دراسة تجريبية لانزيمات البروتييز للزائفة الزنجارية المعزولة من المرضى العراقيين المصابين بتقرح القرنية ودورها في علاج تقرح القرنية المتسبب عن المكورات العنقودية == Experimental Study of Pseudomonas aeruginosa Proteases Isolated from Corneal Ulcer of Iraqi Patients and Their Role in the Treatment of Staphylococcus aureus Keratitis
Author name:
عائدة حسين ابراهيم
Supervisor name:
جاسم محمد كرحوت | منيرة جلوب اسماعيل
General topic:
Medicine
Specific topic:
Microbiology
Degree:
Doctorate
University:
University of Baghdad - Faculty Of Medicine
Language:
English
University location:
Baghdad
First pages:
19T1535 - p.pdf
Abstract:
One - hundred and twenty samples ( corneal scraping) were collected from patients diagnosed to have microbial keratitis (corneal ulcer) who attended Ibn Al - Haitham Teaching Eye Hospital from the period between May 2013 and November 2013, Pseudomonas aeruginosa was reported 26 (21.6%) from the total cases. All bacterial isolates were diagnosed by conventional and biochemical tests, and confirmed by Vitek 2 Compact System.The role of proteases enzymes ( Elastase ( LasB), LasA, Alkaline protease and Protease IV ) of Pseudomonas aeruginosa in the corneal ulcer was studied by using genetic and molecular biological method by real time PCR, and the results indicated that three bacterial isolates of Pseudomonas aeruginosa possessed elastase gene (LasB) (11.5%), and only one bacterial isolate of Pseudomonas aeruginosa harbored LasA protease gene ( 3.8%). All bacterial isolates of Pseudomonas aeruginosa were harbored alkaline protease gene (100%), and twenty bacterial isolates were harbored protease IV (76.9%).The results of real - time PCR analysis indicated that four bacterial isolates of Pseudomonas aeruginosa (15.3%) were harbored more than one gene of different proteases enzymes ( elastase, alkaline protease, and protease IV).On the other hand our results showed that one bacterial isolates (3.8%) harbored both LasA protease and alkaline protease genes, and twenty bacterial isolates of Pseudomonas aerugenosa (76.9%) were harbored alkaline protease and protease IV genes.The LasA protease was extracted from Pseudomonas aeruginosa isolate by cooling centrifuge and precipitated supernatant by ammonium sulfate at saturation (80%). The resulted extracted crude enzyme concentration was 60 μg/ml. Then the crude enzyme was partially purified by dialysis and gel filtration chromatography by using Sephadex G - 100. The concentration of partial purified enzyme reached 40μg/ml.IIISummaryThe results of the experimental treatment of bacterial keratitis ( in vivo) of infected eyes rabbits caused by Methicillin Sensitive Staphylococcus aureus showed that the efficacy of LasA protease was effective was as Lysostaphin in eradicating Methicillin Sensitive Staphylococcus aureus from the infected corneas after approximately 15 h after giving the drug at dose 100 μl ( concentration 1μg / ml ) . While Vancomycin gave us very little potency in eradicating S. aureusfrom corneas in comparison with potency of LasA protease and Lysostaphin duringthis time but showed good potency very late approximately after 3 days of applicationof treatment.The results of the experimental treatment in vitro (in the test tube ) that is caused by Methicillin Sensitive Staphylococcus aureus showed that the efficacy of LasA protease was similar to that of Lysostaphin drug in the killing of Methicillin Sensitive Staphylococcus aureus in the bacterial broth.
