Dataset Information

A New High-Throughput Screening Method for Phages: Enabling Crude Isolation and Fast Identification of Diverse Phages with Therapeutic Potential.

ABSTRACT: Bacteriophage therapy and application of phages for biocontrol necessitate acquisition of suitable phages. The exclusivity of phage-host relations and the risk of phage resistance instigate a need to rapidly isolate and characterize novel phages and continually build sizeable phage libraries. Current methods for phage isolation are both laborious and time consuming, suitable for the isolation of a limited number of phages. The high-throughput screening method for phages upscales and organizes enrichment of phages for fast isolation and identification of potentially hundreds of distinct phages against single hosts. This enables screening of hundreds of samples, in multiple simultaneous setups with varying parameters, increasing the likelihood of isolating multiple distinct phages specific for the given conditions. The efficiency of the method is emphasized by our screening of 200 environmental samples, resulting in the identification of an abundance of unique phage species virulent to Escherichia coli, Salmonella enterica, Enterococcus faecalis, and Pseudomonas aeruginosa.

SUBMITTER: Olsen NS

PROVIDER: S-EPMC9041460 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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A New High-Throughput Screening Method for Phages: Enabling Crude Isolation and Fast Identification of Diverse Phages with Therapeutic Potential.

Olsen Nikoline S NS Hendriksen Niels Bohse NB Hansen Lars H LH Kot Witold W

PHAGE (New Rochelle, N.Y.) 20200916 3

Bacteriophage therapy and application of phages for biocontrol necessitate acquisition of suitable phages. The exclusivity of phage-host relations and the risk of phage resistance instigate a need to rapidly isolate and characterize novel phages and continually build sizeable phage libraries. Current methods for phage isolation are both laborious and time consuming, suitable for the isolation of a limited number of phages. The <i>high-throughput screening</i> method for phages upscales and organ ...[more]

PMID: 36147828

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Project description:Understanding the mechanisms behind chemical susceptibility differences is key to protecting sensitive populations. However, elucidating gene-environment interactions (GxE) presents a daunting challenge. While mammalian models have proven useful, problems with scalability to an enormous chemical exposome and clinical translation faced by all models remain; therefore, alternatives are needed. Zebrafish (Danio rerio) have emerged as an excellent model for investigating GxE. This study used a combined bioinformatic and experimental approach to probe the mechanisms underlying chemical susceptibility differences in a genetically diverse zebrafish population. Starting from high-throughput screening (HTS) data, a genome-wide association study (GWAS) using embryonic fish exposed to 0.6 μM Abamectin revealed significantly different effects between individuals. A hypervariable region with two distinct alleles-one with G at the SNP locus (GG) and one with a T and the 16 bp deletion (TT)-associated with differential susceptibility was found. Sensitive fish had significantly lower sox7 expression. Due to their location and the observed expression differences, we hypothesized that these sequences differentially regulate sox7. A luciferase reporter gene assay was used to test if these sequences, alone, could lead to expression differences. The TT allele showed significantly lower expression than the GG allele in MCF-7 cells. To better understand the mechanism behind these expression differences, predicted transcription factor binding differences between individuals were compared in silico, and several putative binding differences were identified. EMSA was used to test for binding differences in whole embryo protein lysate to investigate these TF binding predictions. We confirmed that the GG sequence is bound to protein in zebrafish. Through a competition EMSA using an untagged oligo titration, we confirmed that the GG oligo had a higher binding affinity than the TT oligo, explaining the observed expression differences. This study identified differential susceptibility to chemical exposure in a genetically diverse population, then identified a plausible mechanism behind those differences from a genetic to molecular level. Thus, an HTS-compatible zebrafish model is valuable and adaptable in identifying GxE mechanisms behind susceptibility differences to chemical exposure.

Dataset Information

A New High-Throughput Screening Method for Phages: Enabling Crude Isolation and Fast Identification of Diverse Phages with Therapeutic Potential.

Publications

A New High-Throughput Screening Method for Phages: Enabling Crude Isolation and Fast Identification of Diverse Phages with Therapeutic Potential.

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