High-Throughput, Massively Parallel Antimicrobial Resistance Surveillance Using Drop-Based Microfluidics

使用基于液滴的微流体进行高通量、大规模并行抗菌药物耐药性监测

• 批准号: 10357953
• 负责人: Connie B Chang
• 金额: $ 10.69万
• 依托单位: MONTANA STATE UNIVERSITY - BOZEMAN
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-22 至 2023-04-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10357953
• 关键词: Agar; Antibiotic Resistance; Antibiotic Therapy; Antibiotic susceptibility; Antibiotics; Antimicrobial Resistance; Antimicrobial susceptibility; Automation; Bacteria; Bacterial Infections; Bar Codes; Biological Assay; Caliber; Cell Proliferation; Cells; Cessation of life; Clinical; Combined Antibiotics; Communicable Diseases; Detection; Devices; Diagnosis; Diagnostic; Drops; Early Diagnosis; Encapsulated; Engineering; Equipment; Fluorescence; Goals; Growth; Hour; Individual; Infection; Laboratories; Libraries; Life; Liquid substance; Methods; Microfluidic Microchips; Microfluidics; Miniaturization; Minimum Inhibitory Concentration measurement; Monitor; Optics; Pharmacotherapy; Phenotype; Predisposition; Pseudomonas aeruginosa; Research; Resistance; Resolution; Sampling; Sepsis; Signal Transduction; Speed; Stains; Technology; Test Result; Testing; Time; Urinary tract infection; Work; bacterial resistance; base; cell growth; combinatorial; cost; design; detection limit; drug standard; effective therapy; high throughput technology; infectious disease treatment; light scattering; microbial; multiplex assay; new technology; novel; operation; optical fiber; pathogen; population based; portability; prevent; rapid detection; rapid test; resistance mechanism; resistant strain; response; screening; single cell analysis

A fundamental challenge in administering effective treatments for infectious diseases caused by bacteria is the rapid identification of antimicrobial resistance and the efficacy of single or combinatorial drug treatments, including the lowest concentrations or combinations of antibiotics required to prevent bacterial growth. We propose to overcome this by engineering an easy-to-use platform based on miniaturization of microbial cultivation using microfluidic drops that can be tagged, tracked, and evaluated in scalable and massively parallel designs. Standard antimicrobial susceptibility testing (AST) platforms typically require 18-72 hours to generate susceptibility results. The primary research objective of this proposal is to apply droplet-based microfluidics to perform rapid AST screening of P. aeruginosa clinical isolates in under four hours. Droplet-based microfluidics is a technology in which picoliter-sized volumes are created and assayed at rates of up to thousands per second. These drops serve as individual microreactors that can contain single cells. The ability to isolate single cells and discrete combinations of antibiotics within picoliter-sized microreactor volumes will allow for rapid and early detection of antibiotic susceptibility with high resolution and fidelity. This method will enable the ability to detect and quantify subpopulations of single cells that are normally below the limit of detection of standard drug assays. To achieve this goal, we have two specific aims: (1) High-throughput minimum inhibitory concentration screening will be performed by multiplexed assaying with a barcoded droplet library, and (2) A microfluidic device with integrated optical fibers will be engineered for sensitive detection of barcode and cell signals, which will enable the platform to be portable and readily adaptable to clinical settings. This novel, scalable technology for high-throughput antimicrobial susceptibility testing in bacteria can greatly decrease the diagnosis time of bacterial infections such as sepsis or urinary-tract infections.

对细菌引起的传染病进行有效治疗的一个基本挑战是快速识别抗菌素耐药性以及单一或组合药物治疗的功效，包括防止细菌生长所需的最低浓度或抗生素组合。我们建议通过设计一个基于微生物培养小型化的易于使用的平台来克服这个问题，该平台使用微流体滴，可以在可扩展和大规模并行设计中进行标记、跟踪和评估。标准抗菌药物敏感性测试 (AST) 平台通常需要 18-72 小时才能生成敏感性结果。该提案的主要研究目标是应用基于液滴的微流体在四小时内对铜绿假单胞菌临床分离株进行快速 AST 筛查。基于液滴的微流体是一种以每秒数千的速率创建和分析皮升大小的体积的技术。这些液滴充当可容纳单个细胞的单独微反应器。在皮升大小的微反应器体积内分离单细胞和抗生素离散组合的能力将允许以高分辨率和保真度快速、早期检测抗生素敏感性。该方法将能够检测和量化通常低于标准药物测定检测限的单细胞亚群。为了实现这一目标，我们有两个具体目标：（1）将通过使用条形码液滴库的多重分析进行高通量最低抑制浓度筛选，（2）将设计具有集成光纤的微流体装置，用于灵敏检测条形码和细胞信号，这将使该平台易于携带并易于适应临床环境。这种用于细菌高通量抗菌药物敏感性测试的新颖、可扩展的技术可以大大缩短脓毒症或尿路感染等细菌感染的诊断时间。