Bacteriophage DNA Organic Extraction

Kunisch, Fabian; Wagemans, Jeroen; Moreno, Mercedes Gonzalez

doi:10.21203/rs.3.pex-1955/v1

Cited by 3 publications

(3 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Genomic DNA of ES10 phage was extracted using the phenolchloroform method (Kunisch et al, 2023) from 10 mL of ES10 phage filtrate (2.2 × 10 11 PFU/mL) treated with DNase I and RNase. Quantification and purification of the extracted DNA was carried out using Nanodrop and the integrity of the extracted DNA was confirmed by gel electrophoresis.…”

Section: Characterization Of Es Dry Labmentioning

confidence: 99%

Characterization of ES10 lytic bacteriophage isolated from hospital waste against multidrug-resistant uropathogenic E. coli

Nawaz,

Zafar,

Alessa

et al. 2024

Front. Microbiol.

View full text Add to dashboard Cite

Escherichia coli is the major causative agent of urinary tract infections worldwide and the emergence of multi-drug resistant determinants among clinical isolates necessitates the development of novel therapeutic agents. Lytic bacteriophages efficiently kill specific bacteria and seems promising approach in controlling infections caused by multi-drug resistant pathogens. This study aimed the isolation and detailed characterization of lytic bacteriophage designated as ES10 capable of lysing multidrug-resistant uropathogenic E. coli. ES10 had icosahedral head and non-contractile tail and genome size was 48,315 base pairs long encoding 74 proteins. Antibiotics resistance, virulence and lysogenic cycle associated genes were not found in ES10 phage genome. Morphological and whole genome analysis of ES10 phage showed that ES10 is the member of Drexlerviridae. Latent time of ES10 was 30 min, burst size was 90, and optimal multiplicity of infection was 1. ES10 was stable in human blood and subsequently caused 99.34% reduction of host bacteria. Calcium chloride shortened the adsorption time and latency period of ES10 and significantly inhibited biofilm formation of host bacteria. ES10 caused 99.84% reduction of host bacteria from contaminated fomites. ES10 phage possesses potential to be utilized in standard phage therapy.

show abstract

Section: Characterization Of Es Dry Labmentioning

confidence: 99%

Characterization of ES10 lytic bacteriophage isolated from hospital waste against multidrug-resistant uropathogenic E. coli

Nawaz,

Zafar,

Alessa

et al. 2024

Front. Microbiol.

View full text Add to dashboard Cite

show abstract

“…Based on plaque morphology and host strain we identified 31 phages that were purified and produced as described above. Of those, 10 (MK.R3-15, MK.R3-30, MK.R57-15, MK.R57-30, MK.R84-15, MK.R84-30, FIM.R60-15, FIM.R60-30, FJK.R9-determined by BLASTn v2.13.0 74 , were concentrated and purified by PEG 8000 precipitation 73 . The name is composed of the ancestral phage name (MK, FIM and FJK), the isolation strain (e.g., R84 for Paer84) and the number of passages denoted as 15 (isolation after round 15) or 30 (isolation after round 30).…”

Section: Bacteriophage Isolationmentioning

confidence: 99%

“…To analyse the phage treated Paer09 strains' genomic data, single nucleotide polymorphisms (SNP), small deletions and insertions (indels) between the ancestral Paer09 genome and each isolated colony were identified using Snippy v4.6.0 84 Phage genomes were extracted 85 and the concentration and purity were determined by NanoDrop ND-1000 UV-Vis Spectrophotometer (PEQLAB, Erlangen, Germany). Phage DNA was subsequently sequenced with Illumina as described in Makalatia et al 86 .…”

Section: Whole Genome Sequencing and Analysismentioning

confidence: 99%

Targeting MDR Pseudomonas aeruginosa biofilm with an evolutionary trained bacteriophage cocktail exploiting phage resistance trade-offs

Kunisch

Wagemans

Yildirim

et al. 2023

Preprint

Self Cite

View full text Add to dashboard Cite

Spread of multidrug-resistant Pseudomonas aeruginosa strains threatens to render currently available antibiotics obsolete, with limited prospects for the development of new antibiotics. Lytic bacteriophages (phages), the natural predators of bacteria, represent a path to combat this threat. In vitro directed evolution is traditionally applied to expand the phage host range or increase bacterial suppression in planktonic cultures. However, while 80% of human microbial infections are biofilm-associated, research towards targeted improvement of phages’ ability to combat biofilms remains scarce. Here, we describe an in vitro evolution assay that improves multiple phage parameters in parallel, while optimizing phage cocktail design by exploiting a bacterial phage resistance trade-off. The obtained evolved phages show an expanded host spectrum, improved antimicrobial efficacy under isothermal microcalorimetric monitoring and enhanced antibiofilm performance, as determined by RT-qPCR. Our two-phage cocktail revealed further improved antimicrobial efficacy without incurring dual-phage-resistance in treated bacteria. We anticipate this assay will allow a better understanding of phenotypic-genomic relationships in phages and enable the training of phages against other desired pathogens. This, in turn, will strengthen phage therapy as a treatment adjunct to improve clinical outcomes of multidrug-resistant bacterial infections.

show abstract

Targeting Pseudomonas aeruginosa biofilm with an evolutionary trained bacteriophage cocktail exploiting phage resistance trade-offs

Kunisch,

Campobasso,

Wagemans

et al. 2024

Nat Commun

View full text Add to dashboard Cite

Spread of multidrug-resistant Pseudomonas aeruginosa strains threatens to render currently available antibiotics obsolete, with limited prospects for the development of new antibiotics. Lytic bacteriophages, the viruses of bacteria, represent a path to combat this threat. In vitro-directed evolution is traditionally applied to expand the bacteriophage host range or increase bacterial suppression in planktonic cultures. However, while up to 80% of human microbial infections are biofilm-associated, research towards targeted improvement of bacteriophages’ ability to combat biofilms remains scarce. This study aims at an in vitro biofilm evolution assay to improve multiple bacteriophage parameters in parallel and the optimisation of bacteriophage cocktail design by exploiting a bacterial bacteriophage resistance trade-off. The evolved bacteriophages show an expanded host spectrum, improved antimicrobial efficacy and enhanced antibiofilm performance, as assessed by isothermal microcalorimetry and quantitative polymerase chain reaction, respectively. Our two-phage cocktail reveals further improved antimicrobial efficacy without incurring dual-bacteriophage-resistance in treated bacteria. We anticipate this assay will allow a better understanding of phenotypic-genomic relationships in bacteriophages and enable the training of bacteriophages against other desired pathogens. This, in turn, will strengthen bacteriophage therapy as a treatment adjunct to improve clinical outcomes of multidrug-resistant bacterial infections.

show abstract

Bacteriophage DNA Organic Extraction

Cited by 3 publications

References 0 publications

Characterization of ES10 lytic bacteriophage isolated from hospital waste against multidrug-resistant uropathogenic E. coli

Characterization of ES10 lytic bacteriophage isolated from hospital waste against multidrug-resistant uropathogenic E. coli

Targeting MDR Pseudomonas aeruginosa biofilm with an evolutionary trained bacteriophage cocktail exploiting phage resistance trade-offs

Targeting Pseudomonas aeruginosa biofilm with an evolutionary trained bacteriophage cocktail exploiting phage resistance trade-offs

Contact Info

Product

Resources

About