Subtilase Cytotoxin from Shiga-Toxigenic Escherichia Coli Impairs the Inflammasome and Exacerbates Enteropathogenic Bacterial Infection

In this study, the cytotoxicity of the recently described subtilase variant SubAB2-2of Shiga toxin-producingEscherichia coliwas determined and compared to the plasmid-encoded SubAB1and the chromosome-encoded SubAB2-1variant. The genes for the respective enzymatic active (A) subunits and binding (B) subunits of the subtilase toxins were amplified and cloned. The recombinant toxin subunits were expressed and purified. Their cytotoxicity on Vero cells was measured for the single A and B subunits, as well as for mixtures of both, to analyze whether hybrids with toxic activity can be identified. The results demonstrated that all three SubAB variants are toxic for Vero cells. However, the values for the 50% cytotoxic dose (CD50) differ for the individual variants. Highest cytotoxicity was shown for SubAB1. Moreover, hybrids of subunits from different subtilase toxins can be obtained which cause substantial cytotoxicity to Vero cells after mixing the A and B subunits prior to application to the cells, which is characteristic for binary toxins. Furthermore, higher concentrations of the enzymatic subunit SubA1exhibited cytotoxic effects in the absence of the respective B1subunit. A more detailed investigation in the human HeLa cell line revealed that SubA1alone induced apoptosis, while the B1subunit alone did not induce cell death.

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Isolation and Identification of Kidney-derived Pathogenic Escherichia coli from Red Panda (Ailurus fulgens)

10.21203/rs.2.20381/v1 ◽

2020 ◽

Author(s):

Songrui Liu ◽

Yunli Li ◽

Chanjuan Yue ◽

Dongsheng Zhang ◽

Xiaoyan Su ◽

...

Keyword(s):

Escherichia Coli ◽

Bacterial Infection ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Ex Situ ◽

Type I ◽

Pathogenicity Test ◽

Whole Genome ◽

Red Panda ◽

Ailurus Fulgens

Abstract Background Disease prevention and control is a significant part during the ex-situ conservation of the red panda (Ailurus fulgens) with bacterial infection being one of the important threats to the health of the captive population. So far, there was no systematic and detailed publications about the red panda-related E. coli disease. This study was conducted for the purpose of determining the cause of death, etiology and pathogenesis on a red panda through clinical symptoms, complete blood count, biochemical analysis, pathological diagnosis, antimicrobial susceptibility test, mouse pathogenicity test, and bacterial whole genome sequencing.Results A bacterial strain confirmed as Uropathogenic Escherichia coli (UPEC) was isolated from one captive dead red panda, which is resistant to most of the β-lactam drugs and a small number of aminoglycoside medications. The mouse pathogenicity test results showed the strains isolated postmortem from mice were the same as from the dead red panda, and the pathological findings were similar to the red panda while they were not completely the same. These pathological differences between red panda and mice may be related to the routes of infection and perhaps species differences and tolerance. The whole genome sequencing results showed that the isolated strain contained P pili, type I pili and iron uptake system related factors, which were closely related to its nephrotoxicity. Conclusion The red panda died of bacterial infection which was identified as Uropathogenic Escherichia coli. The pathogenic mechanisms of the strain are closely related to the expression of specific virulence genes.

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Subtilase Cytotoxin-Coding Genes in Verotoxin-Producing Escherichia coli Strains from Sheep and Goats Differ from Those from Cattle

Applied and Environmental Microbiology ◽

10.1128/aem.05604-11 ◽

2011 ◽

Vol 77 (23) ◽

pp. 8259-8264 ◽

Cited By ~ 11

Author(s):

José A. Orden ◽

Pilar Horcajo ◽

Ricardo de la Fuente ◽

José A. Ruiz-Santa-Quiteria ◽

Gustavo Domínguez-Bernal ◽

...

Keyword(s):

Escherichia Coli ◽

Human Disease ◽

Virulence Genes ◽

Small Ruminants ◽

Human Beings ◽

Content Type ◽

Sheep And Goats ◽

E Coli ◽

Healthy Cattle ◽

Subtilase Cytotoxin

ABSTRACTSubtilase cytotoxin (SubAB) from verotoxin (VT)-producingEscherichia coli(VTEC) strains was first described in the 98NK2 strain and has been associated with human disease. However, SubAB has recently been found in two VT-negativeE. colistrains (ED 591 and ED 32). SubAB is encoded by two closely linked, cotranscribed genes (subAandsubB). In this study, we investigated the presence ofsubABgenes in 52 VTEC strains isolated from cattle and 209 strains from small ruminants, using PCR. Most (91.9%) VTEC strains from sheep and goats and 25% of the strains from healthy cattle possessedsubABgenes. The presence ofsubABin a high percentage of the VTEC strains from small ruminants might increase the pathogenicity of these strains for human beings. Some differences in the results of PCRs and in the association with some virulence genes suggested the existence of different variants ofsubAB. We therefore sequenced thesubAgene in 12 strains and showed that thesubAgene in most of thesubAB-positive VTEC strains from cattle was almost identical (about 99%) to that in the 98NK2 strain, while thesubAgene in most of thesubAB-positive VTEC strains from small ruminants was almost identical to that in the ED 591 strain. We propose the termssubAB1to describe the SubAB-coding genes resembling that in the 98NK2 strain andsubAB2to describe those resembling that in the ED 591 strain.

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BiP (Grp78): A Target for Escherichia coli Subtilase Cytotoxin

Heat Shock Proteins - Moonlighting Cell Stress Proteins in Microbial Infections ◽

10.1007/978-94-007-6787-4_20 ◽

2013 ◽

pp. 309-322

Author(s):

Adrienne W. Paton ◽

Hui Wang ◽

James C. Paton

Keyword(s):

Escherichia Coli ◽

Subtilase Cytotoxin

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Transcription of the Subtilase Cytotoxin Gene subAB1 in Shiga Toxin-Producing Escherichia coli Is Dependent on hfq and hns

Applied and Environmental Microbiology ◽

10.1128/aem.01281-19 ◽

2019 ◽

Vol 85 (20) ◽

Cited By ~ 2

Author(s):

Laura Heinisch ◽

Katharina Zoric ◽

Maike Krause ◽

Herbert Schmidt

Keyword(s):

Gene Expression ◽

Escherichia Coli ◽

Shiga Toxin ◽

Promoter Activity ◽

Deletion Mutants ◽

Content Type ◽

E Coli ◽

Intensive Investigation ◽

Subtilase Cytotoxin

ABSTRACT Certain foodborne Shiga toxin-producing Escherichia coli (STEC) strains carry genes encoding the subtilase cytotoxin (SubAB). Although the mode of action of SubAB is under intensive investigation, information about the regulation of subAB gene expression is currently not available. In this study, we investigated the regulation of the chromosomal subAB1 gene in laboratory E. coli strain DH5α and STEC O113:H21 strain TS18/08 using a luciferase reporter gene assay. Special emphasis was given to the role of the global regulatory protein genes hfq and hns in subAB1 promoter activity. Subsequently, quantitative real-time PCR was performed to analyze the expression of Shiga toxin 2a (Stx2a), SubAB1, and cytolethal distending toxin V (Cdt-V) genes in STEC strain TS18/08 and its isogenic hfq and hns deletion mutants. The deletion of hfq led to a significant increase of up to 2-fold in subAB1 expression, especially in the late growth phase, in both strains. However, deletion of hns showed different effects on the promoter activity during the early and late exponential growth phases in both strains. Furthermore, upregulation of stx2a and cdt-V was demonstrated in hfq and hns deletion mutants in TS18/08. These data showed that the expression of subAB1, stx2a, and cdt-V is integrated in the regulatory network of global regulators Hfq and H-NS in Escherichia coli. IMPORTANCE Shiga toxin-producing Escherichia coli (STEC) strains are responsible for outbreaks of foodborne diseases, such as hemorrhagic colitis and the hemolytic uremic syndrome. The pathogenicity of those strains can be attributed to, among other factors, the production of toxins. Recently, the subtilase cytotoxin was detected in locus of enterocyte effacement (LEE)-negative STEC, and it was confirmed that it contributes to the cytotoxicity of those STEC strains. Although the mode of action of SubAB1 is under intensive investigation, the regulation of gene expression is currently not known. The global regulatory proteins H-NS and Hfq have impact on many cellular processes and have been described to regulate virulence factors as well. Here, we investigate the role of hns and hfq in expression of subAB1 as well as stx2a and cdt-V in an E. coli laboratory strain as well as in wild-type STEC strain TS18/08.

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Draft Genome Sequences of Five Shiga Toxin-Producing Escherichia coli Isolates Harboring the New and Recently Described Subtilase Cytotoxin Allelic Variant subAB2-3

Genome Announcements ◽

10.1128/genomea.01582-16 ◽

2017 ◽

Vol 5 (8) ◽

Cited By ~ 4

Author(s):

Taurai Tasara ◽

Lisa Fierz ◽

Jochen Klumpp ◽

Herbert Schmidt ◽

Roger Stephan

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Sequence Data ◽

Draft Genome ◽

Allelic Variant ◽

Genome Sequences ◽

Content Type ◽

Subtilase Cytotoxin

ABSTRACT We present here the draft genome sequences of five Shiga toxin-producing Escherichia coli (STEC) strains which tested positive in a primary subAB screening. Assembly and annotation of the draft genomes revealed that all strains harbored the recently described allelic variant subAB 2-3 . Based on the sequence data, primers were designed to identify and differentiate this variant.

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Bacteria meets influenza A virus: A bioluminescence mouse model of Escherichia coli O157:H7 following influenza A virus/Puerto Rico/8/34 (H1N1) strain infection

Journal of International Medical Research ◽

10.1177/0300060518778415 ◽

2018 ◽

Vol 46 (7) ◽

pp. 2875-2882

Author(s):

Zhongyi Wang ◽

Hang Chi ◽

Xiwen Wang ◽

Wenliang Li ◽

Zhiping Li ◽

...

Keyword(s):

Escherichia Coli ◽

Puerto Rico ◽

Bacterial Infection ◽

Influenza A Virus ◽

Influenza A ◽

Bacterial Colonization ◽

Infection Model ◽

Escherichia Coli O157 ◽

E Coli ◽

H1n1 Strain

Objective To develop a bioluminescence-labelled bacterial infection model to monitor the colonization and clearance process of Escherichia coli O157:H7 in the lungs of mice following influenza A virus/Puerto Rico/8/34 (H1N1) strain (IAV/PR8) infection. Methods BALB/c mice were administered IAV/PR8 or 0.01 M phosphate-buffered saline (PBS; pH 7.4) intranasally 4 days prior to intranasal administration of 1 × 107 colony-forming units (CFU) of E. coli O157:H7-lux. Whole-body bioluminescent signals were monitored at 10 min, 4 h, 8 h, 12 h, 16 h and 24 h post-bacterial infection. Lung bioluminescent signals and bacterial load (CFU/g) were monitored at 4 h, 8 h, 12 h, 16 h and 24 h post-bacterial infection. Results Prior IAV/PR8 infection of mice resulted in a higher level of bacterial colonization and a lower rate of bacterial clearance from the lungs compared with mice treated with PBS. There were also consistent findings between the bioluminescence imaging and the CFU measurements in terms of identifying bacterial colonization and monitoring the clearance dynamics of E. coli O157:H7-lux in mouse lungs. Conclusion This novel bioluminescence-labelled bacterial infection model rapidly detected bacterial colonization of the lungs and monitored the clearance dynamics of E. coli O157:H7-lux following IAV/PR8 infection.

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Inhibition of Water Absorption and Selective Damage to Human Colonic Mucosa Are Induced by Subtilase Cytotoxin Produced by Escherichia coli O113:H21

Infection and Immunity ◽

10.1128/iai.00287-13 ◽

2013 ◽

Vol 81 (8) ◽

pp. 2931-2937 ◽

Cited By ~ 14

Author(s):

Elizabeth Gerhardt ◽

Mariana Masso ◽

Adrienne W. Paton ◽

James C. Paton ◽

Elsa Zotta ◽

...

Keyword(s):

Escherichia Coli ◽

Water Absorption ◽

Shiga Toxin ◽

Human Colon ◽

Surface Epithelium ◽

Inflammatory Infiltration ◽

Content Type ◽

Uremic Syndrome ◽

Normal Water ◽

Subtilase Cytotoxin

ABSTRACTShiga toxin-producingEscherichia coliO157:H7 (STEC) is by far the most prevalent serotype associated with hemolytic uremic syndrome (HUS) although many non-O157 STEC strains have been also isolated from patients with HUS. The main virulence factor of STEC is the Shiga toxin type 2 (Stx2) present in O157 and non-O157 strains. Recently, another toxin, named subtilase cytotoxin (SubAB), has been isolated from several non-O157 strains and may contribute to the pathogenesis of HUS. Here, we have demonstrated that an O113:H21 STEC strain expressing SubAB and Stx2 inhibits normal water absorption across human colon and causes damage to the surface epithelium, necrosis, mononuclear inflammatory infiltration, edema, and marked mucin depletion. This damage was less marked, but nevertheless significant, when purified SubAB orE. coliO113:H21 expressing only SubAB was assayed. This is the first study showing that SubAB may directly participate in the mechanisms of diarrhea in children infected with non-O157 STEC strains.

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