Abstract
Fluorescent proteins, especially green fluorescent protein (GFP), have been instrumental in understanding urinary tract infection pathogenesis by uropathogenic Escherichia coli (UPEC). We have used a recently developed GFP variant, vsfGFP-9, to create new plasmid- and chromosome-based GFP derivatives of the UPEC strain UTI89. The vsfGFP-9 strains are nearly 10× brighter with no in vitro growth or in vivo virulence defects compared to previously reported GFP-expressing UTI89 strains. The chromosomal vsfGFP-9 strain is equivalent to the wild type UTI89 during in vivo UTI, while both plasmid GFP constructs have an equivalent virulence defect compared to non-plasmid carrying UTI89. These new vsfGFP-9 expressing strains should be useful for further studies of the pathogenesis of UTI89, and similar strategies can be used to create improved fluorescent derivatives of other UPEC strains.
Highlights
Fluorescent proteins (FPs) have been instrumental to our understanding of bacterial pathogenesis [1]
As with other infectious diseases [1,4], fluorescent proteins have been instrumental for many discoveries of the pathogenic mechanisms utilized by uropathogenic E. coli (UPEC), including the development of intracellular bacterial communities (IBCs) [5,6], quiescent intracellular reservoirs (QIRs) [7], and avoidance of neutrophil killing [6] by the cystitis strain UTI89 [8]
The vsfGFP-9 strains had no gross growth defect relative to UTI89 or to their corresponding GFPmut3 strain (SLC-638 compared with UTI89/pANT4; SLC-719 compared with UTI89 att HK022 ::COM-green fluorescent protein (GFP)) (Figure 1a)
Summary
Fluorescent proteins (FPs) have been instrumental to our understanding of bacterial pathogenesis [1]. As with other infectious diseases [1,4], fluorescent proteins have been instrumental for many discoveries of the pathogenic mechanisms utilized by UPEC, including the development of intracellular bacterial communities (IBCs) [5,6], quiescent intracellular reservoirs (QIRs) [7], and avoidance of neutrophil killing [6] by the cystitis strain UTI89 [8] For these studies, two strains are commonly used, both of which express the GFPmut variant of GFP [1]: UTI89 carrying plasmid pANT4 [9] and UTI89 att HK022 ::COM-GFP [10]. These new, brighter strains should be useful in future studies of the pathogenic mechanisms of UTI89, and the strategies employed here can be applied to improve fluorescent derivatives of other UPEC strains
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