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.

Agency
National Institute of Health (NIH)
Institute
National Institute for Occupational Safety and Health (NIOSH)
Type
Research Project (R01)
Project #
5R01OH007364-06
Application #
7256941
Study Section
Safety and Occupational Health Study Section (SOH)
Program Officer
Board, Susan
Project Start
2001-09-30
Project End
2010-06-30
Budget Start
2007-07-01
Budget End
2010-06-30
Support Year
6
Fiscal Year
2007
Total Cost
$265,939
Indirect Cost
Name
University of Cincinnati
Department
Public Health & Prev Medicine
Type
Schools of Medicine
DUNS #
041064767
City
Cincinnati
State
OH
Country
United States
Zip Code
45221
Johansson, Elisabet; Boivin, Gregory P; Yadav, Jagjit S (2017) Early immunopathological events in acute model of mycobacterial hypersensitivity pneumonitis in mice. J Immunotoxicol 14:77-88
Chandra, Harish; Yadav, Jagjit S (2016) Human leukocyte antigen (HLA)-binding epitopes dataset for the newly identified T-cell antigens of Mycobacterium immunogenum. Data Brief 8:1069-71
Chandra, Harish; Yadav, Jagjit S (2016) T-cell antigens of Mycobacterium immunogenum, an etiological agent of occupational hypersensitivity pneumonitis. Mol Immunol 75:168-77
Chandra, Harish; Lockey, James; Yadav, Jagjit S (2015) Novel antigens of Mycobacterium immunogenum relevant in serodiagnosis of occupational hypersensitivity pneumonitis in machinists. Ann Allergy Asthma Immunol 114:525-6
Kapoor, Renuka; Selvaraju, Suresh B; Yadav, Jagjit S (2014) Extended tracking of the microbial community structure and dynamics in an industrial synthetic metalworking fluid system. FEMS Microbiol Ecol 87:664-77
Chandra, Harish; Yadav, Ekta; Yadav, Jagjit S (2013) Alveolar macrophage innate response to Mycobacterium immunogenum, the etiological agent of hypersensitivity pneumonitis: role of JNK and p38 MAPK pathways. PLoS One 8:e83172
Selvaraju, Suresh B; Khan, Izhar U H; Yadav, Jagjit S (2011) Susceptibility of Mycobacterium immunogenum and Pseudomonas fluorescens to formaldehyde and non-formaldehyde biocides in semi-synthetic metalworking fluids. Int J Mol Sci 12:725-41
Kapoor, Renuka; Yadav, Jagjit S (2010) Development of a rapid ATP bioluminescence assay for biocidal susceptibility testing of rapidly growing mycobacteria. J Clin Microbiol 48:3725-8
Gupta, Manish K; Subramanian, Venkataraman; Yadav, Jaglit S (2009) Immunoproteomic identification of secretory and subcellular protein antigens and functional evaluation of the secretome fraction of Mycobacterium immunogenum, a newly recognized species of the Mycobacterium chelonae-Mycobacterium abscessus group. J Proteome Res 8:2319-30
Selvaraju, Suresh B; Kapoor, Renuka; Yadav, Jagjit S (2008) 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. Mol Cell Probes 22:273-80

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