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Optimizing bacteriophage engineering through an accelerated evolution platform.

ABSTRACT: The emergence of antibiotic resistance has raised serious concerns within scientific and medical communities, and has underlined the importance of developing new antimicrobial agents to combat such infections. Bacteriophages, naturally occurring bacterial viruses, have long been characterized as promising antibiotic alternatives. Although bacteriophages hold great promise as medical tools, clinical applications have been limited by certain characteristics of phage biology, with structural fragility under the high temperatures and acidic environments of therapeutic applications significantly limiting therapeutic effectiveness. This study presents and evaluates the efficacy of a new accelerated evolution platform, chemically accelerated viral evolution (CAVE), which provides an effective and robust method for the rapid enhancement of desired bacteriophage characteristics. Here, our initial use of this methodology demonstrates its ability to confer significant improvements in phage thermal stability. Analysis of the mutation patterns that arise through CAVE iterations elucidates the manner in which specific genetic modifications bring forth desired changes in functionality, thereby providing a roadmap for bacteriophage engineering.

SUBMITTER: Favor AH

PROVIDER: S-EPMC7438504 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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Optimizing bacteriophage engineering through an accelerated evolution platform.

Favor Andrew H AH Llanos Carlos D CD Youngblut Matthew D MD Bardales Jorge A JA

Scientific reports 20200819 1

The emergence of antibiotic resistance has raised serious concerns within scientific and medical communities, and has underlined the importance of developing new antimicrobial agents to combat such infections. Bacteriophages, naturally occurring bacterial viruses, have long been characterized as promising antibiotic alternatives. Although bacteriophages hold great promise as medical tools, clinical applications have been limited by certain characteristics of phage biology, with structural fragil ...[more]

PMID: 32814789

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Project description:Several mechanisms that increase the rate of mutagenesis across the entire genome have been identified; however, how the rate of evolution might be promoted in individual genes is unclear. Most genes in bacteria are encoded on the leading strand of replication. This presumably avoids the potentially detrimental head-on collisions that occur between the replication and transcription machineries when genes are encoded on the lagging strand. Here we identify the ubiquitous (core) genes in Bacillus subtilis and determine that 17% of them are on the lagging strand. We find a higher rate of point mutations in the core genes on the lagging strand compared with those on the leading strand, with this difference being primarily in the amino-acid-changing (nonsynonymous) mutations. We determine that, overall, the genes under strong negative selection against amino-acid-changing mutations tend to be on the leading strand, co-oriented with replication. In contrast, on the basis of the rate of convergent mutations, genes under positive selection for amino-acid-changing mutations are more commonly found on the lagging strand, indicating faster adaptive evolution in many genes in the head-on orientation. Increased gene length and gene expression amounts are positively correlated with the rate of accumulation of nonsynonymous mutations in the head-on genes, suggesting that the conflict between replication and transcription could be a driving force behind these mutations. Indeed, using reversion assays, we show that the difference in the rate of mutagenesis of genes in the two orientations is transcription dependent. Altogether, our findings indicate that head-on replication-transcription conflicts are more mutagenic than co-directional conflicts and that these encounters can significantly increase adaptive structural variation in the coded proteins. We propose that bacteria, and potentially other organisms, promote faster evolution of specific genes through orientation-dependent encounters between DNA replication and transcription.

Project description:BackgroundCystic fibrosis (CF) is a chronic multi-systemic disease that requires a complex daily treatment regimen. Therefore, there is sub-optimal adherence to CF therapies, and it was shown to impact its clinical and economic burden. Cystic fibrosis transmembrane conductance regulator modulators (CFTRm) are high-cost medications that demonstrated significant benefit in clinical trials. The aim of this study was to evaluate the safety, usability, and efficacy of the ReX platform in medication management of CFTRm for the treatment of people with CF (pwCF).MethodsReX is a patient engagement platform consisting of a cloud-based management system and a cell-enabled handheld device intended to dispense oral medication into the patient's mouth, following a pre-programmed treatment protocol. It provides real-time adherence data to caregivers and timely, personalized reminders to patients. This is a prospective multi-center open study for pwCFs older than 12 years, who had been prescribed CFTRm [elexacaftor/tezacaftor/ivacaftor (ETI) or tezacaftor/ivacaftor (TI)], and provided consent to use ReX platform to receive CFTRm and record their health condition. Study duration was 12-24 months, with clinic visits where physical examination, body mass index (BMI), and pulmonary function tests were performed, and user experience questionnaires were filled in.ResultsTen pwCFs from two CF centers in Israel were included. The mean age was 31.5 years (range 15-74 years); eight were taking ETI and two TI. Median adherence to CFTRm was 97.5% (range 70%-100%) in the first year and 94% (range 84%-99%) in the second year, which is higher than the previously reported CFTRm adherence of ∼80%. No adverse events related to the use of the platform were reported. Patients reported ReX to be valuable to their treatment management and user friendly. Estimated mean forced expiratory volume in 1 s (FEV1%) increased from 74.4% to 80.8% (p = 0.004) over 2 years. Similarly, estimated BMI percentile increased from 53.5 to 59.0 (p < 0.001).ConclusionsUsing the ReX platform in medication management of pwCF treated by CFTRm is safe, easy to use, and effective in improving the adherence to treatment and the clinical outcomes. Consequently, this device may potentially reduce costs to healthcare providers. Further larger and long-term studies are required to examine the clinical benefits of the ReX platform.

Dataset Information

Optimizing bacteriophage engineering through an accelerated evolution platform.

Publications

Optimizing bacteriophage engineering through an accelerated evolution platform.

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