Mycobacteria in metalworking fluids

金属加工液中的分枝杆菌

• 批准号: 7256941
• 负责人: Jagjit S Yadav
• 金额: $ 26.59万
• 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 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7256941
• 关键词: Mycobacteria; metalworking; fluids

Non-tuberculous mycobacteria (NTM) frequently colonize the modem metalworking fluids (MWFs) and are increasingly being implicated in occupational respiratory illnesses such as hypersensitivity pneumonitis (HP) in machine workers. Their early detection/recovery and characterization will help take pre-emptive measures and develop intervention strategies. Our first funding has led to the development of PCR based protocols for culture-independent detection and quantification of NTM in water-based MWF, coupled with isolation and genotypic identification of several NTM strains from MWF, which belonged to two species M. immunogenum and M. chelonae. Our proposed future goals are to develop next generation of simplified DNA-based approaches that can be adapted in in-house laboratories and to investigate the relative biocide susceptibility and antigenic potential of these MWF isolated NTM strains (genotypes). The specific aims are (1). to develop protocols for efficient NTM recovery and simple DNA-based detection and quantification of NTM from different MWF types; (2). to determine the relative biocide susceptibility of the available and new NTM strains from MWF by optimization and use of a high throughput protocol; (3). to investigate antigenic characteristics of the available and HP-linked NTM strains. Immunomagnetic separation using Mycobacterium-specific polyclonal antibodies will be developed for selective cell recovery for improved NTM detection and cultural isolation. Our current PCR protocol will be adapted for colorimetric detection of NTM. A new protocol based on Fluorescence in-situ hybridization (FISH) will be developed for microscope- assisted culture-independent detection and quantification of the MWF-specific mycobacterial species. A high throughput protocol based on intracellular ATP will be optimized and used for measurement ofbiocide susceptibility. Relative antigenic potential of the MWF-prevalent strains/genotypes will be determined based on cellular immune response in the sensitized blood cells from HP patients or MWF exposed workers. M. immunogenum polyclonal antibodies or HP patient sera will be applied as probes to identify the putative protein antigens in the cell protein lysates and culture filtrates of the identified antigenic strain. The resulting information will help NORA's objectives by providing a set of practical methods for mycobacterial contamination- and exposure- assessment and a strain characteristics database, for facilitating development of intervention strategies related to MWF exposures.

非结核分枝杆菌（NTM）经常定植在现代金属加工液（MWF）中， 越来越多地涉及职业性呼吸道疾病，如过敏性肺炎（HP） 机器工人。及早发现/回收和定性将有助于采取先发制人的措施 制定干预策略。我们的第一笔资金导致了基于PCR的协议的发展， 在水基MWF中NTM的非培养依赖性检测和定量， 对几株MWF中的NTM菌株进行了基因型鉴定，分别属于两个种M. 免疫原菌和M.龟类。我们提出的未来目标是开发下一代简化的 基于DNA的方法，可在内部实验室中进行调整，并研究相关的杀生物剂 这些MWF分离的NTM菌株（基因型）的易感性和抗原潜力。具体目标是 （一）.开发有效的NTM回收和简单的基于DNA的检测和定量的方案， 不同MWF类型的NTM;确定现有和新的生物杀灭剂的相对敏感性， 通过优化和使用高通量方案从MWF获得NTM菌株;（3）。研究抗原性 可用的和HP连锁的NTM菌株的特征。免疫磁性分离 将开发分枝杆菌特异性多克隆抗体，用于选择性细胞回收，以改善 NTM检测和培养分离。我们目前的PCR方案将适用于比色检测 NTM。一种基于荧光原位杂交（FISH）的新方案将被开发用于显微镜- 辅助的不依赖于培养物的检测和MWF特异性分枝杆菌物种的定量。高 基于细胞内ATP的通量方案将被优化并用于测量杀生物剂 易感性将根据以下标准确定MWF流行菌株/基因型的相对抗原潜力 对HP患者和MWF作业工人致敏血细胞免疫功能的影响。M. 免疫原多克隆抗体或HP患者血清将用作探针以鉴定推定的 鉴定的抗原性菌株的细胞蛋白裂解物和培养物中的蛋白抗原。所得 这些信息将有助于诺拉的目标，提供一套实用的方法， 污染和接触评估以及菌株特征数据库，以促进 与MWF暴露相关的干预策略。