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العلاقة بين انتاج الانزيم المحلل للكولاجين وتكوين الغشاء الحياتي بوساطة بكتريا Pseudomonas aeruginosae == The Relationship Between Collagenase Production And Biofilm Formation By Pseudomonas Aeuroginosa
Author name:
امال عزيز كريم السعدي
Supervisor name:
شذى سلمان الطحان
General topic:
Biology
Specific topic:
Microbiology - Bacteria
Degree:
Doctorate
University:
University of Baghdad
Language:
English
University location:
Baghdad
First pages:
24T2915 - p.pdf
Abstract:
A total of 359 samples divided as 228 clinical and 131 non clinical specimens were collected during 2012 from four hospitals in Baghdad city including : Al - Kadhymia Teaching hospital, Baghdad Teaching hospital, The Burn Specialist Hospital and Al - Imam Ali hospital, for isolation of P.aeruginosa to study the correlation between collagenase production and biofilm formation. Eighty two Pseudomonas isolates were screened for biofilm formation, 28 isolates were classified as strong biofilm formers, 25 as moderate and 27 as weak biofilm former. The 28 isolates were identifid by VITEk - 2 Compact system which confirmed that the isolates were P.aeruginosa. Collagenase production assay was used to screen 28 isolates that were strong biofilm formers inorder to detect the ability of these isolates to produce collagenase, the substrate of collagenase (collagen) was purified localy from bovin tendon and the results showed that just 8 isolates could grow in mineral salt media with collagen after 4 days of incubation. The factors affecting biofilm formation and collagenase production were studied to determine the optimual conditions for their production, those factors included : 1 - Nitrogen sources represented higher influence on collagenase production specialy (yeast extract) in media containing collagen than other media without collagen as a substrate. The specific activity differed between the 8 isolates, biofilm formation also became more pronounced with (yeast extract), while NH4Cl and NaNO3 depressed biofilm formation at the same conditions. The statistical analysis between the two parameters (biofilm and collagenase) according to different nitrogen sources demonstrated highly significance at p?0.01 with yeast extract and casein. 2 - pH, results showed that the best pH for production was 7 for both collagenase and biofilm.The statistical analysis for determination the relationshipe between the two parameters showed highly significance at p ?0.01 for different pH. 3 - The maximum production of the two parameters was at 35?C temperature which gave highly significance at p?0.01 with defferent temperature. 4 - Long incubation periods revealed increasing in collagenase production and biofilm formation which represented highly significance detween them when incubation periods were prolonged at p?0.01. Results of this study showed that collagenase production increases when bacteria switch from a planktonic to biofilm phenotype. This indicates that biofilms and collagenase are more virulent and have a greater ability to cause tissue destruction. The REP - PCR analysis using BOX - primer, showed a clusters genetic relatedness among the isolates. The isolates were grouped according to the REP - PCR in 9 different genotypes, named cluster 1 to 3 which included C1, C2, C3 with relatedness : 8 (80%), 8 (86%), 3 (80%) respectively. A19 and A20 both of them were not included in any cluster, they have 78% similarity.The REP - PCR analysis showed that the genotypic relatedness is consistently high between the 8 producer isolates and non producer isolates (13), showed similarity reached 86
