Molecular Analysis of Mycobacteria in Cutting Fluids

切削液中分枝杆菌的分子分析

• 批准号: 6437155
• 负责人: Jagjit S Yadav
• 金额: $ 22.95万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-30 至 2004-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6437155
• 关键词: Mycobacterium; allergic pneumonitis; environmental exposure; genetic strain; metal working fluid; method development; occupational hazard; polymerase chain reaction

DESCRIPTION (provided by applicant): Microbial contaminants including nontuberculous mycobacteria (NTM) in metalworking fluids (MWFS) have been implicated in occupational respiratory illnesses. Our broader goals are to develop and apply practical DNA-based approaches for real-time detection, quantitation, and identification of mycobacteria in MWF for understanding the prevalent strains and their survivability in commercial MWFS. The specific aims are (i) to develop polymerase chain reaction (PCR)-based protocols for real-time detection and quantitation of NTM in water-based MWF; (ii). to PCR screen field samples of different commercial formulations of water-based MWFs for NTM followed by strain-specific identification of the NTM isolates using molecular typing methods; (iii) to investigate viability of the identified NTM strains in MWF using quantitative PCR. A Mycobacterium genus-specific PCR-based method will be optimized for real-time detection of NTM in water-based MWF. The method will be used as a basis for developing a quantitative PCR protocol for quantitation of NTM in field MWF samples. NTM strains will be isolated from MWF samples in use across the industries and in selected plants linked with hypersensitivity pneumonitis cases. The isolates will be identified at the species level by 16S ribosomal gene amplicon sequencing and at the strain level based on genome fingerprinting using pulsed field gel electrophoresis (PFGE) or rapid amplification of polymorphic DNA-PCR (RAPD-PCR) technique. Effect of strain type and biocide load on viability of the selected strains in MWF will be investigated in simulated as well as field conditions using quantitative PCR. The resulting information will help NORA's objectives by providing practical methods for microbial exposure assessment and database for developing intervention strategies related to MWF exposures.

描述（由申请人提供）：微生物污染物包括 金属加工液（MWF）中的非结核分枝杆菌（NTM）已经 与职业呼吸道疾病有关。我们更广泛的目标是 开发并应用基于DNA的实用方法进行实时检测， MWF中的定量和鉴定，以了解 普遍的菌株及其在商业MWF中的生存能力。具体目标 是（i）开发基于聚合酶链反应（PCR）的方案 水基MWF中NTM的实时检测和定量； （ii）。到PCR 水基MWF的不同商业公式的屏幕场样品 对于NTM，然后使用特异性鉴定NTM分离株 分子打字方法； （iii）研究已识别的NTM的生存能力 使用定量PCR中MWF中的菌株。分枝杆菌特异性PCR 方法将被优化，以实时检测水基MWF中的NTM。这 方法将被用作开发定量PCR协议的基础 NTM在现场MWF样品中的定量。 NTM菌株将从MWF中分离出来 在整个行业中使用的样品以及与之相关的选定植物 高敏性肺炎病例。分离株将在 物种水平按16S核糖体基因扩增子测序和应变水平 基于使用脉冲场凝胶电泳（PFGE）或 多态DNA-PCR（RAPD-PCR）技术的快速扩增。效果 在MWF中所选菌株的生存力上的应变类型和杀菌载荷将 使用定量在模拟和现场条件中进行研究 pcr。由此产生的信息将通过提供来帮助Nora的目标 用于开发微生物暴露评估和数据库的实用方法 与MWF暴露有关的干预策略。