scholarly journals Characterization of the role of yadK in the pathogenesis of enterohaemorrhagic E. coli O157:H7

2021 ◽  
Author(s):  
Yijing Yu

Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 can use serious diarrhea and haemolytic uremic syndrome. Various factors including adhesins contribute to pathogenesis of EHEC. Previous studies suggested that yadK gene, which encodes a putative fimbrial adhesin in EHEC, may be involved in response of EHEC to acid stress. To characterize role of yadK protein in the pathogenesis of EHEC, recombinant yadK protein was generated and used to immunize rabbit to obtain anti-yadK antiserum, which was able to specifically recognize over-expressed yadK protein in EHEC. Western blotting with anti-yadK revealed a higher level of yadK expression in EHEC under acid adapted-acid stress compared to EHEC under unstressed conditions, which confirmed earlier yadK mRNA studies and indicated that yadK is upregulated in EHEC under acid stress. Finally, we observed that anti-yadK antiserum was able to specifically reduce adhesion of acid stressed EHEC to human epithelial cells compared to adhesion level of unstressed EHEC.

2021 ◽  
Author(s):  
Yijing Yu

Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 can use serious diarrhea and haemolytic uremic syndrome. Various factors including adhesins contribute to pathogenesis of EHEC. Previous studies suggested that yadK gene, which encodes a putative fimbrial adhesin in EHEC, may be involved in response of EHEC to acid stress. To characterize role of yadK protein in the pathogenesis of EHEC, recombinant yadK protein was generated and used to immunize rabbit to obtain anti-yadK antiserum, which was able to specifically recognize over-expressed yadK protein in EHEC. Western blotting with anti-yadK revealed a higher level of yadK expression in EHEC under acid adapted-acid stress compared to EHEC under unstressed conditions, which confirmed earlier yadK mRNA studies and indicated that yadK is upregulated in EHEC under acid stress. Finally, we observed that anti-yadK antiserum was able to specifically reduce adhesion of acid stressed EHEC to human epithelial cells compared to adhesion level of unstressed EHEC.


2010 ◽  
Vol 55 (No. 8) ◽  
pp. 359-368 ◽  
Author(s):  
M. Atef Yekta ◽  
F. Verdonck ◽  
W. Van Den Broeck ◽  
BM Goddeeris ◽  
E. Cox ◽  
...  

Enterohemorrhagic Escherichia coli (EHEC) serotype O157:H7 strains are associated with haemorraghic colitis and haemolytic uremic syndrome (HUS) in humans. Cattle are a reservoir of E. coli O157:H7. We studied the ability of bovine and human lactoferrin, two natural antimicrobial proteins present in milk, to inhibit E. coli O157:H7 growth and attachment to a human epithelial colorectal adenocarcinoma cell line (Caco-2). The direct antibacterial effect of bLF on E. coli O157:H7 was stronger than that of hLF. Nevertheless, both lactoferrins had bacteriostatic effects even at high concentrations (10 mg/ml), suggesting blocking of LF activity by a yet undefined bacterial defence mechanism. Additionally, both lactoferrins significantly inhibited E. coli O157:H7 attachment to Caco-2 cells. However, hLF was more effective than bLF, probably due to more efficient binding of bLF to intelectin present on human enterocytes leading to uptake and thus removal of bLF from the extracellular environment. Inhibition of bacterial attachment to Caco-2 cells was at least partly due to the catalytic effect of lactoferrins on the type III secreted proteins EspA and EspB


2012 ◽  
Vol 78 (19) ◽  
pp. 6799-6803 ◽  
Author(s):  
Sam Abraham ◽  
David M. Gordon ◽  
James Chin ◽  
Huub J. M. Brouwers ◽  
Peter Njuguna ◽  
...  

ABSTRACTThe role ofEscherichia colias a pathogen has been the focus of considerable study, while much less is known about it as a commensal and how it adapts to and colonizes different environmental niches within the mammalian gut. In this study, we characterizeEscherichia coliorganisms (n= 146) isolated from different regions of the intestinal tracts of eight pigs (dueodenum, ileum, colon, and feces). The isolates were typed using the method of random amplified polymorphic DNA (RAPD) and screened for the presence of bacteriocin genes and plasmid replicon types. Molecular analysis of variance using the RAPD data showed thatE. coliisolates are nonrandomly distributed among different gut regions, and that gut region accounted for 25% (P< 0.001) of the observed variation among strains. Bacteriocin screening revealed that a bacteriocin gene was detected in 45% of the isolates, with 43% carrying colicin genes and 3% carrying microcin genes. Of the bacteriocins observed (H47, E3, E1, E2, E7, Ia/Ib, and B/M), the frequency with which they were detected varied with respect to gut region for the colicins E2, E7, Ia/Ib, and B/M. The plasmid replicon typing gave rise to 25 profiles from the 13 Inc types detected. Inc F types were detected most frequently, followed by Inc HI1 and N types. Of the Inc types detected, 7 were nonrandomly distributed among isolates from the different regions of the gut. The results of this study indicate that not only may the different regions of the gastrointestinal tract harbor different strains ofE. colibut also that strains from different regions have different characteristics.


2020 ◽  
Vol 75 (9) ◽  
pp. 2554-2563 ◽  
Author(s):  
Christopher Fröhlich ◽  
Vidar Sørum ◽  
Sandra Huber ◽  
Ørjan Samuelsen ◽  
Fanny Berglund ◽  
...  

Abstract Background MBLs form a large and heterogeneous group of bacterial enzymes conferring resistance to β-lactam antibiotics, including carbapenems. A large environmental reservoir of MBLs has been identified, which can act as a source for transfer into human pathogens. Therefore, structural investigation of environmental and clinically rare MBLs can give new insights into structure–activity relationships to explore the role of catalytic and second shell residues, which are under selective pressure. Objectives To investigate the structure and activity of the environmental subclass B1 MBLs MYO-1, SHD-1 and ECV-1. Methods The respective genes of these MBLs were cloned into vectors and expressed in Escherichia coli. Purified enzymes were characterized with respect to their catalytic efficiency (kcat/Km). The enzymatic activities and MICs were determined for a panel of different β-lactams, including penicillins, cephalosporins and carbapenems. Thermostability was measured and structures were solved using X-ray crystallography (MYO-1 and ECV-1) or generated by homology modelling (SHD-1). Results Expression of the environmental MBLs in E. coli resulted in the characteristic MBL profile, not affecting aztreonam susceptibility and decreasing susceptibility to carbapenems, cephalosporins and penicillins. The purified enzymes showed variable catalytic activity in the order of &lt;5% to ∼70% compared with the clinically widespread NDM-1. The thermostability of ECV-1 and SHD-1 was up to 8°C higher than that of MYO-1 and NDM-1. Using solved structures and molecular modelling, we identified differences in their second shell composition, possibly responsible for their relatively low hydrolytic activity. Conclusions These results show the importance of environmental species acting as reservoirs for MBL-encoding genes.


2018 ◽  
Vol 102 ◽  
pp. 147
Author(s):  
Wouter Feitz ◽  
Carolina Ortiz ◽  
Dorothea Orth-Hoeller ◽  
Lambert van den Heuvel ◽  
Nicole van de Kar ◽  
...  

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 2804-2804
Author(s):  
Toshihiko Nishimura ◽  
John Morser ◽  
Zhifei Shao ◽  
Lawrence L. Leung

Abstract Hemolytic uremic syndrome (HUS) is characterized by a triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. The most common cause of HUS is Shiga toxin (STX)-producing E. coli, and eculizumab, a monoclonal antibody against complement C5, has shown clinical efficacy in some patients. Carboxypeptidase B2 (CPB2) is a metalloprotease activated by the thrombin/thrombomodulin complex that inactivates a number of inflammatory mediators, including complement C3a, and C5a by removing their C-terminal arginine. We hypothesized that in a murine model of STX-induced HUS, Cpb2-/- mice would have exacerbated disease compared to wild type (WT) mice due to excessive C3a and/or C5a in the absence of CPB2. A mouse model of STX-induced HUS was established by giving STX and LPS toxins intraperitoneally. Cpb2-/- mice had worse survival than WT (37% survival vs. 87% at 48h, p=0.0156). At 48h, severe thrombocytopenia developed in both WT and Cpb2-/- mice (WT: 0.096x106/μL; Cpb2-/-: 0.054x106/μL) compared to controls (1.2x106/μL; p>0.0001 vs. either WT or Cpb2-/-), with Cpb2-/- mice showing worse thrombocytopenia. Renal insufficiency was worse in Cpb2-/- mice than WT mice (BUN at 48h: 85 mg/dL vs. 37 mg/dL, p=0.0074; creatinine: 1.33 mg/dL vs. 0.23 mg/dL; p=0.0112, for Cpb2-/- and WT mice respectively, compared with normal baseline BUN and creatinine of 19 mg/dL and 0.1 mg/dL). Cpb2-/- mice developed worse anemia than WT (hemoglobin 9.8 g/dL vs. 12.4 g/dL, p=0.001 in Cpb2-/- vs. WT mice respectively). At 48h, liver function was worse in Cpb2-/- mice than WT mice, while plasma LDH was increased in Cpb2-/- mice more than WT mice. Using a standardized health score, the Cpb2-/- mice were worse than WT mice at all time points. Thus this model recapitulates STX-induced HUS with the Cpb2-/- mice having worse disease than WT. If the animals were treated with STX alone, there were no deaths in either genotype at 48h and only 37.5% mortality in Cpb2-/- mice by 60h compared with no deaths in WT mice. BUN, creatinine, liver enzymes and LDH were increased in both genotypes treated with STX alone compared to untreated mice, but there was no significant difference between the genotypes. Treatment with LPS alone caused thrombocytopenia in both WT and Cpb2-/- mice and LDH, BUN and creatinine levels were higher in Cpb2-/- mice than in WT mice, but there was no death at 48h and no drop in hemoglobin. Thus while either STX alone or LPS alone caused pathological conditions in the mice, the typical triad of HUS was only present when STX and LPS were given in combination. The Cpb2-/- mice had worse disease than WT mice consistent with our hypothesis on the role of CPB2 in inactivating C3a and/or C5a in STX-induced HUS. The potential efficacy of C3a and/or C5a blockade and anti-thrombotic agents will be tested in this model. Disclosures No relevant conflicts of interest to declare.


2021 ◽  
Vol 15 (11) ◽  
pp. 1755-1760
Author(s):  
Jorge Acosta-Dibarrat ◽  
Edgar Enriquez-Gómez ◽  
Martín Talavera-Rojas ◽  
Edgardo Soriano-Vargas ◽  
Armando Navarro ◽  
...  

Introduction: Commensal Escherichia coli is defined as bacteria without known virulence factors that could be playing a specific role in some diseases; however, they could be responsible to disseminate antimicrobial resistance genes to other microorganisms. This study aimed to characterize the commensal E. coli isolates obtained from slaughtered sheep in the central region of Mexico. Methodology: Isolates were classified as commensal E. coli when distinctive genes related to diarrheagenic pathotypes (stx1, stx2, eae, bfp, LT, stp, ipaH, and aggR) were discarded by PCR. Identification of serotype, phylogenetic group, and antimicrobial resistance was also performed. Results: A total of 41 isolates were characterized. The phylogenetic groups found were B1 in 37 isolates (90.2%), A in 2 (4.8%), and 1 isolate (2.4%) for C and D groups. Serotypes associated with diarrhea in humans (O104:H2 and O154:NM) and hemolytic uremic syndrome (O8:NM) were detected. Thirty-three isolates (80%) were resistant to ceftazidime, 23 (56%), to tetracycline 8 (19.5%) to ampicillin, and 1 to amikacin. Six isolates (14.6%) were multidrug-resistant. Conclusions: This study provides new information about commensal E. coli in slaughtered sheep, high percentages of resistance to antibiotics, and different profiles of antimicrobial resistance were found, their dissemination constitute a risk factor towards the consuming population.


2019 ◽  
Vol 7 (10) ◽  
pp. 452 ◽  
Author(s):  
Xu ◽  
Guo ◽  
Li ◽  
Zhang ◽  
Wu ◽  
...  

Cryptosporidium parvum is a protozoan parasite that can cause moderate-to-severe diarrhea. Insulinase-like proteases (INS) are one of the largest protein families within the small proteome of the pathogen. However, their roles in C. parvum biology remain un-elucidated. In this study, a member of the protein family, INS-15 of C. parvum encoded by cgd3_4260, was cloned, expressed and characterized to understand its function. INS-15 and its domain I were expressed in Escherichia coli and polyclonal antibodies against the domain I and one specific polypeptide were prepared in rabbits. The role of INS-15 protein in the C. parvum invasion was preliminarily studied. Recombinant INS-15 protein and its domain I were successfully expressed in E. coli, together with various degraded products. The cgd3_4260 gene had a peak expression at 2 h of in vitro C. parvum culture, while the INS-15 protein was expressed in the mid-anterior region of sporozoites and the area of merozoites opposite to the nucleus. Anti-INS-15 domain I antibodies reduced the invasion of C. parvum sporozoites by over 40%. The anterior location of INS-15 in invasion stages and partial reduction of in vitro growth indicate that INS-15 plays some roles in the invasion or early development of C. parvum.


1999 ◽  
Vol 62 (7) ◽  
pp. 741-746 ◽  
Author(s):  
FRANÇOIS CAYA ◽  
JOHN M. FAIRBROTHER ◽  
LOUISE LESSARD ◽  
SYLVAIN QUESSY

The purpose of this study was to evaluate the risk for human health associated with pathogenic Escherichia coli isolated from airsacculitis and cellulitis in chickens, by comparing the genotypic and phenotypic profiles of avian E. coli isolates and E. coli strains isolated from sick humans during the same period and in the same geographical area as the avian isolates. A total of 96 isolates and 46 isolates from lesions of cellulitis and airsacculitis, respectively, were obtained. Isolates from the backs of some of the affected and healthy birds and 91 intestinal and extraintestinal isolates from humans with diarrhea, urinary tract infections, or septicemia were examined. The frequency of antimicrobial resistance was in general higher in the avian than in the human isolates. VT1-VT2-Eae and VT2-Eae, pathotypes associated with hemolytic and uremic syndrome and bloody diarrhea in humans, were the most frequently encountered pathotypes in human intestinal isolates but were not recovered from the avian isolates. Aero-Pap-TSH and Aero-TSH were the most frequently encountered pathotypes in avian isolates but were rarely observed in human isolates. No avian isolate was of serogroup O157, whereas many human isolates belonged to this O group. O78 and O2 were the most frequently observed O groups in avian isolates but were rarely found in human isolates. Only two avian isolates demonstrated possible relatedness to human isolates based on pulsed-field gel electrophoresis profiles, but they belonged to different pathotypes. Our results suggest that avian isolates recovered from cellulitis and air sacullitis possess very few of the attributes required to cause diseases in humans. It is also concluded that isolates from cellulitis and airsacculitis do not represent a greater hazard than isolates from the back of healthy birds.


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