MS diagnostic bacterial identification library

MS诊断细菌鉴定文库

• 批准号: 8722128
• 负责人: Robert K Ernst
• 金额: $ 27.74万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8722128
• 关键词: Amino Acid Sequence; Antibiotics; Bacteria; Bacterial Infections; Bar Codes; Biological; Biological Assay; Bronchoalveolar Lavage Fluid; Cell Wall; Chemicals; Clinical; Data; Data Set; Development; Diagnosis; Diagnostic; Diagnostic Procedure; Failure; Fingerprint; Future; Generations; Growth; Hour; Individual; Infection; Intellectual Property; Laboratories; Libraries; Life; Lipids; MALDI-TOF Mass Spectrometry; Mass Spectrum Analysis; Membrane; Membrane Lipids; Methodology; Methods; Microbe; Morbidity - disease rate; Mycoses; Nucleic Acids; Organism; Outcome; Pathogen detection; Patients; Pattern; Peptide Sequence Determination; Phenotype; Physicians; Preparation; Protein Analysis; Proteins; Protocols documentation; Recovery; Resistance profile; Resistance to infection; Ribosomal Proteins; Sample Size; Sampling; Serum; Solid; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Speed; Structure; Technology; Time; Translating; United States Food and Drug Administration; antimicrobial peptide; base; clinically relevant; cost; culture plates; design; fundamental research; fungus; genome sequencing; improved; interest; microbial; microwave electromagnetic radiation; mortality; novel; novel diagnostics; pathogen; public health relevance; research and development; treatment planning; vaccine development; wound

PROJECT SUMMARY Rapid and accurate pathogen detection and identification is needed to allow physicians to react and respond appropriately to potentially life threatening infections. With an increased need to treat life-threatening infections more rapidly, in hours instead of days, improved assays using state-of-the-art technologies that decrease time, diagnosis sample size, and cost per assay are needed. Current Food and Drug Administration (FDA)-approved methods for pathogen detection in a clinical laboratory include biological culture, nucleic acid amplification, ribosomal protein sequence characterization, or genome sequencing. Collectively, these methods suffer from being time intensive, requiring amplification of clinically obtained material, and are often significantly costly and burdensome for diagnostic laboratory support staff. Recent advances in matrix-assisted laser desorption ionization-time of flight-mass spectrometry (MALDI- TOF-MS) have enabled development of an accurate, and precise method for pathogen identification. The method uses extracts containing high abundance proteins, picked directly from colonies on culture plates as a chemical bar code for individual species to detect differences in the composition of ribosomal proteins with ~90% accuracy. To increase the ability to rapidly diagnose bacterial infections using MALDI-TOF-MS, this proposal will develop, refine, and utilize ultra-small scale lipid purification methodologies for the extraction of high abundance lipids from Gram-positive and -negative bacteria, as well as fungi. Essential, high abundance lipids are found in all membranes of these microbes and are a highly diverse set of molecules. This diversity forms the basis of our hypothesis that "essential bacterial and fungal lipids constitute a chemical barcode that can be used to identify pathogens by mass spectrometry profiling". Our preliminary data show that these lipid structures are unique and can be used as novel chemical barcodes for speciation and/or sub-speciation of bacterial and fungal infections and resistance patterns to a subset of antibiotic and antimicrobial peptides. Following a rapid extraction method, lipids will be analyzed by mass spectrometry and the resulting spectra will be used to generate a mass spectral signature library of lipid "fingerprints" from a wide variety of clinically relevant pathogen backgrounds. When combined, the analysis of the protein and lipid phenotype will provide > 99% accuracy in pathogen identification from a variety of samples, such as solid medium, serum, bronchoalveolar lavage fluid and/or wound effluent. Ultimately, treatment plans tailored to specific infections should translate to improved outcomes, such as faster recovery times, decreased complications, and decreased morbidity and mortality infected patients.

项目摘要 需要快速准确的病原体检测和鉴定，以使医生能够做出反应和反应 适当地应对潜在的威胁生命的感染。随着治疗威胁生命的感染的需求增加 更迅速的是，在几个小时而不是几天内，使用降低时间的最先进技术改进了测定， 诊断样本量和每个测定的成本。当前食品药品监督管理局（FDA） - 批准 临床实验室中病原体检测的方法包括生物学培养，核酸扩增， 核糖体蛋白序列表征或基因组测序。这些方法共同遭受 耗时，需要放大临床获得的材料，并且通常是昂贵的，并且 负担诊断实验室支持人员的繁重。 矩阵辅助激光解吸电离时间的最新进展（MALDI- TOF-MS）已经能够开发出一种精确且精确的病原体鉴定方法。这 方法使用含有高丰度蛋白的提取物，直接从培养板上的菌落中挑选为 单个物种的化学条形码检测核糖体蛋白组成的差异 〜90％精度。为了增加使用MALDI-TOF-MS快速诊断细菌感染的能力，这 提案将开发，完善和利用超小规模的脂质纯化方法来提取 革兰氏阳性和阴性细菌以及真菌的高丰度脂质。必不可少的，高丰富 在这些微生物的所有膜中都发现了脂质，并且是一组高度多样化的分子。这种多样性 构成了我们假设的基础，即“必需细菌和真菌脂质构成化学条形码，该化学条形码 可用于通过质谱分析识别病原体”。我们的初步数据表明这些脂质 结构是独特的，可用作新型化学条形码，用于形成和/或子规范 细菌和真菌感染以及抗生素和抗菌肽子集的抗性模式。 遵循快速提取方法，将通过质谱法分析脂质，所得光谱将 用于从多种临床上产生脂质“指纹”的质谱签名库 相关的病原体背景。合并后，蛋白质和脂质表型的分析将提供> 来自各种样品的病原体鉴定精度，例如固体培养基，血清， 支气管肺泡灌洗液和/或伤口废水。最终，针对特定感染量身定制的治疗计划 应该转化为改善的结果，例如更快的恢复时间，并发症减少，并且 发病率降低和死亡率感染患者。