Di Somma, Angela (2020) Regulation of biofilm development in Gram-negative and Gram-positive bacteria: New antimicrobial strategies. [Tesi di dottorato]


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Item Type: Tesi di dottorato
Resource language: English
Title: Regulation of biofilm development in Gram-negative and Gram-positive bacteria: New antimicrobial strategies
Di Somma, Angelaangela.disomma@unina.it
Date: 13 March 2020
Number of Pages: 185
Institution: Università degli Studi di Napoli Federico II
Department: Scienze Chimiche
Dottorato: Scienze chimiche
Ciclo di dottorato: 32
Coordinatore del Corso di dottorato:
Lombardi, Angelinaalombard@unina.it
Date: 13 March 2020
Number of Pages: 185
Keywords: biofilm, antimicrobial peptide, molecular mechanisms, functional and differential proteomics.
Settori scientifico-disciplinari del MIUR: Area 05 - Scienze biologiche > BIO/10 - Biochimica
Date Deposited: 27 Mar 2020 12:13
Last Modified: 05 Nov 2021 13:27
URI: http://www.fedoa.unina.it/id/eprint/13191

Collection description

During evolution, microorganisms developed different forms and organizations according to needs. They can be found in planktonic form or as sessile aggregates anchored to biotic or abiotic surfaces. This phenotype is known as biofilm, an ensemble of microbial cells irreversibly associated to a surface and enclosed in an essentially self-produced matrix (1). This articulated and complex matrix organization can be called “house of biofilm cells” (2), because biofilm protects bacteria cells allowing them to survive in hostile environments. This condition significantly contributes to the onset of diseases, encouraging the adhesion to host cells and preventing access to antimicrobial agents. Currently, any drug targeting crucial processes for bacterial life will inevitably lead to the development of resistant strains. On the other hand, the development of potential molecules designed to prevent biofilm formation and the identification of target proteins involved in biofilm formation will leave pathogens exposed to antibiotic treatments. In the search for new drugs, increasing attention has been devoted to AntiMicrobialPeptides (AMPs), small molecules composed of 10-100 amino acid residues produced by all organisms. AMPs are attractive candidates for the design of new antibiotics because of their specific characteristics and a low propensity for the development of resistance. My PhD project focused on the investigation of biofilm formation under stress conditions and the definition of the mechanism of action of antimicrobial peptides., Two enzymes deeply involved in biofilm formation in E.coli and M. smeg, the N-acetylneuraminate lyase NanA and the Bifunctional protein GlmU, respectively, were identified. When these enzymes are impaired by either inhibitors or genetic deletion, biofilm formation decreases confirming their key role in the process. Since they are present in prokaryotic and not in eukaryotics they represent promising targets for the development of new drugs. The possible development of AMPs as new antibiotics is strictly dependent on the definition of their lethal effect at the molecular level. Our attention has been focused on the elucidation of the mechanism of action of Temporin-L by functional proteomics approaches and the evaluation of its antibacterial and antibiofilm properties. The results showed that Temporin-L (TL) interacts with several proteins belonging to the divisome and elongasome complexes, impairing the natural formation of the septal peptidoglycan and affecting the cell division process. We demonstrated that Temporin-L is able to affect bacterial cell division by inhibiting FtsZ a protein involved in the crucial step of Z-ring formation at the beginning of the division process. Moreover, we studied the mechanism of action of Magainin-2, another peptide deriving from frog skin. Preliminary results showed that a large number of the overexpressed proteins in the presence of the peptide are involved in porin activity and protein insertion into the outer membrane. The antimicrobial activities of Temporin-L and Magainin-2 on the EUCAST panel for susceptibility testing have also been evaluated. Magainin-2 did not exhibit a significant effect on the bacterial strains, while TL showed a strong activity on S. aureus, E. coli and S. pneumonia showing different effects as revealed by TEM microscopy. A differential proteomic experiment has been performed on S. aureus, which suggests that TL treatment might stimulate the synthesis of proteins involved in membrane extroflexion and this phenomenon seems to reduce bacterial pathogenesis as shown by the down regulation of proteins involved in pathogenesis and cell adhesion.


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