اسم المؤلف: هند تحسين ابراهيم

اسم المشرف: حارث جبار فهد المذخوري

الموضوع العام: علوم الحياة

السنة: 2014

الموضوع الدقيق: الاحياء المجهرية

الدرجة: ماجستير

الجامعة: جامعة بغداد

اللغة: الانكليزية

مكان الجامعة: بغداد

الصفحات الاولى:

المستخلص: SDS - PAGE showed that a single sharp band with an approximately 16.8 KDa molecular weight has been seen. A matter indicates that the enzyme is consistently pure. PCR technique was applied to approve the existence of nuc gene in S.aureus Nevertheless, only those which depicted positive results on DNase agar harboured nuc gene, as it is specified by single band appearance of nuc at assumed molecular weight (ca. 255 pb) of marker that being used. The current study findings highlighted the participation of SNase purified from S. aureus in significant (P< 0.05) preventing biofilm formation by E. coli and K. pneumoniae compared with untreated controls. Evidently, the inhibitory effect of SNase on biofilm formation is undoubtedly perceived, indicating the degradation of the structural major component of biofilm formation (i.e. extracellular DNA). Results revealed that SNase was able to significantly (P< 0.05) reduce the number of the uropathogens; E. coli and K. pneumoniae attached to the uroepithelial cells. 1 Chapter one : Introduction and literature Review 1. Introduction and Literature Review 1.1. Introduction Staphylococcus aureus is a persistent human pathogen that is responsible for a range of diseases that vary widely in clinical presentation and severity. The pathogenesis of S. aureus infection is a complex process involving a diverse array of secreted and surface - associated virulence determinants that are coordinately expressed at different stages of infection (Loughman et al., 2009). Escherichia coli is a genetically diverse species that causes a variety of infections which fulfill many or all of the proposed criteria for biofilmassociated infections (Kaper et al., 2004). Klebsiella pneumoniae is an opportunistic pathogen responsible for a wide range of nosocomial infections. One important factor associated with virulence in K. pneumoniae is its capacity to adhere to surfaces and form biofilms (Cruz et al., 2012). Bacteria often exist as sessile communities called biofilms which are exquisite structures caused by a genetically programmed developmental process. It is estimated that biofilms are involved in 65% of human bacterial infections, since cells in biofilms are 1000 times more resistant than cells in the planktonic state, making medical treatments fail (Shal? et al., 2011). Extracellular DNA (eDNA) plays a significant role in biofilm formation, as revealed by studies in several bacteria including E. coli (Nakao, 2012) and K. pneumoniae (Whitchurch et al., 2002); however, there is no definite proof on a cause - and - effect relationship between DNA release and biofilm formation (Beenken et al., 2012) or becomes a key component of the macromolecular scaffold in many different biofilms (Jakubovics et al., 2013). In rare cases eDNA has been shown to inhibit bacterial settlement. Therefore, it is possible that nuclease mediated eDNA degradation would therefore promote adhesion. Consequently, it is not clear whether microbial nucleases contribute to the gross biofilm structure in clinically relevant situations (Sheilds et al., 2013). 2 Chapter one : Introduction and literature Review All previous studies used both commercial bovine and recombinant human DNaseI in the disruption of medically important biofilms; whilst, extracellular nuclease of S. aureus (SNase) was used against biofilm of S. aureus themselves (Benenken et al., 2012b ; Kiedrowski et al., 2014). Thus, to date, the role of S. aureus exonucleases in biofilm of other bacteria remains unclear. However, in the present study SNase is used against biofilm of other bacterial species (viz. K. pneumoniae and E. coli). To address this, the following steps were undertaken : 1 - Investigating the negative impact of SNase extracted from S. aureus on K. pneumoniae and E.coli biofilm formation. 2 - Studying the preventive activity of SNase on the adherence of K. pneumoniae and E. coli on uroepithelial cell.