Rapid Antibiotic Susceptibility Testing for Neonatal Intensive Units

新生儿重症监护病房的快速抗生素敏感性测试

• 批准号: 7903816
• 负责人: Vincent Jen-Jr Gau
• 金额: $ 21.85万
• 依托单位: GENEFLUIDICS, INC.
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7903816
• 关键词: Acute; Address; Agreement; Ampicillin; Antibiotic Resistance; Antibiotic susceptibility; Antibiotics; Antimicrobial susceptibility; Arizona; Automation; Award; Bacteremia; Bacterial Infections; Biological Assay; Biosensor; Biotechnology; Blood; Blood Cells; Blood Tests; Blood specimen; Body Fluids; California; Cefotaxime; Centrifugation; Childhood; Clinical; Clinical Microbiology; Clinical Research; Collaborations; Communicable Diseases; Computer Systems Development; Development; Devices; Diagnosis; Diagnostic; Doctor of Medicine; Doctor of Philosophy; Engineering; Enrollment; Ensure; Escherichia coli; Escherichia coli Infections; Excision; Filtration; Gases; Gentamicins; Goals; Growth; Hospitals; Hour; Imipenem; Infection; Laboratories; Lead; Libraries; Los Angeles; Maps; Measures; Microfluidics; Molecular; Molecular Analysis; Molecular Diagnosis; Multi-Drug Resistance; Neonatal; Neonatal Intensive Care Units; Organism; Outcome; Outsourcing; Patients; Phase; Preparation; Prevalence; Principal Investigator; Process; Protocols documentation; Reporting; Reproducibility; Research; Research Proposals; Resistance profile; Ribosomal RNA; Saliva; Sample Size; Sampling; Screening procedure; Small Business Innovation Research Grant; Specimen; System; Technical Expertise; Techniques; Technology; Testing; Time; Universities; Urine; Validation; Whole Blood; Work; antimicrobial; base; blood filter; clinical practice; clinically relevant; design; electric impedance; evidence base; improved; innovation; instrument; multidisciplinary; novel; pathogen; phase 1 study; point of care; point-of-care diagnostics; product development; programs; public health relevance; sample collection; validation studies

DESCRIPTION (provided by applicant): Standard culture-based diagnosis of bacteremia, including pathogen identification (ID) and antimicrobial susceptibility testing (AST), requires 2-3 days for clinical sample acquisition to result reporting. The absence of definitive microbiological diagnosis at the point of care has largely driven the over- and misuse of antibiotics in the neonatal intensive care unit (NICU), resulting in an increase in proportion and prevalence of antibiotic-resistance organisms. While microbiological diagnosis has improved with the availability of high throughput, automated instruments in the larger clinical microbiology laboratories, the process remains time-consuming and requires significant technical expertise. Standard automation instruments are bulky and typically require a priori isolation of the pathogens from the body fluid samples prior to AST. The significant work burden of a modern clinical microbiology laboratory has led to an increase in outsourcing practice of clinical laboratory tests. Development of a point-of-care (POC) platform capable of rapid pathogen identification and AST can provide clinicians with evidence-based information to start patient-specific antimicrobial treatment only when necessary. Even short-term alterations in the use of antibiotics have been found to favorably impact the antibiotic resistance profiles. Furthermore, such platform could potentially expedite the screening of novel class of antibiotics. In this proposal, we will leverage our ongoing development on the point-of-care diagnostic platform for urine and saliva testing (U01 AI082457 and U01 DE017790) to create an integrated diagnostic cartridge specifically for rapid blood testing by incorporating a complementary rapid blood cell removal cross-flow filter and an electrokinetic (EK) concentrator. The proposed study will utilize cross-flow filtration to replace centrifugation, employ high aspect-ratio gas-permeable microchannels to obtain optimal conditions for rapid phenotypic assessment of bacterial growth, exploit EK sample preparation techniques for on-chip matrix management, and develop an electrochemical-based fluidic cartridge to obtain pathogen identification and antimicrobial susceptibility assessment from infected blood samples in 90 minutes. The ultimate goal of this project is to leverage the advancement of the established microfluidic cartridge technology and the phenotypic assay to develop a POC platform for diagnosing bacteremia in the NICU. While the goal for Phase 1 is to develop a microfluidic cartridge for diagnosing E. coli infection, this platform will be extended to diagnose infections caused by other prevalent pathogens found in the NICU in Phase 2 upon completion of Phase 1. Specific Aims 1 and 2 of this project is to investigate and develop cross-flow filtration and EK manipulation for matrix management, and rapid antibiotic susceptibility testing in fluidic channels. The outcome of the proposed Aim 1 will remove 95% of blood cells with a PDMS-based cross-flow filter with a two-tier micro-channel design. The focus of Specific Aim 2 is to measure the impedance of the cross-flow filtered blood and apply the optimal EK manipulation conditions to each blood specimen based on the impedance analysis. The flow channel geometry, materials and fabrication details will be comparable with the fluidic cartridge to be built in Specific Aim 3. The design inputs from the Specific Aim 1 and 2 will be incorporated into the antibiotic susceptibility testing (RAST) cartridge. The goal of Specific Aim 3 is to develop and validate the RAST fluidic cartridge with standard and fresh blood samples spiked with known E. coli concentrations. The passing criteria of Specific Aim 3 is to achieve 100% agreement when comparing blood culture results with results acquired by the microfluidic cartridge and associated control system in 90 minutes under optimal RAST assay conditions obtained in Specific Aim 1 and 2. We will validate the RAST cartridge with 10 spiked whole blood samples to demonstrate the ability to obtain antibiotic susceptibility in 90 minutes with known antibiotic-resistant E. coli provided by Childrens Hospital Los Angeles in Phase I study. In Phase II, we plan to incorporate the pathogen identification and RAST into an integrated fluidic cartridge for a multi-center validation study with an expanded panel of other common pathogens. The clinical study in Phase II will be led by Dr. Grace Aldrovani at Childrens Hospitals Los Angeles. The sample size and enrollment plan will be finalized toward the end of the Phase I study. PUBLIC HEALTH RELEVANCE: Standard culture-based diagnosis of bacterial infections, including pathogen identification and antimicrobial susceptibility testing require 2-3 days for clinical sample acquisition to result reporting. The absence of definitive microbiological diagnosis at the point of care has led to over- and misuse of antibiotics in neonatal intensive care units. We proposed to develop an integrated diagnostic cartridge specifically for rapid bacteremia diagnosis by utilizing high aspect-ratio gas-permeable microchannels to obtain optimal conditions for rapid phenotypic assessment of bacterial growth, employing cross-flow filtration and electrokinetic manipulation techniques for on-chip matrix management, and developing an electrochemical- based fluidic cartridge to achieve pathogen identification and obtain antimicrobial susceptibility assessment from infected blood samples in 90 minutes.

描述（由申请人提供）：基于标准培养的菌血症的诊断，包括病原体鉴定（ID）和抗菌易感性测试（AST），需要2-3天的临床样本获取才能报告结果。在护理点上没有明确的微生物诊断，这在很大程度上驱动了新生儿重症监护病房（NICU）的过度和滥用抗生素，从而导致抗生素抗性生物体的比例和患病率增加。虽然微生物诊断有所改善，随着较大的临床微生物实验室中高通量，自动化仪器的可用性，该过程仍然耗时，需要大量的技术专业知识。标准自动化仪器很大，通常需要先验从体液样品中先验地隔离病原体。现代临床微生物学实验室的重大工作负担导致临床实验室测试的外包实践增加。开发能够快速病原体鉴定和AST的护理点（POC）平台可以为临床医生提供基于证据的信息，以便仅在必要时就开始患者特异性抗菌治疗。甚至发现使用抗生素的短期改变也会对抗生素耐药性概况产生有益的影响。此外，这种平台可能会加快对新型抗生素类别的筛查。在这项建议中，我们将利用我们在尿和唾液测试的护理诊断平台上正在进行的开发（U01 AI082457和U01 DE017790）来创建一个集成的诊断墨盒，专门用于快速血液测试，通过结合互补的迅速血液再生过滤过滤过滤过滤器和电子浓缩剂（Ek）浓度（Ek）浓度（Ek）。拟议的研究将利用跨流过滤替换离心，采用高方面比率可渗透的可渗透微通道来获得最佳条件，以快速对细菌生长的快速表型评估，利用EK样品制备技术来获得芯片基质的管理，并获得基于电化学的流动性鉴定，并开发出静电性及其静态的固定型，并进行了触及的静态化病态鉴定。在90分钟内。该项目的最终目标是利用已建立的微流体墨盒技术的进步和表型测定法，以开发一个POC平台，用于诊断NICU中的菌血症。 While the goal for Phase 1 is to develop a microfluidic cartridge for diagnosing E. coli infection, this platform will be extended to diagnose infections caused by other prevalent pathogens found in the NICU in Phase 2 upon completion of Phase 1. Specific Aims 1 and 2 of this project is to investigate and develop cross-flow filtration and EK manipulation for matrix management, and rapid antibiotic susceptibility testing in fluidic channels.拟议的AIM 1的结果将用基于PDMS的跨流滤波器以两层微通道设计去除95％的血细胞。特定目标2的重点是测量跨流过滤血液的阻抗，并根据阻抗分析将最佳的EK操纵条件应用于每个血样。流道通道的几何形状，材料和制造细节将与要在特定目标3中构建的流体墨盒相媲美。特定目标1和2的设计输入将纳入抗生素敏感性测试（RAST）墨盒中。特定目标3的目的是开发和验证带有已知大肠杆菌浓度的标准和新鲜血液样品的rast液墨盒。特定目标3的通过标准3是在将血液培养结果与由微流体弹药型和相关的控制系统中获得的结果进行比较，在90分钟内在90分钟内在特定目标1和2中获得的最佳RAST分析条件下，我们将通过10个刺激的全血液样本验证RAST弹药筒，以证明具有抗抗生素的能力，可在90分钟内提供抗生素的能力。洛杉矶一阶段研究。在第二阶段，我们计划将病原体鉴定和RAST纳入整合的液体墨盒中，以进行多中心验证研究，并扩展了其他常见病原体的面板。第二阶段的临床研究将由洛杉矶儿童医院的Grace Aldrovani博士领导。样本量和注册计划将在I阶段研究结束时最终确定。 公共卫生相关性：基于标准的细菌感染的标准诊断，包括病原体鉴定和抗菌易感性测试，需要2-3天的临床样本获取才能进行报告。在护理点上没有明确的微生物诊断，导致新生儿重症监护病房中过度和滥用抗生素。我们提出，通过利用高方面比例可渗透的可渗透微渠道来开发专门用于快速细菌诊断的综合诊断弹药筒，以获取最佳的细菌生长的快速表型评估，采用交叉流量过滤，并采用跨流量过滤，并通过基于电动机的管理和开发电动机的电视和电动机，并实现了电动型和电动机的电脑，并实现了电动机的流动性。在90分钟内对感染血液样本的抗菌敏感性评估。