Biology subjects

Frick-Cheng, A. E.

Publications and source records attributed to Frick-Cheng, A. E..

4 recordsLinked to original sources

bioRxiv2023

Altered motility in response to iron-limitation is regulated by lpdA in uropathogenic E. coli CFT073

More than half of all women will experience a urinary tract infection (UTI) in their lifetime with most cases caused by uropathogenic Escherichia coli (UPEC). Bacterial motility enhances UPEC pathogenicity, resulting in more severe disease outcomes including kidney infection. Surprisingly, the connection between motility and iron limitation is mostly unexplored, despite the lack of free iron available in the host. Therefore, we sought to explore the potential connection between iron restriction and regulation of motility in UPEC. We cultured E. coli CFT073, a prototypical UPEC strain, in media containing an iron chelator. Under iron limitation, CFT073 had elevated fliC (flagella) promoter activity, driving motility on the leading edge of the colony. Furthermore, this iron-specific response was repressed by the addition of exogenous iron. We confirmed increased flagella expression in CFT073 by measuring fliC transcript, FliC protein, and surface-expressed flagella under iron-limited conditions. To define the regulatory mechanism, we constructed single knockouts of eight master regulators. The iron-regulated response was lost in crp, arcA, and fis mutants. Thus, we focused on the five genes regulated by all three transcription factors. Of the five genes knocked out, the iron-regulated motility response was most strongly dysregulated in an lpdA mutant, which also resulted in significantly lowered fitness in the murine model of ascending UTI. Collectively, we demonstrated that iron-mediated motility in CFT073 is regulated by lpdA, which contributes to the understanding of how uropathogens differentially regulate motility mechanisms in the iron-restricted host. ImportanceUrinary tract infections (UTIs) are ubiquitous and responsible for over five billion dollars in associated health care costs annually. Both iron acquisition and motility are highly studied virulence factors associated with uropathogenic E. coli (UPEC), the main causative agent of uncomplicated UTI. This work is innovative by providing mechanistic insight into the synergistic relationship between these two critical virulence properties. Here, we demonstrate that iron limitation has pleiotropic effects with consequences that extend beyond metabolism, and impact other virulence mechanisms. Indeed, targeting iron acquisition as a therapy may lead to an undesirable enhancement of UPEC pathogenesis through increased motility. It is vital to understand the full breadth of UPEC pathogenesis to adequately respond to this common infection, especially with the increase of antibiotic resistant pathogens.

microbiology↗

bioRxiv2022

Phenotypic Assessment of Clinical Escherichia coli Isolates Predicts Uropathogenic Potential

For women in the United States, urinary tract infections (UTI) are the most frequent diagnosis in emergency departments, comprising 21.3% of total visits. Uropathogenic Escherichia coli (UPEC) causes [~]80% of uncomplicated UTI. To combat this public health issue, it is vital to characterize UPEC strains as well as differentiate them from commensal strains to reduce the overuse of antibiotics. Surprisingly, no genetic signature has been identified which clearly separates UPEC from other E. coli. Therefore, we examined whether phenotypic data could be predictive of uropathogenic potential. We screened 13 clinical strains of UPEC, isolated from cases of uncomplicated UTI in young otherwise healthy women, in a series of microbiological phenotypic assays using UPEC prototype strain CFT073 and non-pathogenic E. coli strain MG1655 K12 as controls. Phenotypes included adherence, iron acquisition, biofilm formation, human serum resistance, motility, and stress resistance. These data were able to predict the severity of bacterial burden in both the urine and bladders using a well-established experimental mouse model of UTI. Multiple linear regression using three different phenotypic assays, growth in minimal medium, siderophore production, and type 1 fimbrial expression, was predictive of bladder colonization (adjusted r2=0.6411). Growth in ex vivo human urine, hemagglutination of red blood cells, and motility modeled urine colonization (adjusted r2=0.4821). These results showcase the utility of phenotypic characterization to predict the severity of infection these strains may cause. We predict that these methods will also be applicable to other complex, genetically redundant, pathogens. ImportanceUrinary tract infections are the second leading infectious disease worldwide, occurring in over half of the female population during their lifetime. Most infections are caused by uropathogenic Escherichia coli (UPEC). These strains can commensally colonize the gut, but upon introduction to the urinary tract, can infect the host and cause disease. Clinically, it would be beneficial to screen patient E. coli strains to understand their pathogenic potential, which may lead to the administration of prophylactic antibiotic treatment for those with increased risk. Others have proposed the use of PCR-based genetic screening methods to detect UPEC and differentiate them from other E. coli pathotypes; however, this method has not yielded a consistent uropathogenic signature. Here, we have used phenotypic characteristics such as growth rate, siderophore production, and expression of fimbriae to successfully predict uropathogenic potential.

microbiology↗

bioRxiv2021

Ferric Citrate Uptake is a Virulence Factor in Uropathogenic Escherichia coli

More than half of women will experience a urinary tract infection (UTI) with uropathogenic Escherichia coli (UPEC) causing [~]80% of uncomplicated cases. Iron acquisition systems are essential for uropathogenesis, and UPEC encode functionally redundant iron acquisition systems, underlining their importance. However, a recent UPEC clinical isolate, HM7 lacks this functional redundancy and instead encodes a sole siderophore, enterobactin. To determine if E. coli HM7 possesses unidentified iron acquisition systems, we performed RNA-sequencing under iron-limiting conditions and demonstrated that the ferric citrate uptake system (fecABCDE and fecIR) was highly upregulated. Importantly, there are high levels of citrate within urine, some of which is bound to iron, and the fec system is highly enriched in UPEC isolates compared to environmental or fecal strains. Therefore, we hypothesized that HM7 and other similar strains use the fec system to acquire iron in the host. Deletion of both enterobactin biosynthesis and ferric citrate uptake ({Delta}entB/{Delta}fecA) abrogates use of ferric citrate as an iron source and fecA provides an advantage in human urine in absence of enterobactin. However, in a UTI mouse model, fecA is a fitness factor independent of enterobactin production, likely due to the action of host Lipocalin-2 chelating ferrienterobactin. These findings indicate that ferric citrate uptake is used as an iron source when siderophore efficacy is limited, such as in the host during UTI. Defining these novel compensatory mechanisms and understanding the nutritional hierarchy of preferred iron sources within the urinary tract are important in the search for new approaches to combat UTI. ImportanceUPEC, the primary causative agent of uncomplicated UTI is responsible for five billion dollars in healthcare costs in the US each year. Rates of antibiotic resistance are on the rise therefore it is vital to understand the mechanisms of UPEC pathogenesis to uncover potential targets for novel therapeutics. Iron acquisition systems used to obtain iron from sequestered host sources are essential for UPEC survival during UTI and have been used as vaccine targets to prevent infection. This study reveals the ferric citrate uptake system is another important iron acquisition system that is highly enriched in UPEC strains. Ferric citrate uptake has not previously been associated with these pathogenic isolates, underlining the importance of the continued study of these strains to fully understand their mechanisms of pathogenesis.

microbiology↗

bioRxiv2020

The Gene Expression Profile of Uropathogenic Escherichia coli in Women with Uncomplicated Urinary Tract Infections Is Recapitulated in the Mouse Model

Uropathogenic Escherichia coli (UPEC) is the primary causative agent of uncomplicated urinary tract infections (UTIs). UPEC fitness and virulence determinants have been evaluated in a variety of laboratory settings that include a well-established mouse model of UTI. However, the extent to which bacterial physiology differs between experimental models and human infections remains largely understudied. To address this important question, we compared the transcriptomes of three different UPEC isolates in human infection and a variety of laboratory conditions including LB culture, filter-sterilized urine culture, and the UTI mouse model. We observed high correlation in gene expression between the mouse model and human infection in all three strains examined (Pearson correlation coefficient of 0.86-0.87). Only 175 of 3,266 (5.4%) genes shared by all three strains had significantly different expression levels, with the majority of them (145 genes) down-regulated in patients. Importantly, gene expression of both canonical virulence factors and metabolic machinery were highly similar between the mouse model and human infection, while the in vitro conditions displayed more substantial differences. Interestingly, comparison of gene expression between the mouse model and human infection hint at differences in bladder oxygenation as well as nutrient composition. In summary, our work strongly validates the continued use of this mouse model for the study of the pathogenesis of human UTI. ImportanceDifferent experimental models have been used to study UPEC pathogenesis including in vitro cultures in different media, tissue culture, as well as mouse models of infection. The latter is especially important since it allows evaluation of mechanisms of pathogenesis and potential therapeutic strategies against UPEC. Bacterial physiology is greatly shaped by environment and it is therefore critical to understand how closely bacterial physiology in any experimental model relates to human infection. In this study, we found a very strong correlation in bacterial gene expression between the mouse model and human UTI using identical strains, suggesting that the mouse model accurately mimics human infection, definitively supporting its continued use in UTI research.

microbiology↗