جداسازي و شناسائي باكتريوفاژهاي اختصاصي سودوموناس آئروژينوزا و بررسي اثرات ضد باكتريايي آن¬ها در شرايط آزمايشگاهي

Isolation and identification of Pseudomonas aeruginosa phages and in-vitro eva‎luation of their antibacterial effects.

زيست شناسي سلولي مولكولي و ميكروبيولوژي

The skin is one of the largest organs of the body, which has several vital functions, including: maintaining homeostasis, regulating body temperature, immunological functions, metabolic function (vitamin D synthesis). It also provides primary protection against infections by acting as a physical barrier.When this barrier is damaged, pathogens can directly enter the body and cause infection. Despite the progress in wound management in recent years, burn infection is still one of the main causes of death. Although the leading bacteria in burn wounds is Staphylococcus aureus, the main cause of death is multi-drug resistant bacteria, including Pseudomonas aeruginosa. Due to the increase of antibiotic resistance, the need for alternative methods to control infections is felt more than before. Today, bacteriophages are considered as a new and effective tool in the treatment of microbial infections. In the present study, isolation and identification of Pseudomonas aeruginosa strains causing burn infection from patients referred to Imam Musa Kazem Accident and Burn Hospital were performed using biochemical and molecular methods. Then the antibiotic resistance of the isolated bacteria was investigated using disk diffusion method. Acute bacteriophages against Pseudomonas aeruginosa bacteria were isolated from urban and hospital wastewater in Isfahan province and their characteristics such as host range, morphology, EOP, stability in difference temperatures, pH values and salt concentrations, MOI and genome digesting pattern were determined. The killing activity of the studied phages in different concentrations against specific bacteria was investigated in laboratory conditions.Also, the antibacterial effects of the phage was eva‎luated in Galleria mellonella larvae. The genome of specific phage vB-PaS-HSN4 was sequenced and its similarity with other phages was analyzed by Mauve, Easyfig, and Circolleto software. The genetic map of the phage genome was drawn using Linear genome plot and CG Viewer software. MEGA 7.1 software was used to draw the phylogenetic tree and determine the kinship relationship. The results of the drug resistance of the isolated strains showed that the isolated strains has the highest resistance to piperacillin and levofloxacin and the lowest resistance to amikacin. Based on the morphological characteristics, the phages isolated against Pseudomonas aeruginosa belonged to the formerMyoviridae, Podoviridae, and Siphoviridae families that were recently categorized as Caudoviricetes class. The isolated phages had high progeny production, fast adsorption time and significant stability to temperature, salt and pH changes.The results of phage vB-PaS-HSN4 genome analysis showed that this phage has a double-stranded DNA genome of 44534 bp, which encodes 61 ORFs. Phage genome lacks genes encoding bacterial virulence factors and drug resistance. Examining the antibacterial effects of the phage vB-PaS-HSN4 in an animal model showed that the phage increased the survival of larvae (60%, 66%, 80%) (100MOI), (26%, 40%, 60%) (1MOI (respectively in Pre-treatment, co-treatment and post-treatment tests performed. Based on the characteristics of the bacteriophages, it seems that the isolated phages can be used as a substitute or complement to the common methods of treating burn infections caused by Pseudomonas aeruginosa.

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