The Streptomycin-Treated Mouse Intestine Selects
            <i>Escherichia coli envZ</i>
            Missense Mutants That Interact with Dense and Diverse Intestinal Microbiota

Leatham-Jensen, Mary; Frimodt-Møller, Jakob; Adediran, Jimmy; Mokszycki, Matthew E.; Banner, Megan; Caughron, Joyce E.; Krogfelt, Karen A.; Conway, Tyrrell; Cohen, Paul S.

doi:10.1128/iai.06193-11

Cited by 50 publications

(106 citation statements)

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“…Interestingly, studies of MG1655 colonization in both germfree and streptomycin-treated mouse models have found that adaptation of this strain to the mouse intestine selects for envZ missense mutations that produce high levels of OmpR phosphorylation (83)(84)(85). We have not looked for the emergence of such mutants in our mouse colonization experiments.…”

Section: Discussionmentioning

confidence: 99%

Escherichia coli Isolate for Studying Colonization of the Mouse Intestine and Its Application to Two-Component Signaling Knockouts

et al. 2014

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The biology of Escherichia coli in its primary niche, the animal intestinal tract, is remarkably unexplored. Studies with the streptomycin-treated mouse model have produced important insights into the metabolic requirements for Escherichia coli to colonize mice. However, we still know relatively little about the physiology of this bacterium growing in the complex environment of an intestine that is permissive for the growth of competing flora. We have developed a system for studying colonization using an E. coli strain, MP1, isolated from a mouse. MP1 is genetically tractable and does not require continuous antibiotic treatment for stable colonization. As an application of this system, we separately knocked out each two-component system response regulator in MP1 and performed competitions against the wild-type strain. We found that only three response regulators, ArcA, CpxR, and RcsB, produce strong colonization defects, suggesting that in addition to anaerobiosis, adaptation to cell envelope stress is a critical requirement for E. coli colonization of the mouse intestine. We also show that the response regulator OmpR, which had previously been hypothesized to be important for adaptation between in vivo and ex vivo environments, is not required for MP1 colonization due to the presence of a third major porin. Escherichia coli is one of the most extensively studied and bestcharacterized organisms. Its high growth rate, facile genetics, and simple nutritional requirements have made this bacterium an excellent model system for studying basic aspects of molecular biology and bacteriology and the primary host for DNA and protein engineering. The physiology of E. coli growth and survival under diverse conditions has been intensively studied, and a significant fraction of E. coli gene products and regulatory networks have been characterized. However, for such a well-studied organism, we know remarkably little about the biology of E. coli in its primary niche: the animal gastrointestinal tract.E. coli is generally the most abundant aerobe in the intestines of warm-blooded vertebrates, although its numbers vary considerably with animal host and geography (1-3). As a species, this bacterium has a remarkable genetic diversity; the number of genes in common among fully sequenced isolates is less than half the number of genes in any individual strain (4-6). Some E. coli strains are pathogenic, depending on the host and site of infection (3, 7-9), and have been intensively studied to understand the factors controlling their virulence. However, the majority of E. coli strains associated with animals are believed to be part of the normal flora of the intestine, growing asymptomatically as commensals.Most of our knowledge about E. coli colonization of the animal intestine comes from studies with streptomycin-resistant strains colonizing mice fed streptomycin continuously in their drinking water (10, 11). This streptomycin-treated mouse model has played a key role in the characterization of the growth of E. coli in the intestine a...

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Section: Discussionmentioning

confidence: 99%

Escherichia coli Isolate for Studying Colonization of the Mouse Intestine and Its Application to Two-Component Signaling Knockouts

et al. 2014

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“…Mouse colonization experiments. The specifics of the streptomycintreated mouse model, used to compare the large-intestine-colonizing abilities of E. coli strains in mice, have been described previously (12). Briefly, sets of three male CD-1 mice (5 to 8 weeks old) were given drinking water containing streptomycin sulfate (5 g/liter) for 24 h to eliminate resident facultative bacteria (31).…”

Section: Methodsmentioning

confidence: 99%

“…More recently, it has been shown that E. coli Nissle 1917 inhibits the adherence of E. coli EDL933 to human epithelial cells in vitro and simultaneously inhibits its growth and Shiga toxin 2 (Stx2) production (23). It has also been shown that in the intestines of streptomycin-treated mice, E. coli Nissle 1917 is better than a number of other commensal E. coli strains at limiting the growth of E. coli EDL933 (12,13). In this study, we examine whether E. coli Nissle 1917 uses both glycolytic and gluconeogenic nutrients to colonize the intestines of streptomycin-treated mice and whether E. coli EDL933 switches to gluconeogenic nutrients in order to maintain itself in the intestines in the presence of E. coli Nissle 1917.…”

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confidence: 99%

“…The streptomycin-treated mouse model is used routinely for this purpose, since streptomycin-treated mice fulfill these criteria (10,11). Using this model, we have shown that for E. coli to colonize, it must obtain nutrients in the mucus layer, which overlies the epithelium (10,11), that it resides in the mucus layer as a member of mixed biofilms (12,13), and that each E. coli strain displays a unique nutritional program in the intestine (14).…”

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Escherichia coli EDL933 Requires Gluconeogenic Nutrients To Successfully Colonize the Intestines of Streptomycin-Treated Mice Precolonized with E. coli Nissle 1917

et al. 2015

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bEscherichia coli MG1655, a K-12 strain, uses glycolytic nutrients exclusively to colonize the intestines of streptomycin-treated mice when it is the only E. coli strain present or when it is confronted with E. coli EDL933, an O157:H7 strain. In contrast, E. coli EDL933 uses glycolytic nutrients exclusively when it is the only E. coli strain in the intestine but switches in part to gluconeogenic nutrients when it colonizes mice precolonized with E. -12) reported that E. coli 86-24, an O157:H7 strain, activates the expression of virulence genes under gluconeogenic conditions, suggesting that colonization of the intestine with a probiotic E. coli strain that outcompetes O157:H7 strains for gluconeogenic nutrients could render them nonpathogenic. Here we report that E. coli Nissle 1917, a probiotic strain, uses both glycolytic and gluconeogenic nutrients to colonize the mouse intestine between 1 and 5 days postfeeding, appears to stop using gluconeogenic nutrients thereafter in a large, long-term colonization niche, but continues to use them in a smaller niche to compete with invading E. coli EDL933. Evidence is also presented suggesting that invading E. coli EDL933 uses both glycolytic and gluconeogenic nutrients and needs the ability to perform gluconeogenesis in order to colonize mice precolonized with E. coli Nissle 1917. The data presented here therefore rule out the possibility that E. coli Nissle 1917 can starve the O157:H7 E. coli strain EDL933 of gluconeogenic nutrients, even though E. coli Nissle 1917 uses such nutrients to compete with E. coli EDL933 in the mouse intestine. While much attention has been paid to the role of specific virulence factors in bacterial pathogenesis, until recently little attention has been paid to the roles of specific metabolic pathways in the ability of bacterial pathogens to initiate the pathogenic process. Yet clearly, if a bacterial pathogen is unable to initiate pathogenesis because nutrients essential for its growth are unavailable in the host, it will be unable to establish itself and cause disease. Therefore, it is important to determine whether pathogens require glycolytic nutrients, gluconeogenic nutrients, or both for success in invading and subsequently colonizing the host. Indeed, recent studies have suggested that some pathogens use glycolytic nutrients, some use gluconeogenic nutrients, and some use both to adapt to diverse host habitats (1-7).We are interested in the nutritional basis of Escherichia coli colonization of the intestine and particularly in whether precolonization by commensal E. coli strains can be used to prevent invading E. coli intestinal pathogens from colonizing. Commensal E. coli strains colonize the human intestine in the presence of a dense and diverse intestinal microbiota comprising at least 500 cultivable species and 10 13 to 10 14 total bacteria (8). Unfortunately, E. coli colonization cannot be studied experimentally in conventional animals due to colonization resistance, which occurs when all niches are filled by the microbiota (9)....

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“…For example, envZ mutations were created by hydroxylamine (Tokishita and Mizuno, 1994) or nitrosoguanadine mutagenesis (Russo et al, 1993) or by UV irradiation (Hsing et al, 1998). Another study isolated missense mutations in envZ that were better able than wildtype E. coli MG1655 to colonize the streptomycin-treated mouse (Adediran et al, 2014;Leatham-Jensen et al, 2012). These envZ missense mutations were located in TM1 and TM2, and were more resistant to bile salts and colicin V compared to E. coli MG1655.…”

Section: Tm Domains Modulate Envz Activity But Are Not Essentialmentioning

confidence: 99%

Cytoplasmic sensing by the inner membrane histidine kinase EnvZ

Foo

Gao

Zhang

et al. 2015

Progress in Biophysics and Molecular Biology

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Two-component regulatory systems drive signal transduction in bacteria. The simplest of these employs a membrane sensor kinase and a cytoplasmic response regulator. Environmental sensing is typically coupled to gene regulation. The histidine kinase EnvZ and its cognate response regulator OmpR regulate expression of outer membrane proteins (porins) in response to osmotic stress. We used hydrogen:deuterium exchange mass spectrometry to identify conformational changes in the cytoplasmic domain of EnvZ (EnvZc) that were associated with osmosensing. The osmosensor localized to a seventeen amino acid region of the four-helix bundle of the cytoplasmic domain and flanked the His 243 autophosphorylation site. High osmolality increased autophosphorylation of His 243 , suggesting that these two events were linked. The transmembrane domains were not required for osmosensing, but mutants in the transmembrane domains altered EnvZ activity. A photoactivatable fusion protein composed of EnvZc fused to the fluorophore mEos2 (EnvZc-mEos2) was as capable as EnvZc in supporting OmpR-dependent ompF and ompC transcription. Over-expression of EnvZc reduced activity, indicating that the EnvZ/OmpR system is not robust. Our results support a model in which osmolytes stabilize helix one in the four-helix bundle of EnvZ by increased hydrogen bonding of the peptide backbone, increasing autophosphorylation and downstream signaling. The likelihood that additional histidine kinases use similar cytoplasmic sensing mechanisms is discussed.

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The Streptomycin-Treated Mouse Intestine Selects Escherichia coli envZ Missense Mutants That Interact with Dense and Diverse Intestinal Microbiota

Abstract: ABSTRACTPreviously, we reported that the streptomycin-treated mouse intestine selected nonmotileEscherichia coliMG1655flhDCdeletion mutants ofE. coliMG1655 with improved colonizing ability … Show more

Cited by 50 publications

References 63 publications

Escherichia coli Isolate for Studying Colonization of the Mouse Intestine and Its Application to Two-Component Signaling Knockouts

Escherichia coli Isolate for Studying Colonization of the Mouse Intestine and Its Application to Two-Component Signaling Knockouts

Escherichia coli EDL933 Requires Gluconeogenic Nutrients To Successfully Colonize the Intestines of Streptomycin-Treated Mice Precolonized with E. coli Nissle 1917

Cytoplasmic sensing by the inner membrane histidine kinase EnvZ

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