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Examination of Salmonella gene expression in an infected mammalian host using the green fluorescent protein and two‐colour flow cytometry
Author(s) -
Bumann Dirk
Publication year - 2002
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.2002.02821.x
Subject(s) - biology , green fluorescent protein , flow cytometry , promoter , virulence , orfs , reporter gene , gene , salmonella , microbiology and biotechnology , genetics , gene expression , bacteria , open reading frame , peptide sequence
Summary Quantitative data on Salmonella gene expression in infected hosts are largely lacking because of technical problems. One attractive reporter, the green fluorescent protein (GFP), is widely used in vitro but is difficult to quantify in infected tissues because of the preponderance of background particles with similar fluorescence. Here, bacterial GFP emission was spectrally distinguished from host autofluorescence by two‐colour flow cytometry. Using this technique, the in vivo activity of three well‐characterized promoters (P sicA , P ssaH and P pagC ) was determined. Their spatial and temporal activity patterns are in close agreement with predictions based on previous data and the colonization defects of corresponding deletion strains. To identify additional Salmonella promoters that are induced in infected animals, a genomic library was sorted by flow cytometry yielding four independent promoters. Genes expressed from P pibB and P sifA contribute to virulence, and chorismate mutase expressed from P aroQ might participate in aromatic acid biosynthesis, which is also required for virulence. Promoter P 3g appears to be part of a mobile genetic element that is lacking in the completely sequenced strain LT2.