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基于液滴的方法用于研究细菌群体对噬菌体暴露的动态响应。

Droplet-based methodology for investigating bacterial population dynamics in response to phage exposure.

作者信息

Nikolic Nela, Anagnostidis Vasileios, Tiwari Anuj, Chait Remy, Gielen Fabrice

机构信息

Living Systems Institute, Faculty of Health and Life Sciences, University of Exeter, Exeter, United Kingdom.

Department of Physics and Astronomy, Faculty of Environment, Science and Economy, University of Exeter, Exeter, United Kingdom.

出版信息

Front Microbiol. 2023 Nov 21;14:1260196. doi: 10.3389/fmicb.2023.1260196. eCollection 2023.


DOI:10.3389/fmicb.2023.1260196
PMID:38075890
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10703435/
Abstract

An alarming rise in antimicrobial resistance worldwide has spurred efforts into the search for alternatives to antibiotic treatments. The use of bacteriophages, bacterial viruses harmless to humans, represents a promising approach with potential to treat bacterial infections (phage therapy). Recent advances in microscopy-based single-cell techniques have allowed researchers to develop new quantitative methodologies for assessing the interactions between bacteria and phages, especially the ability of phages to eradicate bacterial pathogen populations and to modulate growth of both commensal and pathogen populations. Here we combine droplet microfluidics with fluorescence time-lapse microscopy to characterize the growth and lysis dynamics of the bacterium confined in droplets when challenged with phage. We investigated phages that promote lysis of infected cells, specifically, a phage species with DNA genome, T7 () and two phage species with RNA genomes, MS2 () and Qβ (). Our microfluidic trapping device generated and immobilized picoliter-sized droplets, enabling stable imaging of bacterial growth and lysis in a temperature-controlled setup. Temporal information on bacterial population size was recorded for up to 25 h, allowing us to determine growth rates of bacterial populations and helping us uncover the extent and speed of phage infection. In the long-term, the development of novel microfluidic single-cell and population-level approaches will expedite research towards fundamental understanding of the genetic and molecular basis of rapid phage-induced lysis and eco-evolutionary aspects of bacteria-phage dynamics, and ultimately help identify key factors influencing the success of phage therapy.

摘要

全球范围内抗菌素耐药性的惊人上升促使人们努力寻找抗生素治疗的替代方法。使用对人类无害的细菌病毒——噬菌体,是一种有前景的治疗细菌感染的方法(噬菌体疗法)。基于显微镜的单细胞技术的最新进展使研究人员能够开发新的定量方法,以评估细菌与噬菌体之间的相互作用,特别是噬菌体根除细菌病原体群体以及调节共生菌和病原体群体生长的能力。在这里,我们将微滴微流控技术与荧光延时显微镜相结合,以表征当受到噬菌体攻击时,限制在微滴中的细菌的生长和裂解动态。我们研究了促进感染细胞裂解的噬菌体,具体来说,一种具有DNA基因组的噬菌体T7( )和两种具有RNA基因组的噬菌体MS2( )和Qβ( )。我们的微流控捕获装置产生并固定了皮升大小的微滴,从而能够在温度可控的环境中对细菌的生长和裂解进行稳定成像。记录了长达25小时的细菌群体大小的时间信息,这使我们能够确定细菌群体的生长速率,并帮助我们揭示噬菌体感染的程度和速度。从长远来看,新型微流控单细胞和群体水平方法的发展将加快对噬菌体诱导快速裂解的遗传和分子基础以及细菌 - 噬菌体动态的生态进化方面的基础理解的研究,并最终有助于确定影响噬菌体治疗成功的关键因素。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/79fc/10703435/d556e08fda83/fmicb-14-1260196-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/79fc/10703435/bd930bb03d4c/fmicb-14-1260196-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/79fc/10703435/e860a2c322f4/fmicb-14-1260196-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/79fc/10703435/ed1201000829/fmicb-14-1260196-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/79fc/10703435/d556e08fda83/fmicb-14-1260196-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/79fc/10703435/bd930bb03d4c/fmicb-14-1260196-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/79fc/10703435/e860a2c322f4/fmicb-14-1260196-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/79fc/10703435/ed1201000829/fmicb-14-1260196-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/79fc/10703435/d556e08fda83/fmicb-14-1260196-g004.jpg

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Droplet-based methodology for investigating bacterial population dynamics in response to phage exposure.

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引用本文的文献

[1]
Droplet microfluidics for single-cell studies: a frontier in ecological understanding of microbiomes.

FEMS Microbiol Rev. 2025-1-14

[2]
A bacterial toxin-antitoxin system as a native defence element against RNA phages.

Biol Lett. 2025-6

[3]
Gut phages and their interactions with bacterial and mammalian hosts.

J Bacteriol. 2025-2-20

本文引用的文献

[1]
Deep learning with microfluidics for on-chip droplet generation, control, and analysis.

Front Bioeng Biotechnol. 2023-6-7

[2]
Translating phage therapy into the clinic: Recent accomplishments but continuing challenges.

PLoS Biol. 2023-5

[3]
Immunogenicity of bacteriophages.

Trends Microbiol. 2023-10

[4]
Ultrahigh-Throughput Enzyme Engineering and Discovery in Compartments.

Chem Rev. 2023-5-10

[5]
Global trends and hotspots of phage therapy for bacterial infection: A bibliometric visualized analysis from 2001 to 2021.

Front Microbiol. 2023-1-9

[6]
Monitoring phage-induced lysis of gram-negatives in real time using a fluorescent DNA dye.

Sci Rep. 2023-1-16

[7]
Phage therapy: From biological mechanisms to future directions.

Cell. 2023-1-5

[8]
Global mortality associated with 33 bacterial pathogens in 2019: a systematic analysis for the Global Burden of Disease Study 2019.

Lancet. 2022-12-17

[9]
Phenotyping single-cell motility in microfluidic confinement.

Elife. 2022-11-23

[10]
E. coli bacteraemia and antimicrobial resistance following antimicrobial prescribing for urinary tract infection in the community.

BMC Infect Dis. 2022-10-28

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