Resequencing microarray for rapid detection & antimicrobial resistance profiling

描述（由申请人提供）： 本申请的长期目标是创建和验证诊断工具（重测序微阵列），其将允许快速检测医院病原体，艰难梭菌、铜绿假单胞菌、不动杆菌属，肠球菌属，克雷伯氏菌属，粘质沙雷氏菌、奇异变形杆菌和嗜麦芽窄食单胞菌的耐药基因多态性。参与这项应用的研究人员团队代表了一个多学科的科学家小组，他们参与了环境和临床样本中细菌的分子检测。LBNL的Andersen，Brodie和DeSantis博士已经创建了许多微阵列，例如检测所有已知细菌和古细菌的16 S rRNA PhyloChip。加州大学旧金山分校的Lynch和Flanagan博士此前曾使用PhyloChip来描述在抗生素给药期间重症患者临床样本中与抗生素耐药病原体增殖相关的细菌群落动态。加州大学旧金山分校的维纳-克罗尼什博士和米勒博士是医学科学家。加州大学旧金山分校的麦克杜格尔博士和杨博士是抗菌药剂师，他们将获得临床样本进行测试和验证。Foxall博士将带来微阵列开发领域的世界领导者Affyoung的工业专业知识，使拟议的工具成为基于最新科学技术的临床设备。加州大学旧金山分校的西格尔博士是一位统计学家和生物信息学专家，在微阵列分析方面具有特殊的专业知识。所提出的方法包括选择生物标志物和抗生素抗性决定簇中的遗传变异区域，用于病原体鉴定和抗生素抗性表型预测;设计和制造Affytechnic平台重测序微阵列，命名为细菌抗生素抗性基因芯片或BARChip，其将询问所选择的区分性基因座处的每个核苷酸位置;优化目标制备和杂交过程，以缩短获得结果的时间;在CLIA认证的实验室中收集临床样本，用于最终重测序阵列验证，然后在拟议项目的最后一年使用这些前瞻性收集的临床样本对BARChip进行盲法验证。这一应用将导致诊断测试的发展，可以快速确定患者是否有细菌感染。该测试将允许医生快速选择适当的抗生素来治疗细菌感染;该工具还将识别感染患者的细菌菌株，因此可以快速识别和控制感染的爆发。

DESCRIPTION (provided by applicant): The long-term objective of this application is to create and validate a diagnostic tool (a resequencing microarray), that will permit rapid detection of nosocomial pathogens, Clostridium difficile, Pseudomonas aeruginosa, Acinetobacterspp., Enterobacterspp., Klebsiella spp., Serratia marcescens, Proteus mirabilis and Stenotrophomonas maltophila as well as their antibiotic resistance gene polymorphisms within 8 hours. The team of investigators involved in this application represents a multidisciplinary group of scientists involved in the molecular detection of bacteria in environmental and clinical samples. Drs. Andersen, Brodie and DeSantis of LBNL have created a number of microarrays e.g. the 16S rRNA PhyloChip that detects all known bacteria and archaea. Drs. Lynch and Flanagan of UCSF have previously used the PhyloChip to describe bacterial community dynamics associated with antimicrobial resistant pathogen proliferation in clinical samples from critically ill patients during antimicrobial administration. Drs. Wiener-Kronish and Miller of UCSF are physician-scientists. Drs. MacDougall and Yang of UCSF are antimicrobial pharmacists, who will obtain clinical samples for testing and validation. Dr. Foxall will bring the industrial expertise of Affymetrix, world leaders in microarray development, to make the proposed tool a clinical device based on the latest state-of-the-science technology. Dr. Segal of UCSF is a statistician and bioinformatics expert with particular expertise in microarray analysis. The proposed process involves selection of regions of genetic variation in biomarkers and antibiotic resistance determinants for pathogen identification and antibiotic resistance phenotype prediction; design and fabrication of an Affymetrix platform resequencing microarray, to be named the Bacterial Antibiotic Resistance GeneChip or BARChip which will interrogate each nucleotide position at the discriminatory loci selected; optimization of target preparation and hybridization processes to decrease time to result; clinical sample collection for final resequencing array validation in a CLIA certified laboratory followed by blinded validation of the BARChip with these prospectively collected clinical samples in the final year of the proposed project. This application would lead to the development of a diagnostic test that could rapidly determine whether a patient has a bacterial infection. The test would permit physicians to rapidly choose appropriate antibiotic(s) to treat the bacterial infection; this tool will also identify the strain of bacteria infecting patients, so outbreaks of infection could be rapidly identified and controlled.