In Silico Prediction and Design of Uropathogenic Escherichia coli Alpha-Hemolysin Generate a Soluble and Hemolytic Recombinant Toxin

The secretion of α-hemolysin by uropathogenic Escherichia coli (UPEC) is commonly associated with the severity of urinary tract infections, which makes it a predictor of poor prognosis among patients. Accordingly, this toxin has become a target for diagnostic tests and therapeutic interventions. However, there are several obstacles associated with the process of α-hemolysin purification, therefore limiting its utilization in scientific investigations. In order to overcome the problems associated with α-hemolysin expression, after in silico prediction, a 20.48 kDa soluble α-hemolysin recombinant denoted rHlyA was constructed. This recombinant is composed by a 182 amino acid sequence localized in the aa542–723 region of the toxin molecule. The antigenic determinants of the rHlyA were estimated by bioinformatics analysis taking into consideration the tertiary form of the toxin, epitope analysis tools, and solubility inference. The results indicated that rHlyA has three antigenic domains localized in the aa555–565, aa600–610, and aa674–717 regions. Functional investigation of rHlyA demonstrated that it has hemolytic activity against sheep red cells, but no cytotoxic effect against epithelial bladder cells. In summary, the results obtained in this study indicate that rHlyA is a soluble recombinant protein that can be used as a tool in studies that aim to understand the mechanisms involved in the hemolytic and cytotoxic activities of α-hemolysin produced by UPEC. In addition, rHlyA can be applied to generate monoclonal and/or polyclonal antibodies that can be utilized in the development of diagnostic tests and therapeutic interventions.

The Yersinia High-Pathogenicity Island Encodes a Siderophore-Dependent Copper Response System in Uropathogenic Escherichia coli

Interactions between bacteria and transition metal ions play an important role in encounters between humans and bacteria. Siderophore systems have long been prominent mediators of these interactions.

Dictamnine Inhibits the Adhesion to and Invasion of Uropathogenic Escherichia Coli (UPEC) to Urothelial Cells

Uropathogenic Escherichia coli (UPEC) is the most common pathogenic bacteria associated with urinary tract infection (UTI). UPEC can cause UTI by adhering to and invading uroepithelial cells. Fimbriae is the most important virulence factor of UPEC, and a potentially promising target in developing novel antibacterial treatments. In this study, the antibacterial properties and effects of the compound dictamnine, extracted from the traditional Chinese medicine Cortex Dictamni, on the bacterial morphology, cell adhesion, and invasion of UPEC were studied. Dictamnine exhibited no obvious antibacterial activity against UPEC, but significantly impeded the ability of UPEC to adhere to and invade uroepithelial cells. RT-qPCR analysis showed that treatment downregulated the expression of type 1 fimbriae, P fimbriae, and curli fimbriae adhesion genes, and also downregulated adhesion-related receptor genes of uroepithelial cells. Transmission electron microscopy showed that dictamnine destroyed the structure of the fimbriae and the surface of the bacteria became smooth. These results suggest that dictamnine may help to prevent UTI by simultaneously targeting UPEC fimbriae and urothelial adhesin receptors, and may have a potential use as a new anti-UPEC drug.

In vitro growth of uropathogenic Escherichia coli isolated from a snow leopard treated with homeopathic and isopathic remedies: a pilot study

This paper reports the results of incubation of a strain of uropathogenic Escherichia coli (UPEC) isolated from a snow leopard - which had died of septicemia secondary to necro-hemorrhagic cystitis - with homeopathic and isopathic remedies. Methods: UPEC was isolated from heart blood and previously typified for virulence factors; it was incubated with homeopathic remedies Cantharis vesicatoria (urinary tract infection affinity), Mercurius solubilis (from symptoms analysis) and nosode prepared from the actual strain, all in dilution 12cH. Results: 2 patterns of bacterial growth were observed, associated to the quality of nutrients in the culture medium; in rich-nutrient medium, nosode of E. coli 12cH had a significant inhibitory effect; in poor-nutrient medium, Merc 12cH exerted significant inhibitory effect. Conclusion: results suggest that the previous conditions of prokaryote systems may influence the in vitro response to homeopathic and isopathic remedies. Keywords: urinary tract infection; Felines; Uropathogenic Escherichia coli; Homeopathy; Isopathy.  Estudo do crescimento (in vitro) de Escherichia coli uropatogénica isolada a partir de um leopardo das neves e tratada com medicamentos homeopáticos e isopáticos: estudo piloto. ResumoEste artigo relata os resultados da incubação de uma linhagem de Escherichia coli uropatogénica (UPEC) isolada a partir de um leopardo das neves, que morreu de septicemia secundária a cistite necrótica-hemorrágica. A UPEC foi tratada com preparados homeopáticos e isopáticos. Métodos: UPEC foi isolada de sangue cardíaco e previamente tipificada para fatores de viruléncia; foi incubada com o medicamento homeopático Cantharis vesicatoria (afinidade com infecção do trato urinário), Mercurius solubilis (a partir da análise de sintomas) e nosódio preparado a partir da mesma linhagem de bactérias, todas em 12 cH. Resultados: 2 padrões de crescimento bacteriano foram observados, associados àqualidade dos nutrientes do meio de cultura; em meios ricos em nutrientes, nosódio de E. coli 12 cH teve um significativo efeito inibitório; em meio pobre de nutrientes, Merc 12 cH exerceu efeito inibitório significativo. Conclusão: os resultados sugerem que as condições prévias do sistema procarioto estudado podem influenciar as respostas proliferativas in vitro para preparados homeopáticos e isopáticos. Palavras-chave: infecção do trato urinário, felinos, Escherichia coli uropatogénica; homeopatia, isopatia.  Crecimiento in vitro de Escherichia coli uropatogenica, aisladas de un leopardo de la nieve, tratadas con remedios homeopáticos y isopáticos: un estudio preliminar ResumenEste trabajo presenta los resultados de la incubación de una cepa de Escherichia coli uropatogenica (UPEC) aislada de un leopardo de la nieve, que había muerto de septicemia secundaria a la necro-hemorrágica cistitis, con remedios homeopáticos y isopáticos. Métodos: Se aisló UPEC de la sangre del corazón y caracterizado por factores de virulencia, y se incubó con los remedios homeopáticos Cantharis vesicatoria (afinidad con la infección del tracto urinario), Mercurius solubilis (a partir de la análisis de los síntomas) y nosódio preparado a partir de la cepa, todos em la dilución 12cH. Resultados: 2 patrones de crecimiento de las bacterias se han observado, asociado a la calidad de los nutrientes en el medio de cultivo. En medio rico en nutrientes, nosódios de E. coli 12cH tuvo un importante efecto inhibitorio; en médios pobres en nutrientes, Merc 12cH haz ejercido importante efecto inhibitorio. Conclusión: los resultados sugieren que las condiciones anteriores de los sistemas procariotos pueden influir en la respuesta in vitro a los remedios homeopáticos y isopáticos. Palabras-clave: Infección del tracto urinario; Felinos; Escherichia coli Uropatogenica ; Homeopatía; Isopatía  Correspondence author: Leoni Villano Bonamin, [email protected] How to cite this article: Kawakami AP; Osugui L; César AT; Priven SW; Carvalho VM; Bonamin LV. In vitro growth of uropathogenic Escherichia coli isolated from a snow leopard treated with homeopathic and isopathic remedies: a pilot study. Int J High Dilution Res [online]. 2009 [cited YYYY Month dd]; 8 (27): 41-44. Available from: http://journal.giri-society.org/index.php/ijhdr/article/view/341/394. ÂÂÂ

Antibacterial and antibiofilm effect of silver and gold nanoparticles in Uropathogenic Escherichia coli

Nosocomial infections of the urinary tract are the most common nosocomial infections. Infectious diseases currently pose a life threatening due to the increasing phenomenon of infectious agent resistance. One of the factors known to cause resistance to broad-spectrum antimicrobials agent is the susceptibility to biofilm formation. The use of silver nanoparticles (NPs) is now regarded as one of the most promising strategies to target infections associated with biofilms. This study aims to determine the application of nanotechnology in the form of metal nanoparticles in targeting the formation of biofilms on infectious agents. The antibacterial and antibiofilm activity test was performed on a 96-well microtiter plate on uropathogenic Escherichia coli (UPEC) at concentrations of 0, 10, 20, 30, 40, 50, 60, 70, 80, 90, and 100 ppm. The results showed the addition of silver and gold nanoparticles at different concentrations affected and inhibited the growth of UPEC and prevented the formation of biofilms, as indicated by a decrease in the OD value. In conclusion, silver and gold nanoparticles have antibacterial and antibiofilm effects on UPEC.

Human Renal Fibroblasts, but Not Renal Epithelial Cells, Induce IL-1β Release during a Uropathogenic Escherichia coli Infection In Vitro

Understanding how uropathogenic Escherichia coli (UPEC) modulates the immune response in the kidney is essential to prevent UPEC from reaching the bloodstream and causing urosepsis. The purpose of this study was to elucidate if renal fibroblasts can release IL-1β during a UPEC infection and to investigate the mechanism behind the IL-1β release. We found that the UPEC strain CFT073 induced an increased IL-1β and LDH release from renal fibroblasts, but not from renal epithelial cells. The UPEC-induced IL-1β release was found to be NLRP3, caspase-1, caspase-4, ERK 1/2, cathepsin B and serine protease dependent in renal fibroblasts. We also found that the UPEC virulence factor α-hemolysin was necessary for IL-1β release. Conditioned medium from caspase-1, caspase-4 and NLRP3-deficient renal fibroblasts mediated an increased reactive oxygen species production from neutrophils, but reduced UPEC phagocytosis. Taken together, our study demonstrates that renal fibroblasts, but not renal epithelial cells, release IL-1β during a UPEC infection. This suggest that renal fibroblasts are vital immunoreactive cells and not only structural cells that produce and regulate the extracellular matrix.