Peptide Nucleic Acid-Fluorescence In Situ Hybridization (PNA-FISH) Assay for Specific Detection of Mycobacterium immunogenum and DNA-FISH Assay for Analysis of Pseudomonads in Metalworking Fluids and Sputum

Details

• Personal Author:
  Kapoor R ; Selvaraju SB ; Yadav JS
• Description:
  Specific and rapid detection and quantification of mycobacteria in contaminated metalworking fluid (MWF) are problematic due to complexity of the matrix and heavy background co-occurring microflora. Furthermore, cross-reactivity among neighboring species of Mycobacterium makes species differentiation difficult for this genus. Here, we report for the first time a species-specific peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH) method for Mycobacterium immunogenum, a non-tuberculous Mycobacterium species prevalent in MWF and implicated in occupational lung disease hypersensitivity pneumonitis and pseudo-outbreaks. A novel species-specific 14-bp PNA probe was designed for M. immunogenum based on its 16S rRNA gene sequence and was validated for specificity, by testing against a panel of other phylogenetically closely related rapidly growing mycobacteria and representative species of gram-positive, gram-negative, and acid fast organisms. In addition, a DNA-FISH protocol was optimized for co-detection of Pseudomonas, the most predominantly co-occurring genus in contaminated MWF. Reliable quantification for both the test organisms was achieved at or above a cell density of 10(3)cellsml(-1), a recognized minimum limit for microscopic quantification. The mycobacterial PNA-FISH assay was successfully adapted to human sputum demonstrating its potential for clinical diagnostic applications in addition to industrial MWF monitoring, to assess MWF-associated exposures and pseudo-outbreaks. [Description provided by NIOSH]
• Subjects:
  Bacterial Infections Communicable Diseases Hypersensitivity Lung Lung Diseases Metals Microbiology Occupational Diseases Occupational Exposure Occupational Health Respiratory System Respiratory Tract Diseases Safety

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• Keywords:
  Author Keywords: Mycobacterium Immunogenum Bacteria Bacterial-disease Bacterial-infections Bacteriology Cell-function Cell-metabolism Cellular-function Cellular-structures Hypersensitivity Pneumonitis Infection-control Infectious-diseases Lung-cells Lung-disorders Lung-irritants Metalworking-fluids Metalworking Fluids Microbial-test-systems Microorganisms Microscopic-analysis Occupational-diseases Occupational-exposure Occupational-health PNA-FISH Pseudomonads Pulmonary-system-disorders Quantitative-analysis Respiratory-system-disorders Ribonucleic-acids Sputum

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• ISSN:
  0890-8508
• Document Type:
  Text
• Funding:
  Grant
• Genre:
  Journal Article
• Place as Subject:
  Ohio ; OSHA Region 5
• CIO:
  National Institute for Occupational Safety and Health (NIOSH)
• Topic:
  Workplace Safety & Health
• Location:
  North America
• Pages in Document:
  273-280
• Volume:
  55
• Issue:
  5
• NIOSHTIC Number:
  nn:20035171
• Citation:
  Mol Cell Probes 2008 Oct-Dec; 22(5-6):273-280
• Contact Point Address:
  Jagjit S. Yadav, Microbial Pathogenesis and Toxicology Laboratory, Department of Environmental Health, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0056
• Email:
  jagjit.yadav@uc.edu
• Federal Fiscal Year:
  2009
• Performing Organization:
  University of Cincinnati
• Peer Reviewed:
  True
• Start Date:
  20010930
• Source Full Name:
  Molecular and Cellular Probes
• End Date:
  20170731
• Collection(s):
  National Institute for Occupational Safety and Health
• Main Document Checksum:
  urn:sha-512:a1af195609eeecd2a8fd5772fa4ee1779a8db65c3a272e524ce5b59dd2f7d218f48cb73c9fb45e07342250af770ded0158f78f452fdd9d3f0f9a2b24ce6bcc05
• Download URL:
  https://stacks.cdc.gov/view/cdc/187109/cdc_187109_DS1.pdf
• File Type:
  [PDF - 607.44 KB ]