Proteomic analysis of uropathogenic Escherichia coli

Urinary tract infections (UTIs) are among the most common of bacterial infections in humans. Although a number of Gram-negative bacteria can cause UTIs, most cases are due to infection by uropathogenic E. coli (UPEC). Genomic studies have shown that UPEC encode a number of specialized activities that allow the bacteria to initiate and maintain infections in the environment of the urinary tract. Proteomic analyses have complemented the genomic data and have documented differential patterns of protein synthesis for bacteria growing ex vivo in human urine or recovered directly from the urinary tracts of infected mice. These studies provide valuable insights into the molecular basis of UPEC pathogenesis and have aided the identification of putative vaccine targets. Despite the substantial progress that has been achieved, many future challenges remain in the application of proteomics to provide a comprehensive view of bacterial pathogenesis in both acute and chronic UTIs.     

Role of collectin-11 in innate defence against uropathogenic Escherichia coli infection

Classical collectins (surfactant protein A and D) play a significant role in innate immunity and host defence in uropathogenic Escherichia coli (UPEC)-induced urinary tract infection (UTI). However, the functions of collectin-11 (CL-11) with respect to UPEC and UTI remain largely unexplored. This study aimed to investigate the effect of CL-11 on UPEC and its role in UTI. We further examined its modulatory effect on inflammatory reactions in proximal tubular epithelial cells (PTECs). The present study provides evidence for the effect of CL-11 on the growth, agglutination, binding, epithelial adhesion and invasion of UPEC. We found increased basal levels of phosphorylated p38 MAPK and human cytokine homologue (keratinocyte-derived chemokine) expression in CL-11 knockdown PTECs. Furthermore, signal regulatory protein α blockade reversed the increased basal levels of inflammation associated with CL-11 knockdown in PTECs. Additionally, CL-11 knockdown effectively inhibited UPEC-induced p38 MAPK phosphorylation and cytokine production in PTECs. These were further inhibited by CD91 blockade. We conclude that CL-11 functions as a mediator of innate immunity via direct antibacterial roles as well as dual modulatory roles in UPEC-induced inflammatory responses during UTI. Thus, the study findings suggest a possible function for CL-11 in defence against UTI.     

Vaginal Lactobacillus isolates inhibit uropathogenic Escherichia coli

The purpose of this study was to investigate the antibacterial activities of Lactobacillus jensenii KS119.1 and KS121.1, and Lactobacillus gasserii KS120.1 and KS124.3 strains isolated from the vaginal microflora of healthy women, against uropathogenic, diffusely adhering Afa/Dr Escherichia coli (Afa/Dr DAEC) strains IH11128 and 7372 involved in recurrent cystitis. We observed that some of the Lactobacillus isolates inhibited the growth and decreased the viability of E. coli IH11128 and 7372. In addition, we observed that adhering Lactobacillus strains inhibited adhesion of E. coli IH11128 onto HeLa cells, and inhibited internalization of E. coli IH11128 within HeLa cells.     

Prevalence of virulence genes in Escherichia coli strains isolated from Romanian adult urinary tract infection cases

A total of 78 E. coli strains isolated from adults with different types of urinary tract infections were screened by polymerase chain reaction for prevalence of genetic regions coding for virulence factors. The targeted genetic determinants were those coding for type 1 fimbriae ( fimH ), pili associated with pyelonephritis ( pap ), S and F1C fimbriae ( sfa and foc ), afimbrial adhesins ( afa ), hemolysin ( hly ), cytotoxic necrotizing factor ( cnf ), aerobactin ( aer ). Among the studied strains, the prevalence of genes coding for fimbrial adhesive systems was 86 %, 36%, and 23% for fimH, pap , and sfa/foc , respectively. The operons coding for Afa afimbrial adhesins were identified in 14% of strains. The hly and cnf genes coding for toxins were amplified in 23% and 13% of strains, respectively. A prevalence of 54% was found for the aer gene. The various combinations of detected genes were designated as virulence patterns. The strains isolated from the hospitalized patients displayed a greater number of virulence genes and a diversity of gene associations compared to the strains isolated from the ambulatory subjects. A rapid assessment of the bacterial pathogenicity characteristics may contribute to a better medical approach of the patients with urinary tract infections.     

Escherichia coli from retail meats carry genes associated with uropathogenic Escherichia coli, but are weakly invasive in human bladder cell culture

Aims: The aim of this study was to determine the uropathogenic potential of Escherichia coli isolated from retail meats. Methods and Results: Two hundred E. coli isolates recovered from retail meats, which were previously identified molecularly as extraintestinal pathogenic E. coli, were investigated for the presence of 21 uropathogenic E. coli (UPEC) virulence‐associated genes. Twenty‐three E. coli isolates were selected based on their serogroups and the number of virulence genes they contained, and further characterized using multilocus sequence typing, and by tissue culture assays for adherence to and invasion of T‐24 human bladder cells and for their induction of interleukin (IL)‐6 secretion. All virulence genes tested, except afa/dra and hlyD, were detected among the E. coli isolates. Multilocus sequence typing analysis of 23 selected isolates revealed that 17 isolates belonged to STs associated with human UPEC. Nearly all 23 isolates exhibited lower level of adherence and invasion compared to a clinical strain, UPEC CFT073. Conclusions: These observations suggested that a small proportion of E. coli isolates from retail meats carry uropathogenic associated virulence genes and thus may serve as a reservoir of these genes to UPEC in the human intestine. Their virulence potential seemed limited as they were only weakly invasive in human bladder cell culture. Significance and Impact of the Study: These findings support the hypothesis that retail meat E. coli may play a role in relation to urinary tract infection (UTI) and may be considered in development of a UTI prevention strategy.     

致肾盂肾炎大肠杆菌的毒力因子和调控

致肾盂肾炎大肠杆菌引起人的尿路感染,它的毒力因子包括表面毒力因子和分泌毒力因子两大类。表面毒力因子包括菌毛、鞭毛、黏附素和多糖类物质,主要在细菌的侵染过程中起作用。分泌毒力因子主要是溶血素、细胞毒性坏死因子等毒素蛋白,主要对宿主细胞产生毒力作用。本文简要综述致肾盂肾炎大肠杆菌毒力因子分泌所需要的5种分泌机制,并论及毒力因子的宏观调控和影响毒力调控的因素。     

致病性大肠杆菌UPEC CFT073全基因组分析及致病机制的新认识

尿道致病性大肠杆菌UPEC CFT073菌株(uropathogenic Escherichia coli CFT073)于2002年被完全测序并注释。但是,对其基因组的研究还很不完善,首先表现在基因组注释的系统性错误和滞后性。作者运用一系列生物信息学方法和工具,从编码蛋白质基因、编码RNA基因等角度对RefSeq数据库的基因组注释进行了系统的修正和增补,并在此基础上鉴别了一批新的候选致病因子基因。进一步的分析表明,得到的基因组注释对CFT073致病相关的一些重要调控关系和机制能够给出更准确、完整的描述。     

Depletion of multidrug‐resistant uropathogenic Escherichia coli BC1 by ebselen and silver ion

Ebselen, an organo‐selenium compound with well‐characterized toxicology and pharmacology, recently exhibited potent antibacterial activity against glutathione (GSH)‐negative bacteria by disrupting redox homeostasis. In this paper, we show that ebselen and silver ion in combination exert strong bactericidal activity against multidrug‐resistant (MDR) uropathogenic Escherichia coli (UPEC) BC1, a model MDR GSH‐positive bacterium. The mechanisms were found to involve consumption of total intracellular GSH and inhibition of thioredoxin reductase activity, which was highly related to reactive oxygen species up‐regulation. Furthermore, the therapeutic efficacy of ebselen and silver ion against UPEC‐induced cystitis was assessed in a mouse model. Treatment with ebselen and silver ion significantly reduced bacterial loads, down‐regulated the expression levels of tumour necrosis factor‐α (TNF‐α) and interferon‐γ (IFN‐γ) on‐site and decreased white/red blood cell counts in mild cystitis model mice, which demonstrated the anti‐inflammatory property of these agents. In addition, ebselen and silver ion also exhibited significantly high protective ability (100%) against acute cystitis infections. These results together may lay the foundation for further analysis and development of ebselen and silver ion as antibacterial agents for treatment of MDR UPEC infections.     

Susceptibility of Escherichia coli to L-selenaproline and other L-proline analogues in laboratory culture media and normal human urine

AIMS: The aims of this study were to identify analogues of L-proline which inhibit the growth of Escherichia coli in both laboratory culture media and normal human urine and to study their mechanisms of uptake. METHODS AND RESULTS: The susceptibility of E. coli to L-proline analogues was studied by radial streak assays on agar plates and by minimal inhibitory concentration determinations in liquid media. Only L-selenaproline (SCA) inhibited growth in Mueller-Hinton medium and human urine as well as in glucose minimal medium. L-Proline did not prevent the inhibition of growth by SCA and strains defective in L-proline transport were as susceptible to SCA as wild-type strains. However, E. coli was resistant to SCA in the presence of L-cysteine and L-cystine. Spontaneous mutants selected for resistance to SCA or L-selenocystine were resistant to the other compound and had reduced growth in minimal medium containing L-cysteine or L-cystine as the sole sulfur source. CONCLUSIONS: L-selenaproline inhibited the growth of E. coli under conditions that may occur in the urinary tract and appeared to be taken up by the L-cystine transport system. SIGNIFICANCE AND IMPACT OF THE STUDY: Although urinary tract infections caused by E. coli can be treated with sulfamethoxazole/trimethoprim and quinolones, resistance to these antibiotics has been increasing. These results suggest that L-selenaproline may represent a new class of compounds that could be used to treat these infections.     

P fimbriae on uropathogenic Escherichia coli O16:K1 and O18 strains

Abstract The fimbrial composition of 12 P-fimbriate uropathogenic Escherichia coli O16 and O18 strains was analysed by immunoprecipitation with 14 fimbria-specific antisera. All the O16 strains possessed a P fimbrial serovariant with an apparent M _r of 17500. One strain had an additional, serologically closely related P fimbria with an apparent M _r of 19 800. Two groups were found among the O18 strains; one possessing a type 1C fimbria and a 19800-Da P fimbria, the other lacking type 1C fimbriae and possessing a P-fimbrial variant with an apparent M _r of 17 800. Fimbriae on strains within the groups were serologically similar by immunoprecipitation assays. Also, the fimbriae on the O16 and O18 strains were mutually cross-reactive. The grouping of the O18 strains by fimbrial serology corresponded to the previous clonal grouping based on other phenotypic characters.     

Ferritinophagy drives uropathogenic Escherichia coli persistence in bladder epithelial cells

Autophagy is a cellular recycling pathway, which in many cases, protects host cells from infections by degrading pathogens. However, uropathogenic Escherichia coli (UPEC), the predominant cause of urinary tract infections (UTIs), persist within the urinary tract epithelium (urothelium) by forming reservoirs within autophagosomes. Iron is a critical nutrient for both host and pathogen, and regulation of iron availability is a key host defense against pathogens. Iron homeostasis depends on the shuttling of iron-bound ferritin to the lysosome for recycling, a process termed ferritinophagy (a form of selective autophagy). Here, we demonstrate for the first time that UPEC shuttles with ferritin-bound iron into the autophagosomal and lysosomal compartments within the urothelium. Iron overload in urothelial cells induces ferritinophagy in an NCOA4-dependent manner causing increased iron availability for UPEC, triggering bacterial overproliferation and host cell death. Addition of even moderate levels of iron is sufficient to increase and prolong bacterial burden. Furthermore, we show that lysosomal damage due to iron overload is the specific mechanism causing host cell death. Significantly, we demonstrate that host cell death and bacterial burden can be reversed by inhibition of autophagy or inhibition of iron-regulatory proteins, or chelation of iron. Together, our findings suggest that UPEC persist in host cells by taking advantage of ferritinophagy. Thus, modulation of iron levels in the bladder may provide a therapeutic avenue to controlling UPEC persistence, epithelial cell death, and recurrent UTIs.     

Isolation and characterisation of dog uropathogenic Escherichia coli strains and their fimbriae

A number of Escherichia coli strains have been isolated from dogs with urinary tract infections. These strains have been characterised with respect to their O, K, H, and fimbrial antigens, colicin production, antibiotic resistance, plasmid content and their ability to haemagglutinate erythrocytes from various species. Crossed immunoelectrophoresis of fimbrial extracts, as well as the reaction of partly purified fimbriae of a number of these strains with monoclonal antibodies revealed homology or a strong crossereaction with an F12 fimbrial subunit protein of human uropathogenic E. coli strains. Unlike human F12 fimbriae producing strains, the dog isolates did agglutinate dog erythrocytes in the presence of D-mannose but not human erythrocytes, indicating that the adhesin carried by these strains is different from the adhesin on fimbriae of human uropathogenic E. coli. Similar indications were obtained from experiments with latex beads coated with the receptor for P-fimbriae. These beads were agglutinated by Escherichia coli strains from human urinary tract infections, but not by the dog isolates described here. Preliminary adhesion experiments of human and dog Escherichia coli to human bladder epithelial and canine kidney epithelial cells also showed differences in adhesion depending on the origin of the strain tested.     

Escherichia coli outer membrane protease OmpT confers resistance to urinary cationic peptides

Escherichia coli OmpT, located in the outer membrane, has been characterized as a plasminogen activator, with the ability to hydrolyze protamine and block its entry. In this investigation, a complex of low molecular weight cationic peptides purified from human urine by a combination of membrane ultrafiltration and weak cation exchange chromatography was characterized. The impact of OmpT on E. coli resistance to urinary cationic peptides was investigated by testing ompT knockout strains. The ompT mutants were more susceptible to urinary cationic peptides than ompT⁺ strains, and this difference was abolished by complementation of the mutants with pUC19 carrying the ompT gene. The urinary protease inhibitor ulinastatin greatly decreased the resistance of the ompT⁺ strains. Overall, the data indicate that OmpT may help E. coli persist longer in the urinary tract by enabling it to resist the antimicrobial activity of urinary cationic peptides.