| 1. |
- Euler, Chad W., et al.
(författare)
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PlyKp104, a Novel Phage Lysin for the Treatment of Klebsiella pneumoniae, Pseudomonas aeruginosa, and Other Gram-Negative ESKAPE Pathogens
- 2023
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Ingår i: Antimicrobial Agents and Chemotherapy. - 0066-4804. ; 67:5
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Tidskriftsartikel (refereegranskat)abstract
- Klebsiella pneumoniae and Pseudomonas aeruginosa are two leading causes of burn and wound infections, pneumonia, urinary tract infections, and more severe invasive diseases, which are often multidrug resistant (MDR) or extensively drug resistant. Due to this, it is critical to discover alternative antimicrobials, such as bacteriophage lysins, against these pathogens. Unfortunately, most lysins that target Gram-negative bacteria require additional modifications or outer membrane permeabilizing agents to be bactericidal. We identified four putative lysins through bioinformatic analysis of Pseudomonas and Klebsiella phage genomes in the NCBI database and then expressed and tested their intrinsic lytic activity in vitro. The most active lysin, PlyKp104, exhibited .5-log killing against K. pneumoniae, P. aeruginosa, and other Gram-negative representatives of the multidrug-resistant ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, K. pneumonia, Acinetobacter baumannii, P. aeruginosa, and Enterobacter species) without further modification. PlyKp104 displayed rapid killing and high activity over a wide pH range and in high concentrations of salt and urea. Additionally, pulmonary surfactants and low concentrations of human serum did not inhibit PlyKp104 activity in vitro. PlyKp104 also significantly reduced drug-resistant K. pneumoniae .2 logs in a murine skin infection model after one treatment of the wound, suggesting that this lysin could be used as a topical antimicrobial against K. pneumoniae and other MDR Gram-negative infections.
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| 2. |
- Lood, Rolf, et al.
(författare)
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Determining bacteriophage endopeptidase activity using either fluorophore-quencher labeled peptides combined with liquid chromatography-mass spectrometry (LC-MS) or Förster resonance energy transfer (FRET) assays
- 2017
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 12:3
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Tidskriftsartikel (refereegranskat)abstract
- The necessity of identifying novel methods to combat infections caused by antibiotic resistant bacteria is increasing each year. Recent advancements in the development of peptidoglycan hydrolases (e.g. lysins) from bacterial viruses (bacteriophages) have revealed the efficiency of this class of enzymes in treating serious bacterial infections. Though promising results have been obtained regarding the lethal action of lysin on bacterial pathogens both in vitro and in vivo, an often-overlooked factor in these studies is precisely identifying their peptidoglycan cleavage site. This knowledge would be useful for following the activity of the enzyme during development, without the need for whole-organism lytic assays. However, more importantly, it would enable the selection of lysins with different cleavage activities that would act synergistically for enhanced efficacy. Here, we have developed two new methods to accurately identify the cleavage site of lysins using liquid chromatography mass spectrometry (LC-MS) on peptidoglycan-like fluorophore-quencher modified synthetic peptides, as well as determining the enzymatic action and kinetics of the enzymes on modified peptides in a FoÈrster resonance energy transfer (FRET) assay. These methods should facilitate progress within the lysin field, accelerating the development of therapeutic lysins to combat antibiotic resistant bacterial infections.
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