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In vitro activity and mode of action of the antimicrobial peptide CPF-P2 against Escherichia coli ATCC 700928
Dissertation (MSc (Biochemistry))--University of Pretoria, 2024.
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| Format: | Thesis |
| Language: | English |
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University of Pretoria
2025
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| _version_ | 1867613663885524992 |
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| access_status_str | Open Access |
| author2 | Gaspar, A.R.M. (Anabella Regina Marques) |
| author_browse | Gaspar, A.R.M. (Anabella Regina Marques) |
| author_facet | Gaspar, A.R.M. (Anabella Regina Marques) |
| collection | Thesis |
| dc_rights_str_mv | © 2023 University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. |
| description | Dissertation (MSc (Biochemistry))--University of Pretoria, 2024. |
| format | Thesis |
| id | oai:repository.up.ac.za:2263/100045 |
| institution | University of Pretoria (South Africa) |
| language | English |
| last_indexed | 2026-06-10T12:39:44.170Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UPSpace — University of Pretoria Institutional Repository |
| publishDate | 2025 |
| publishDateRange | 2025 |
| publishDateSort | 2025 |
| publisher | University of Pretoria |
| publisherStr | University of Pretoria |
| record_format | dspace |
| source_str | UPSpace — University of Pretoria Institutional Repository |
| spelling | oai:repository.up.ac.za:2263/100045 In vitro activity and mode of action of the antimicrobial peptide CPF-P2 against Escherichia coli ATCC 700928 Gaspar, A.R.M. (Anabella Regina Marques) molebogengonkaetse@gmail.com Bester, Megan J. Ibrahim, Mohammed Auwal Onkaetse, Molebogeng Refemetswe UCTD Sustainable Development Goals (SDGs) Antimicrobial resistance Antimicrobial peptides Biofilms Escherichia coli Membrane permeabilisation Dissertation (MSc (Biochemistry))--University of Pretoria, 2024. A major challenge associated with antimicrobial resistance (AMR) is the lack of new drugs especially for the treatment of infections caused by Gram-negative bacteria. Antimicrobial peptides (AMPs) present an alternate solution to the crisis of AMR, as AMPs are novel antimicrobials with unique modes of action and potentially reduced risk of developing antibiotic resistance. Previous studies have identified the antimicrobial potential of the frog skin-derived AMP, the caerulein-precursor fragment-P2 (CPF-P2). To advance the development of this AMP for therapeutic applications, the aim of this study was to further evaluate the antimicrobial activity, mode of action, cytotoxicity and stability against Escherichia coli ATCC 700928. Against E. coli ATCC 700928, the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of CPF-P2 was 16 ± 0.7 μg/mL (6.08 ± 0.27 μM), with an MIC/MBC ratio of 1 indicating that this AMP is bactericidal. Against E. coli biofilms, the minimum biofilm prevention concentration (MBPC) of CPF-P2 was 640 μg/mL (243.2 μM), whereas both the minimum biofilm inhibitory concentration (MBIC) and minimum biofilm eradication concentration (MBEC) were >2560 μg/mL (> 972.8 μM). This implies that although some biofilm prevention occurred, against established biofilms CPF-P2 was ineffective. Only at 128 μg/mL (48.64 ± 2.64 μM) and 256 μg/mL (97.28 ± 2.25 μM) CPF-P2, did human erythrocyte haemolysis occur while some cytotoxicity was observed against HaCaT cells at 256 μg/mL (97.28 ± 4.81 μM) after 24 hours exposure. The mode of action studies included the determination of the killing kinetics, kinetics of membrane permeabilisation and the subsequent effects on bacteria ultrastructure. At the MBC of CPF-P2 (i.e. 16 μg/mL) the killing time was 120 minutes, mediated by membrane permeabilisation which was time- and concentration-dependent. Ultrastructural studies with scanning electron microscopy, confirmed membrane targeting with structural changes to cell shape, ruptured cell membranes and leakage of cellular content with evidence of blebbing. The stability of CPF-P2 in physiological environments was evaluated. CPF-P2 retained activity in 5% foetal bovine serum (FBS) but lost activity in 25% FBS. Pre-incubation with trypsin also resulted in complete loss of CPF-P2 activity. In conclusion, CPF-P2 was identified as bactericidal, selective for planktonic E. coli with a killing time of 120 minutes at its MBC. The bactericidal activity is mediated by membrane permeabilisation leading to associated changes in cell morphology. Its activity against biofilms was limited and was compromised in physiological environments. Nonetheless, this study identified CPF-P2, as a promising template for the further development of analogues for topical infections. South African National Department of Health and the SA-UK Newton Fund Antibiotic Accelerator (MC-PC-MRT/T029552/1) Medical Research Council (MRC) and the South African Medical Research Council (SAMRC). Biochemistry, Genetics and Microbiology (BGM) MSc (Biochemistry) Restricted Faculty of Natural and Agricultural Sciences SDG-03: Good health and well-being SDG-09: Industry, innovation and infrastructure SDG-12: Responsible consumption and production SDG-15: Life on land SDG-17: Partnerships for the goals 2025-01-15T06:36:00Z 2025-01-15T06:36:00Z 2025-04-30 2024-09-16 Dissertation * A2025 http://hdl.handle.net/2263/100045 https://doi.org/10.25403/UPresearchdata.27961263 en © 2023 University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. application/pdf University of Pretoria |
| spellingShingle | UCTD Sustainable Development Goals (SDGs) Antimicrobial resistance Antimicrobial peptides Biofilms Escherichia coli Membrane permeabilisation In vitro activity and mode of action of the antimicrobial peptide CPF-P2 against Escherichia coli ATCC 700928 |
| title | In vitro activity and mode of action of the antimicrobial peptide CPF-P2 against Escherichia coli ATCC 700928 |
| title_full | In vitro activity and mode of action of the antimicrobial peptide CPF-P2 against Escherichia coli ATCC 700928 |
| title_fullStr | In vitro activity and mode of action of the antimicrobial peptide CPF-P2 against Escherichia coli ATCC 700928 |
| title_full_unstemmed | In vitro activity and mode of action of the antimicrobial peptide CPF-P2 against Escherichia coli ATCC 700928 |
| title_short | In vitro activity and mode of action of the antimicrobial peptide CPF-P2 against Escherichia coli ATCC 700928 |
| title_sort | in vitro activity and mode of action of the antimicrobial peptide cpf p2 against escherichia coli atcc 700928 |
| topic | UCTD Sustainable Development Goals (SDGs) Antimicrobial resistance Antimicrobial peptides Biofilms Escherichia coli Membrane permeabilisation |
| url | http://hdl.handle.net/2263/100045 https://doi.org/10.25403/UPresearchdata.27961263 |