Document Detail


Alternative methods to limit extracellular bacterial activity for enumeration of intracellular bacteria.
MedLine Citation:
PMID:  15927288     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The gentamicin survival assay, a method routinely used to estimate bacterial infection of eukaryotic host cells, depends on the presumed limited penetration of gentamicin across the eukaryotic cell membrane. However, some studies have suggested that gentamicin may in fact enter eukaryotic cells and kill intracellular bacteria. In this study we devised alternative methods to enumerate intracellular Salmonellae using a lytic bacteriophage, SP6, and an amino acid auxotroph, Pro- mutant, which replicates selectively within host cells in the presence of its uptake inhibitor, 3,4-dehydro-L-proline. The conventional gentamicin survival assay was systematically compared with the alternative methods for the enumeration of intracellular Salmonellae. We found that gentamicin decreases the survival of intracellular Salmonellae when added to extracellular media at concentrations above 20 microg/ml. The alternative methods do not suffer from this disadvantage, suggesting that they should be used to replace the gentamicin survival assay. In addition, the proline auxotroph method could be applied to detect bacterial release from host cells.
Authors:
Hyun-Ju Kim; Eun Young Kim; Yeongjin Hong; Joon Haeng Rhee; Hyon E Choy
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Publication Detail:
Type:  Comparative Study; Journal Article; Research Support, Non-U.S. Gov't     Date:  2005-05-31
Journal Detail:
Title:  Journal of microbiological methods     Volume:  64     ISSN:  0167-7012     ISO Abbreviation:  J. Microbiol. Methods     Publication Date:  2006 Jan 
Date Detail:
Created Date:  2005-12-19     Completed Date:  2006-10-19     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  8306883     Medline TA:  J Microbiol Methods     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  17-26     Citation Subset:  IM    
Affiliation:
Genome Research Center for Enteropathogenic Bacteria and Research Institute of Vibrio Infection, South Korea.
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MeSH Terms
Descriptor/Qualifier:
Anti-Bacterial Agents / pharmacology*
Colony Count, Microbial / methods*
Extracellular Space / microbiology*
Gentamicins / pharmacology
Green Fluorescent Proteins / biosynthesis
Hela Cells / microbiology
Humans
Intracellular Space / microbiology*
Microscopy, Confocal
Proline / analogs & derivatives,  pharmacology
Salmonella Phages
Salmonella typhimurium / drug effects,  growth & development*,  virology
Survival Analysis
Chemical
Reg. No./Substance:
0/Anti-Bacterial Agents; 0/Gentamicins; 147-85-3/Proline; 147336-22-9/Green Fluorescent Proteins; 3220-74-4/3,4-dehydroproline

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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