 Rockville Pike, Bethesda MD, 20894 USA.
• https://www.sigmaaldrich.com/technical-documents/articles/biology/sanger-sequencing.html, Sanger Sequencing Steps & Method,© 2020 Merck KGaA, Darmstadt, Germany and/or its affiliates. All Rights Reserved. Reproduction of any materials from the site is strictly forbidden without permission. Sigma-Aldrich Products are sold exclusively through Sigma-Aldrich, Inc. Site Use Terms | Privacy
• Kaszab, E., Szoboszlay, S., Dobolyi, C., Háhn, J., Pék, N., & Kriszt, B. (2011). Antibiotic resistance profiles and virulence markers of Pseudomonas aeruginosa strains isolated from composts. Bioresource technology, 102(2), 1543-1548.‏
• Lari, A. R., Azimi, L., Soroush, S., & Taherikalani, M. (2015). Low prevalence of metallo-beta-lactamase in Pseudomonas aeruginosa isolated from a tertiary burn care center in Tehran. International journal of immunopathology and pharmacology, 28(3), 384-389.‏
• Liew, S. M., Rajasekaram, G., Puthucheary, S. A., & Chua, K. H. (2019). Antimicrobial susceptibility and virulence genes of clinical and environmental isolates of Pseudomonas aeruginosa. PeerJ, 7, e6217.‏
• Lister, P. D., Wolter, D. J., & Hanson, N. D. (2009). Antibacterial-resistant Pseudomonas aeruginosa: clinical impact and complex regulation of chromosomally encoded resistance mechanisms. Clinical microbiology reviews, 22(4), 582-610.‏
• MacFaddin, J. F. (2000). Biochemical Tests for Identification of Medical Bacteria, Williams and Wilkins. Philadelphia, PA, 113.‏
• Mukadasi, B. (2018). Mixed Cropping Systems for Sustainable Domestic Food Supply of the Smallholder Farming Communities in Nakasongola District, Central Uganda. Canadian Journal of Agriculture and Crops, 3(1), 42-54.
• Noreen, H., Zaman, B., ur Rahman, A., & Hassan, W. (2016). Antioxidant and Antimicrobial Efficacies of Withania Coagulans Seed Extract against Pathogenic Bacteria and Fungi. The International Journal of Biotechnology, 5(3), 45-51.
• Ohno, M., Sakumi, K., Fukumura, R., Furuichi, M., Iwasaki, Y., Hokama, M., ... & Nakabeppu, Y. (2014). 8-oxoguanine causes spontaneous de novo germline mutations in mice. Scientific reports, 4(1), 1-9.‏
• Pazos, M. A., Lanter, B. B., Yonker, L. M., Eaton, A. D., Pirzai, W., Gronert, K., ... & Hurley, B. P. (2017). Pseudomonas aeruginosa ExoU augments neutrophil transepithelial migration. PLoS pathogens, 13(8), e1006548.‏
• Tacconelli, E., Carrara, E., Savoldi, A., Harbarth, S., Mendelson, M., Monnet, D. L., ... & Ouellette, M. (2018). Discovery, research, and development of new antibiotics: the WHO priority list of antibiotic-resistant bacteria and tuberculosis. The Lancet Infectious Diseases, 18(3), 318-327.‏Klockgether J, Cramer N, Wiehlmann L, Davenport C F, Tümmler B (2011). Pseudomonas aeruginosa genomic structure and diversity. Frontiers in Microbiology | Cellular and Infection Microbiology, 2(150), 1–18. https://doi.org/10.3389/fmicb.2011.00150
• Tadesse, A. and Alem, M. (2006). Medical Bacteriology. EPHTI. Gondar University.
• Todar K (2011). Pseudomonas aeruginosa. Textbook of Bacteriology. Science Magazine V. 304:1421.
• Wilfinger W, Mackey K, Chomczynski P. (1997). Effect of pH and ionic strength on the spectrophotometric assessment of nucleic acid purity.Biotechniques. Mar;22(3):474-6, 478-81.

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